Slide 1

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 2

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 3

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 4

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 5

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 6

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 7

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 8

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 9

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 10

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 11

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 12

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 13

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 14

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 15

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 16

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 17

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 18

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 19

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 20

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 21

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 22

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 23

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 24

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 25

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 26

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 27

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 28

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 29

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 30

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 31

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 32

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 33

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 34

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 35

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 36

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 37

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 38

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 39

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 40

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 41

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 42

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 43

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
43
and are directed against either viruses or bacterial toxins.

Thank
ou

44


Slide 44

Advanced Pharmacology-I
(PHR5001)
Lecture 12:
Anti-ulcer Agents
(H2 receptor antagonists)
Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
1

Introduction
Definition: Anti-ulcer (Antisecretory) agents are the drugs
which decreases the secretion of gastric acid in the stomach.

 Histamine - Powerful stimulant of Hydrochloric acid
secretion from parietal (oxyntic)cells of gastric mucosa.
 Histamine (In larger dose )- increase secretion of pepsin.
These actions are mediated by H2-receptor.
 The secretion of acid by these parietal cells are regulated by
various mediators or receptors.
a) Histamine H2 receptors
b) G receptors-release gastrin & release gastric acid
c) Acetylcholine (Ach) M3 receptor
2

What is Peptic Ulcer ?
• A peptic ulcer disease or PUD is an ulcer (defined as mucosal
erosions ≥ 0.5 cm) of an area of the gastrointestinal tract
exposed to the acid and pepsin secretion
• Gastritis is the precursor to PUD and it is clinically difficult to
differentiate the two
– Stomach (called gastric ulcer): Relieved by food but pain may
persist even after eating. Infrequent or absent remissions (>55
yrs)
– Duodenum (called duodenal ulcer): burning upper abdomen pain
relieved by food but reappears 1-3 hrs after meals. Worse pain
when stomach empty
– Esophagus (called Esophageal ulcer)
Causes of ulcers:
 Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.
Pylori.
 Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs?
• High [H+] in the gastric lumen
• Require defense mechanisms to protect oesophagus and stomach
– Mucus secretion: slows ion diffusion
– Prostaglandins: I2 and E2 (alcohol, aspirin )
– Bicabonate ions
– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors
& Defensive factors:

Gastroesophageal Reflux Disease (GERD)
•
•
•
•

Common and GI motility disorder
Acidity of Gastric contents – most common factor
Acid contents reflux back into esophagus
Intense burning, sometimes belching

• Can lead to esophagitis, and esophageal ulcers
– Barrett’s esophagus (A complication of severe chronic GERD
involving changes in the cells of the tissue that line the bottom
of the esophagus. These esophageal cells become irritated
when the contents of the stomach back up (refluxes) and
there is a small but definite increased risk of adenocarcinoma
of the esophagus.)

• Commonly associated with obesity
• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell.
ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1,
M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+
dependent pathway. Both pathway activates the H+/K+ ATPase. Ach
release from postganglionic vagal fibres can stimulate directly gastric
acid secretion though M3 (a specific muscarnic cholinergic receptor
subtype). Ach. also indirectly affects the parietal cells though the
stimulation of histamine release from the entrochromaffin-like cells
in the fundus and the stimulation of gastric release from G cells in the
gastric antrum.
• Histamine is released from ECL cells through the multifactorial
pathway and is critical regulator of acid production through the H2
sub type of receptor.
• The release of gastric is regulated through the multifactorial pathway
and it stimulates acid secretion predominantly in an indirect manner
by causing the release of histamine from ECL cells; a direct effect of
gastric or parietal cell is also seen but is less important.

