Transcript SINGAPORE’S R&D FRAMEWORK and the TECHNOLOGY DEVELOPMENT

Lecture 8: Computational Drug Target
Identification
Y.Z. Chen
Department of Computational Science
National University of Singapore
E-mail: [email protected]
Web-page: http://www.cz3.nus.edu.sg/~yzchen/
Outline
 Potential Application:
 Unknown and secondary therapeutic targets of drugs, leads,
natural products.
 Targets related to toxicity, side-effect, delivery.
 Ligand-protein interactions involved in pathways.
 Methodology:
 Ligand-protein inverse docking
 Tests:
 Known targets for therapeutics, toxicity and side effect.
Y.Z. Chen, National University of Singapore
E-mail: [email protected]
Web-page: http://www.cz3.nus.edu.sg/~yzchen/
Why
protein targets?
Novel or Unknown
Therapeutic Targets
Nature 1998, 396:15
Why protein targets?
Toxicity, side effect,
pharmacokinetics and pharmacogenetics
Annu. Rev. Pharmacol Toxicol 2000, 40:353-388
1997, 37:269-296
Pharmacological Rev. 2000, 52:207-236
Why protein targets?
Towards the prediction of side effect, toxicity,
pharmacokinetics in early stages of drug
discovery
 Most drug candidates fail
to reach market
 Pharmacokinetics, side
effect and toxicity are the
main reason.
Drug Candidates
in Different Stages of Development
Majority of Them Fail to Reach Market
Clin Pharmacol Ther. 1991; 50:471
 Large portion of money
($350 million per drug)
and time (6-12 years for a
drug) has been wasted on
failed drugs.
Drug Discov Today 1997; 2:72
Why protein
targets?
Drugs from Natural
Products
From natural products
to therapeutic drugs
TIPS, May 1999, 20:190
Traditional medicines
Pharmacology & Therapeutics 2000, 86:191



Screening or extraction of
bioactive compounds
Mechanism and standardization
Further development
Drug targets and new drug discovery
Drugs from Traditional Medicines


Mixture of multiple
herbs etc.
Therapeutic action +
maintaining and
restoring balance:
Mutual accentuation,
mutual enhancement,
mutual counteraction,
mutual suppression,
mutual antagonism,
mutual incompatibility

Multiple targets:
therapeutic effect,
symptom treatment,
toxicity modulation,
drug delivery,
harmonization
Pharmacology & Therapeutics 2000, 86:191
Why searching protein targets
of a molecule?
Applications in pathways
EGF Pathway
From Signaling Pathway Database
http://www.grt.kyushu-u.ac.jp/spad/
Strategy
Existing Methods:
Given a Protein,
Find Potential Binding Ligands
From a Chemical Database
New Method:
Given a Ligand,
Find Potential Protein Targets
From a Protein Database
Compound Database
Protein Database
Compound 1
...
Compound n
Protein 1
...
Protein n
Protein
Ligand
Successfully Docked Compounds
as Putative Ligands
Successfully Docked Proteins
as Putative Targets
Science 1992;257: 1078
Proteins 2001;43:217
Feasibility
Proteins
 Database: >12,000 3D structures in PDB.
 Protein diversity: 17% in PDB with unique sequence.
 Advances in structural genomics: 10,000 unique proteins within 5 years.
Ann. Rev. Biophys. Biomol. Struct. 1996; 25:113
Nature Struct. Biol. 1998; 5:1029
Method
 Ligand-protein docking docking algorithms capable of finding binding
conformations.
Proteins. 1999; 36:1
Proteins 2001; 43:217
Additional information
 Rapid accumulation of knowledge in proteomics, pathways, protein
functions (functional genomics).
Computer resources
 Increasing power and decreasing cost (Linux PC, Multi-processor
Machine)
Automated Protein Targets Identification Software
INVDOCK
Ligand
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Automated Process to inversely dock the Lignad to each entry in
a Built-In Biomolecular Cavity Database (10,000 Protein and Nucleic Acid Entries)
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Step 1: Vector-based docking of a ligand to a cavity
Step 2: Limited conformation optimization on the ligand and side chain of biomolecule
Step 3: Energy minimization for all atom in the binding site
Step 4: Docking evaluation by molecular mechanics energy functions and comparison with other ligands
Successfully Docked Proteins and Nucleic Acids
as Putative Targets of a Ligand
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Potential Applications:
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Protein function, Proteomics, Ligand transport, Metabolism
Therapeutic Targets, Side-Effects, Metabolism, Toxicity
Function in Pathways
INVDOCK Cavity Models
HIV-1 Protease
INVDOCK Cavity Models
Estrogen
Receptor
Energy Functions