Mechanism

8

Comparison
H1 receptor antagonist
 Two aryl, heteroaryl rings in
place of imidazole (bulky
groups)
 Connecting chain of aryl
and terminal nitrogen is of 2
to 3 atoms
 Ionic flexible chain at end.
Ionized at pH 7.4
 Hydrophobic (due to aryl
rings). High partition
coefficient value
 Low dipole moment

H2 receptor antagonist
 Imidazole or other 5
membered heterocyclic ring
(Imidazole is not necessary)
 Connecting chain of ring and
terminal nitrogen is of 4
atoms
 Polar π e containing systems,
amphoteric, unionized at pH
7.4
 Hydrophobic (due to polar
grp). Low partition coefficient
 High dipole moment
9

Treatment of ulcer
 H2 receptor antagonists are the most popular
drug for the treatment of peptic ulcer. H2 receptor
antagonists Must bind but not activate H2 receptor
site
 They bring about sympathomimetic relief and
promote ulcer healing.
 Gastric acid secretion involve H2 receptor which is
competitively blocked by H2 blockers.
 In 1972, Black and co-workers first described selective
H2 receptor blokade for acid secretion. With
successful introduction of cimetidine, in 1977
other analogs like ranitidine, famotidine &
roxatidine has been synthesized.
10

Historical development

11

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to
optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.
Substitution in side chain with different basic as well as neutral groups were
carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for
better activity.
Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine
12
(Famotidine) groups on ring showed better activity.

H2 receptor antagonists

13

Classification of Anti-ulcer Drugs
I.

Gastric Antisecretory Drugs
HCl secretion
1.
2.

Antagonize Rs. on Parietal Cell--- H2,M3, G
Inhibitor of H+-Pump

II. Antacids--- Neutralizations of secreted acid
and pepsin activity
III. Agents kill HP : Eradication of Helicobacter

pylori by triple therapy: Almost all duodenal and
2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd
Clarithromycin
Amoxycillin / Metronidazole

- 500 mg bd
- 1gm / 500 mg bd

Given for 14 days

Probanthine
Pirenzepine

PGE2
ACh

+

Histamine

+
_

M3

Ranitidine
Gastrin
_
Proglumide
_ Compete

Misoprostol

HCl & pepsin

PGE
receptor

Ca++

Adenyl
cyclase

ATP

+

Gastrin-R

H2

+

cAMP

+

+

Gastrin
receptor

Ca++
+

Protein Kinase
(Activated)
K+ + H+
K

_
Omeprazole

Proton pump

Gastric acid

Parietal cell
Lumen of stomach

_

Antacid

Classification of Anti-ulcer Drugs
1. Reduction in Gastric acid secretion:
•

H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine
and Roxatidine
Proton pump inhibitors: Omeprazole, Lansoprazole Pantoprazole,
Rabeprazole and Esomeprazole
Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium
Prostaglandin analogue: Misoprostol

•
•
•

2. Acid Neutralizing agents: (ANTACIDS)
•
•

3.
4.

Systemic: Sodium Bicarbonate and Sod. Citrate
Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium
hydroxide gel, Magaldrate and calcium carbonate

Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate
Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,
metronidazole, tinidazole and tetracycline

H2 Antagonists
• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine

• MOA:
–
–
–
–

Reversible competitive inhibitors of H2 receptor
Highly selective, no action on H1 or H3 receptors
All phases of gastric acid secretion
Very effective in inhibiting nocturnal acid secretion (as it
depends largely on Histamine )
– Modest impact on meal stimulated acid secretion (as it depends
on gastrin, acetylcholine and histamine)
– Volume of pepsin content and IF are also reduced
– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:
Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors
• Most effective drugs in antiulcer therapy
• Prodrugs requiring activation in acidic pH.
Omeprazole
• Block enzymes responsible for secreting HCl - binds
irreversibly to H+K+ATPase
• Prototype: Omeprazole
• Substituted Benzimidazole derivative
• Diffuses into G. canaliculi = accumulation
pH < 5 (proton catalyzed )=
tetracyclic sulfenamide + sulphenic acid
• Covalent binding with sulfhydryl cysteines of
H⁺K⁺ ATPase
• Irreversible inactivation of the pump molecule
• Acid suppressants regardless of stimulating factors
• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:
1. H2 –histamine receptor antagonist eg.
Ranitidine,Cimetidine,Famotidine, Nizatidine
- Peptic ulcer (Gastric ulcer and duodenal ulcer)
- Reflex oesophagitis
2. Proton Pump Inhibitor : Omeprazole Lansoprazole,
Esomeprazole,Rabeprazole,Pantoprazole
- Peptic ulcer
- Reflex oesophagitis
- As one component Of therapy for H-pylori infection
- Zollinger-Ellison syndrome
3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates
- Dyspepsia
- Symptomatic relief of peptic ulcer
- Esophageal reflex
4. Bismuth chelate
-One component of therapy for H-pylori infection.

Screening methods:1. Pyloric ligation induced gastric ulceration
2. Ethanol induced mucosal damage in rats
3. Stress ulcer through immobilization stress
Lower & upper extremities fixed, wrapped in a wire
gaze, kept at dark for 24 hr.
4. Indomethacin induced ulcers in rats
After 10 min of admistering 20 mg/kg
indomethacin, test drug is given orally. 6hr later ,
sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration
• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,
animals are anaesthetized with the help of anesthetic ether; the
abdomen is opened by a small midline incision. Pyloric portion of
the stomach is slightly lifted out and ligated.
• The stomach is replaced carefully and the abdominal wall is
closed by interrupted sutures. Rats are sacrificed by an over dose
of anaesthetic ether after four hours of pyloric ligation.
• Abdomen is opened and ligature is placed around esophagus.
Stomach is removed and contents are drained to centrifuge tube.
• The volume of the gastric juice is measured and centrifuged at
2000 rpm for 10 min. From the supernatant, aliquots (1 ml of
each) are taken for the determination of pH, total and free
acidity. Each stomach is examined (by a 10Χ magnifier lens) for
lesions in the fore stomach portion & indexed according to
severity.
0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation
% Inhibition of Ulceration =

Ethanol induced ulcer model
The ulcer is induced by administering ethanol to Albino rats
(fasted for 36 h).
One group represented the control group, which received
ethanol Second & Third
• Groups received methanolic extract of A. Indicum 250 and
• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were
• administered orally for Four group as reference standard
• drug. The gastric ulcers were induced in rats by
• administrating absolute ethanol (90%) (0.5 ml/100g) orally,
• after 45 min of methanolic extract and ranitidine treatment.
• They were kept in specially constructed cages to prevent
• coprophagia during and after the experiment. The animals
• were anaesthetized 1h latter with anaesthetic ether and
• stomach was incised along the greater curvature and
• ulceration will be scored

Advanced Pharmacology-I
(PHR5001)
Lecture 13:

Dr. M G Azam
Asstt. Professor
Dept. of Pharmacy, NSU
23



Antibody is a class of immunological proteins (globular
glycoproteins/ “Immunoglobulin” produced by plasma cells (B
lymphocytes) in response to a specific antigen.

• Antibodies function as markers,
binding to the antigen so that the
antigen molecules can be
recognized and destroyed by
phagocytes.
• The part of the antigen that the
antibody binds to is called the
epitope. The epitope is thus a short
amino acid sequence that the
antibody is able to recognize.
• Antibodies are useful tools for the
immune system to combat
pathogens.
24

 MAbs are identical. They are directed toward same epitope. So more
efficient and specific.

* Used in

Biomedical research
Diagnosis of diseases
Treatment of diseases such as infection, cancer, etc.

Polyclonal Abs

• Mixture of antibodies similar to sera
 Antibodies that are obtained from different B cell
resources. They are a combination of immunoglobulin
molecules secreted against a specific antigen, each
25
identifying a different epitope.

Structure of antibody
Consist of four polypeptide chains, somewhat resembling to a Y shape.
i) 2 identical Light chains ( L chains)

ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino
acids; globular domain
used in many proteins.
Variable domains,
Constant domains, Hinge.
Fab: fragment antigen
binding
Fc: fragment crystallizable
(effector functions)
CDR: Complementarity-Determining Regions
26

Immunoglobulin

(Ig) Class

Heavy
Chain

IgA

α

IgD

IgE

IgG

δ

ε

γ

Diagram

or

Distribution

External Secretions

B Cell surface receptor

Cells that secrete
histamines

Main antibody in
serum
Most Stable

IgM

μ

First antibody
secreted in
development
27

Production of Monoclonal Antibodies
• A mouse is immunized with the specific antigen“X”. This will allow
the mouse to produce antibodies.
• The spleen cells producing specific antibodies are removed.
• Fusion of spleen cells with tumor cells (myeloma cells) in a culture
medium is done. The resulting cell is called a hybridoma.
• Hybridoma cells are continuously growing cell line. Each hybridoma
will produce relatively large quantities of monoclonal antibody
molecules. These hybridomas have antibody producing capability from
lymphocytes & have ability to grow continuously (immortal) from malignant
cells.