Chemical bonds
Hydrogen bonding
van der Waals interactions
Electrostatic interactions
Empirical solvation free energy
V = Vbonds +
H bonds [ V0 (1-e-a(r-r0) )2 - V0 ] +
non bonded [ Aij/rij12 - Bij/rij6 + qiqj /r rij] +
atoms i Dsi Ai
INVDOCK Testing Results
Molecule
Docked Protein
PDB
Id
RMSD
Energy
Description of Docking Quality
Indinavir
HIV-1 Protease
1hsg
1.38
Match
-70.25
Xk263 Of Dupont
Merck
HIV-1 Protease
1hvr
2.05
Match
-58.07
Vac
HIV-1 Protease
4phv
0.80
Match
-88.46
Folate
Dihydrofolate Reductase
1dhf
2.41
5-Deazafolate
Dihydrofolate Reductase
2dhf
1.48
Match
-65.49
Estrogen
Estrogen Receptor
1a52
1.30
Match
-45.86
4-Hydroxytamoxifen
Estrogen Receptor
3ert
0.97
Guanosine-5'-[B,GMethylene]
Triphosphate
H-Ras P21
121p
0.94
Glycyl-*L-Tyrosine
Carboxypeptidase A a
3cpa
2.19
One end match, the other in
slightly different orientation
Match
Match
Match
-63.02
-55.15
-80.20
-44.84
INVDOCK Testing Results
Compound
Potential Targets
Identified
Experimentally
Confirmed
Experimentally
Implicated
4H-Tamoxifen
17
4
4
Aspirin
52
4
16
Vitamin C
46
4
9
Vitamin E
26
2
11
INVDOCK Identified Protein Targets For an Anticancer Drug Tamoxifen
PDB
Putative Protein Target
Experimental Finding
1a52
Estrogen Receptor
1akz
Uracil-DNA Glycosylase
1ayk
Collagenase
1az1
Aldose Reductase
1bnt
Carbonic Anhydrase
1boz
Dihydrofolate Reductase
1dht,
1fdt
17b -Hydroxysteroid Dehydrogenase
1gsd
,
3ljr
Glutathione Transferase A1-1,
Glutathione S-Transferase
Target
Status
Clinical Implication
Ref
Drug target
Confirmed
Treatment of breast cancer
36
Inhibited activity
Confirmed
Tumor cell invasion and cancer
metastasis
38
Decreased level
Inhibitor
Suppressed enzyme
and activity
Combination therapy for cancer
Confirmed
Promotion of tumor regression
43
39
41
Genotoxicity and carcinogenicity
INVDOCK Identified Protein Targets For an Anticancer Drug Tamoxifen
1mch
Immunoglobulin l Light Chain
Temerarily enhanced
Ig level
44
Modulation of immune response
1p1g
Macrophage Migration Inhibitory factor
1ulb
Purine Nucleoside Phosphorylase
1zqf
DNA Polymerase b
2nll
Retinoic Acid Receptor
1a25
Protein Kinase C
1aa8
D-Amino Acid Oxidase
1afs
1pth
Inhibition
3a -Hydroxysteroid Dehydrogenase
Effect on androgen
induced activity
Prostaglandin H2 Synthase-1
Direct inhibition
Confirmed
Confirmed
Anticancer
37
Hepatic steroid metabolism
42
Prevention of vasoconstriction
40
INVDOCK Testing on Toxicity Targets
Compound
Number of
experimentally
confirmed or
implicated toxicity
targets
Number of
toxicity targets
predicted by
INVDOCK
Number of
toxicity
targets
missed by
INVDOCK
Number of
toxicity targets
without 3D
structure or
involving
covalent bond
4
Number of
INVDOCK
predicted toxicity
targets without
experimental
finding
2
Aspirin
15
9
2
Gentamicin
17
5
2
10
2
Ibuprofen
5
3
0
2
2
Indinavir
6
4
0
2
2
Neomycin
14
7
1
6
6
Penicillin G
7
6
0
1
8
Tamoxifen
2
2
0
0
4
Vitamin C
2
2
0
0
3
Total
68
38
5
25
29
Toxicity and side effect targets of Aspirin
identified from INVDOCK search of protein database
PDB
Protein
1a42
Carbonic anhydrase II
1a6a
HLA-DR3
1a7c
Plasminogen activator
inhibitor
1d6n
Hypoxanthine-guanine
phosphoribosyltransferase
1hdy
Alcohol dehydrogenase
Experimental
Finding
Target
Status
Toxicity/Side Effect
Ref
Activate enzyme
activity that
may lead to
increase in
plasma
bicarbonate
concentration.
Change in HLA
level
Implicated
Metabolic alkalosis
(hypoventilation).
Puscas I
Implicated
Aspirin-induced
asthma
Dekker JW
Tissuedependent
response of