• Separate the hybridomas from unfused cells.
• Culture selected hybridoma cells for the production of monoclonal
antibodies in large quantity.
• Remain indefinitely and maintained by in vitro; that is, in culture
vessels or in vivo-in the body of an animal so that antibodies will
28
be produced in body and can be recovered later from body fluid.

29

 Purification of monoclonal antibody :
 Affinity chromatography is used for
purification of mAbs.
 This method separates molecules on
their differing affinities for a ligand.
Antigen can be bound to the support
matrix (small, chemically reactive beads) in
order to purify antigen-specific antibody
from a polyclonal antiserum.
Specific antibody is eluted by altering
the pH, which disrupt antibody-antigen
bonds.
30

 Application of monoclonal antibody :
1. Research tools:
 Identification of cell surface marker
 Purification of proteins and enzymes
 Detection assays
 Genetic engineering
 Structure of cell membrane

Sufix of Antibody

2. Clinical diagnosis:
 Cancers
 Leukemia
 Viral disease
 Allergic reaction
 AIDS

3. Treatment of diseases:

31

• Diagnostic tests
Monoclonal antibodies are widely used as diagnostic and
research reagents.
• Once monoclonal antibodies for a given substance have been
produced, they can be used to detect the presence of this substance.
The Western blot test detect the protein on a membrane.
• Monoclonal antibodies can also be used to purify a substance with
techniques called affinity chromatography.
• Other monoclonal antibodies allow rapid diagnosis of hepatitis,
influenza, herpes and streptococcal infections.
• Antibodies are used in several diagnostic tests to detect small
amounts of drugs, toxins or hormones.
• Monoclonal Antibodies also used in ELISA (enzyme linked
immunosorbant Assay) for detection of viruses.
32

Monoclonal antibody in cancer therapy:
• By means of genetic engineering, so-called chimeric
antibodies are prepared that contain both human
and mouse sections.
• The mouse section is located in the variable region
of the antibody that contains the antigen- binding
site, while the human section is in the constant
region.
• After the variable region binds to the cancer antigen,
the human section activates complement and
cytotoxic T cells to destroy the cancer cell.
A. Naked mAbs: Without any drugs, radioactive materials or toxins.
B. Conjugated mAbs: They are combined with chemotherapy
drugs (Chemolabeled Ab) or radioactive material or toxins
(Immunotoxin)

33

34

When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more visible to the immune
system
• Block growth signals
• Deliver radiation to cancer cells.
• Slip powerful drugs into cancer cells
35

mAbs as antitumor agents
Trastuzumab is effective only in cancers where
HER2 is over-expressed. After binding to HER2,
the Mab causes an increase in p27, a protein
that halts cell proliferation. Another MAb,
Pertuzumab, which inhibits dimerization of HER2
and HER3 receptors, was approved by the FDA
for use in combination with trastuzumab in June
2012
HER2 (Human Epidermal Growth Factor
Receptor 2) also known as CD340 (cluster of
differentiation340) /p185 is a protein that in
humans is encoded by the ERBB2 gene.
Amplification or over-expression of this gene
has been shown to play an important role in
the pathogenesis and progression of certain
aggressive types of breast cancer

Trastuzumab

36

Antitumor therapy
Treatment(s)