protein.
Implicated
Hypertension,
thrombolysis
Smokovitis
A
Excess uric acid in
serum*
Inhibition of
activity
Confirmed
Increased blood
alcohol level
Gentry RT
Toxicity and side effect targets of Aspirin
identified from INVDOCK search of protein database
1hdy
Alcohol dehydrogenase
Inhibition of
activity
Confirmed
Increased blood
alcohol level
Gentry RT
1hiq
Insulin
Tissue
insensitivity to
insulin
Implicated
Impaired glucose
metabolism in
insulin-sensitive
cells.
Newmann
WP
1hmr
Fatty acid binding protein
Increased
binding capacity
and protein
content.
Implicated
Effect on
peroxisomal beta
oxidation activity.
Kawashima
Y
1mch
Immunoglobulin lambda light
chain
Ig reactivities
Implicated
Allergic reaction to
aspirin.
Zhu DX
1pah
Phenylalanine hydroxylase
2ant
Antithrombin
Irreversibly
acetylate
antithrombin.
Confirmed
Blood coagulation,
thrombolysis.
Villanueva
GB
2hdh
L-3-hydroxyacyl CoA
dehydrogenase
Reversible
inhibition of
enzyme activity.
Confirmed
Effect on Reye's
syndrome patients.
Glasgow JF
Phenylketonurea*
Molecular targets of Chinese natural products
Chinese
Natural Product
Number of Identified
Therapeutic Targets
Number Confirmed or
Implicated
Therapeutic Targets
by experiment
Number of Identified
Toxicity/Side effect
Targets
Number Confirmed
or Implicated
Toxicity/Side Effect
Targets by
experiment
Acronycine
3
1
4
-
Allicin
5
2
1
1
Baicalin
14
4
6
-
Catechin
17
12
5
-
Camptothecine
9
6
3
2
Dicoumarin
7
1
3
1
Emodin
6
3
5
1
Genistin
22
7
12
1
Putative and known therapuetic targets of Camptothecine identified from
INVDOCK search of human and mammalian proteins
PDB
Protein
Experimental
Finding
Target
Status
Theraputic
Effect
Ref
1ads
Aldose Reductase
2gss
Glutathione STransferase p1-1
7ice
DNA Polymerase
Beta
1a25
Protein Kinase C
1cdk
CAMP-Dependent
Protein Kinase
Anti-cancer
3bct
Beta-Catenin
Anti-cancer
1dvi
Calpain
Inhibition of calpain activities.
Implicated
Induces apoptosis in leukemic
cells.
Eymin
1yfo
Receptor Protein
Tyrosine
Phosphatase
Causes elevation of PTPase in the
cytosol and the nucleus which
play a critical role in the
induction of the differentiation of
IW32 erythroleukemia cells.
Implicated
Anti-cancer
Wang MC
1a35
Topoisomerase I
Inhibitor
Confirmed
Anti-cancer
Wang MC
Diabetes treatment
Increases intracellular
glutathione
Implicated
Enhance radical scavenging
activities that may useful in
cancer treatment
Matsumoto
Anti-cancer
Inhibitor
Confirmed
Induction of apoptosis in
tumor.
Martelli
Nieves-Neira
Putative and known therapuetic targets of Camptothecine identified from
INVDOCK search of human and mammalian proteins
PDB
Protein
Experimental
Finding
Target
Status
Theraputic
Effect
Ref
1ads
Aldose Reductase
2gss
Glutathione STransferase p1-1
7ice
DNA Polymerase
Beta
1a25
Protein Kinase C
1cdk
CAMP-Dependent
Protein Kinase
Anti-cancer
3bct
Beta-Catenin
Anti-cancer
1dvi
Calpain
Inhibition of calpain activities.
Implicated
Induces apoptosis in leukemic
cells.
Eymin
1yfo
Receptor Protein
Tyrosine
Phosphatase
Causes elevation of PTPase in the
cytosol and the nucleus which
play a critical role in the
induction of the differentiation of
IW32 erythroleukemia cells.
Implicated
Anti-cancer
Wang MC
1a35
Topoisomerase I
Inhibitor
Confirmed
Anti-cancer
Wang MC
Diabetes treatment
Increases intracellular
glutathione
Implicated
Enhance radical scavenging
activities that may useful in
cancer treatment
Matsumoto
Anti-cancer
Inhibitor
Confirmed
Induction of apoptosis in
tumor.
Martelli
Nieves-Neira