Approved
in

Antibody

Target

Trastuzumab

HER2

Breast cancer & lymphomas 1998

Rituximab

CD20

B cell lymphomas

1997

Tositumomab

CD20

B cell lymphomas

2003

Alemtuzumab

CD52

Chronic lymphocytic
leukemia

2001

Bevacizumab

Vascular
endothelial
growth factor

Colorectal cancer

2004

Cetuximab

Epidermal
growth factor
receptor

Colorectal cancer

2004

Panitumumab

EGF receptor

Colorectal cancer

Gemtuzumab

CD33

Acute myelogenous
leukemia

2000

Ibritumomab

CD20

Non Hodgkin lymphoma

2002
37

Rituximab
Pharmacology and M/A
• Rituxan is the first MAb approved in 1997 by FDA. Chimeric
MAb that binds to the antigen CD20 found on B-lymphocytes
(normal B cell and most malignant B cells).
• CD20 is involved in cell cycle initiation, regulation and
differentiation by activation of B-cells from the G0 to G1 phase.
It also operates as a calcium channel.
• Used in low, intermediate and high grade lymphomas/NonHodgkin
Lymphoma
(NHL)
(in combination
with
cyclophosphamide, doxorubicin, vincristine and prednisone).
• It can also be given with radioimmunotherapy given directly to
the tumor site
• M/A: Rituximab causes CD20-positive cell death by antibody
dependent cell-mediated cytotoxicity, direct effects via CD20
ligation, and complement-mediated lysis.
38

Therapeutic agents:
Antibody

Target

Treatment(s)

Supportive Care

OTK3 (muromonab-CD3)

T-cell CD3 receptor

Transplant Rejection

Daclizumab

IL2 Receptor

Transplant Rejection

Infliximab

TNF α

Rheumatoid arthritis

Omalizumab

Ig E

Allergy related Asthma

Natalizumab

T cell VLA4 receptor

Multiple sclerosis therapy

Adalimumab

TNF α

Inflammatory diseases

Basiliximab

IL2 Receptor

Transplant Rejection

Other uses
Abciximab

Glycoprotein IIb/IIIa

Antiplatelet/

Efalizumab

CD11a

Psoriasis

Palivizumab

An epitope of F protein
of RSV

Respiratory Syncytial
39
Virus

Tissue transplantation
Muromonoab-CD3:
Muromonoab-CD3 is used for the
treatment of acute organ
transplant rejection. It is effective
in preventing graft rejection after
kidney, heart or liver
transplantation.
Muromonoab-CD3 is effective in
patients who after acute cardiac
or liver allograft rejection do not
respond to steroid therapy.
CD3 (cluster of differentiation 3) T-cell coreceptor is a protein complex and is composed of
four distinct chains. These chains associate with a
molecule known as the T-cell receptor (TCR)

40

It is a chimeric monoclonal
antibody produced by recombinant
DNA technology and is directed
against TNF-α.
It is composed of human constant
and mouse variable regions.
Infliximab binds to the soluble and
the membrane bound form of TNFα,
resulting in the neutralization of its
biological activity.
Rheumatoid Arthritis is a chronic, autoimmune disease
characterized by Severe joint inflammation, Increased synovial
fluid , and thickened synovial membrane. It occurs due to
destruction of bone and cartilage in several joints & Elevated
levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)41
α, IL-1, IL-6

Efalizumab
Efalizumab is a humanized IgG1 antibody produced by
recombinant DNA technology.
It exhibits immunosuppressive function. It binds to CD11a, which
is the α-subunit of leukocyte function antigen (LFA)-1.
Efalizumab decreases the cell surface expression of CD11a, which
is expressed on all leukocytes.

• Psoriasis is a disease of the immune system that
involves T lymphocytes.
• It results from complex communications that cause
activation of T lymphocytes and trafficking to the
skin.
• Further reactivation causes inflammation and
overproduction of skin, resulting in lesions and
plaques

42

In the treatment of diabetes, novel and improved therapeutic
modalities for those individuals with impaired insulin secretory
function would be helpful. Currently, long-acting basal insulins are
given daily, or more often, and are associated with both
hypoglycemia and weight gain.
Therefore, a highly specific, ultra-long-acting activator of INSR,
such as a monoclonal antibody, would represent a new paradigm in
diabetes therapy.
XMetA, an allosteric activator of INSR both in vitro and in vivo,
has the potential to normalize glycemic control in a model of
insulinopenic diabetes without causing hypoglycemia or promoting
weight gain.
• Of the more than 20 monoclonal antibodies generated to combat infectious
diseases that are in clinical development in 2011, most are in phase 1 or 2
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and are directed against either viruses or bacterial toxins.

Thank
ou

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