Transcript Photodynamic Therapy - UFCH JH

CZECH TECHNICAL UNIVERSITY IN PRAGUE
FACULTY OF BIOMEDICAL ENGINEERING
Photodynamic Therapy
Martin Hof, Radek Macháň
Photodynamic Therapy (PDT)
• Origin of tumors and the principles of their treatment
• Principles and history of PDT
• Photo-physical and -chemical aspects
• Photosensitizers (PS) of 1. generation
• Endogenous photosensitizer
• Photosensitizers of 2. and 3. generation
• Summary
Origin of tumors
A. Individual cells with modified
genome
A
B
B. Hyperplasia: mutated cells are
phenotypically identical with the healthy
ones but they multiply faster
C. Dysplasia: abnormalities in
cell shape and orientation
C
Origin of tumors
D. Noninvasive carcinoma: The cells
differ more in appearance and
multiplication rate. The tumor does
not spread to other tissues
E. Invasive carcinoma: Spreading out of
the tissue of origin: individual cells are
transported by cardiovascular and
lymphatic system; a malignant tumor can
lead to metastasis over the whole body
D
E
Principles of tumor treatment
Traditional
New developments
• Surgery
• Radiotherapy
• Chemotherapy
• Boron neutron capture
therapy
• Monoclonal antibody
therapy
• Antigene or antisense
therapy
• Photodynamic therapy
(PDT)
Principles of photodynamic therapy of
tumors (PDT)
• Photosensitizers (PS) are not
toxic “at dark”
• PS accumulate in tumors
• Illumination of the tumor
leads to
a) Fluorescence: diagnosis of
the tumor
b) Killing of tumor cells
(Apoptosis, Necrosis) - PDT
History of photodynamic therapy of tumors
• 1900 Acridin exhibits photo-toxicity
(Raab)
• 1903 Eosin applied against skin cancer
(von Tappeiner)
• 1908 Photo-toxicity of porphyrins (Hausmann)
• 1913 Mayer-Betz tested photodynamic therapy with porphyrins on his
skin
• 1924 Porphyrin enriched tissue exhibits red fluorescence upon
illumination with UV radiation (Policard)
• 1942 Different retention of porphyrins in helathy and malignant
tissues (Auler, Banzer)
History of photodynamic therapy of tumors
• 1948 Diagnosis and treatment of cancer by
hematoporphyrin and its complexes with zinc
(Figge)
• 1961 Lipson developed
derivative (HpD).
a
hematoporphyrin
• 1966 first successful breast cancer treatment by
HpD (Lipson)
• 1978 first systematic clinical studies (Dougherty)
• Today a few thousand patients treated by HpD
Photosensitizers of the first generation
Photosensitizers of the first generation
are oligomers (HpD) of Hematoporphyrin (Hp)
Hp
Dihematoporphyrinether
• Absorption at 405, 505, 525, 565 and
630 nm
• Emission at 635 and 700 nm
• Accumulation in tumors
Freie Radikale
O2
Photophysics (Jablonski)
Intersystem
ISC Crossing: kisc
S1
Energie
Energy
des
of the
Zustandes
states
Fluorescence: kf
Fluoreszenz
kPDD
nrS
T1
O2
SUPEROXID
Nonradiative transitions: knrT
S
Phosphorescence:1 kp
Singulett-Sauerstoff
Absorption
S0
Quantum Yields:
F:
f = kf / (kf + knrS + kisc)
REAKTIONEN TY
ISC: T0 isc = kisc / (kf +Oxidationen
knrS + kisc)
P:
p = isc  kp / Cycloadditionen
(kp + knrT)
an:
Triplett-Sauerstoff
ungesättigte Lipide,
Cholesterin, Proteine
Excited state reactions of photosensitizer in T1 state represent an
additional nonradiative decay pathway. Reaction with O2 gives rise to
singlet oxygen 1O2. Electron transfer reactions give rise to free radicals
What is 1O2?
• O2 is paramagnetic (in triplet
state) in the ground state
(according to Hund rule)
• Because of spin restriction
triplet oxygen 3O2 participates
only in non-selective radical
reactions
E
Electron
configuration
of 3O2
• Singlet oxygen 1O2 is very
reactive and selective
95 KJ/mol
3O
2
1269 nm
1O
2
REAKTIONEN
REAKCE TYP I
Photochemistry
Freie
Radikale
Volné
radikály
O2
FREE RADICALS
S1
ISC
Energie
Energy
des
of the
Zustandes
states
Fluoreszenz
PDD
Absorption
S0
Type 1 reaction
(electron transfer)
T1
k
+ O2
O2
SUPEROXID
Reactive Oxygen
Species (ROS):
Superoxide ·O2S1
Hydroxyl rad. ·OH
O2
Singulett-Sauerstoff
…
Type 2 reaction
(energy transfer)
T0
Triplett-Sauerstoff
 (1O2 ) = isc  k [3O2 ] / (k [3O2 ] +
REAKTIONEN TYP II
Oxidationen
Cycloadditionen an:
ungesättigte Lipide,
kp Cholesterin,
+ knrT) Proteine
HpD accumulates
preferentially in
membranes
Plasma membrane
Affected sites
Nuclear
membrane
Mitochondria
outer
membrane
Endoplasmatic
reticulum
Lysosomes
inner
membrane
Reactions
of 1O2 and ROS
with biomolecules
Cause oxidative damage,
which can lead ultimately to
cell death
Peroxidation
Addition
on cycles
Oxidation
BEFORE
AFTER
BEFORE – The photodynamic diagnostics (PDD) of a
tumor
AFTER – The tumor tissue has been removed by PDT
Pros and cons of the 1. generation of
photosensitizers
• Photophysics: high isc and  (1O2 ), but relatively short
wavelength absorption with a low absorption coefficient
• in vivo activity: low dark activity, high photodynamic
activity, but relatively low selectivity of absorption in tumors
An example of HpD-PDT
INJECTION
INJEKTION
intravenous
intravenös, 1 – 5 mg/kg
AKKUMULATION IM
EWEBE
ACUMULATION
INGTISSUE
ELIMINATION
E LIMINIERUNGFROM
AUS :
normales Gewebe
Healthy
tissue
Haut
Skin
Serum
T UMORGEWEBE
IN TUMOR
Serum ACUMULATION
16 –30 h
B ESTRAHLUNG
DES T UMORS
TUMOR ILLUMANTION
( 50-800 Jcm-2 )
1 – 2 Wochen
weeks
NEKROSE
APOPTOSIS
NECROSIS
Endogenous PS:
Cells produce their own PS
Pp IX
ALA
COOCH2
• The physiological
concentration of Pp IX
is low because of a
controlled expression
of its precursor 5aminolevulinic acid
(ALA)
O
CH2 NH2
ALA-synthase
succinyl-CoA
+
glycine
MITOCHONDRIA
CH2
C
• Photosesitizer
protoporphyrin IX (Pp
IX) is an intermediate
of heme synthesis
Ferrochelatase
Heme
• The expression of ALA
is feedback controlled
via heme
concentration
Endogenous PS:
Cells produce their own PS
Pp IX
ALA
COOCH2
C
O
CH2 NH2
ALA-synthase
succinyl-CoA
+
glycine
MITOCHONDRIA
CH2
Ferrochelatase
Heme
• Administration of
exogenous ALA breaks
the feedback control
and results in
accumulation of Pp IX
• Concentration of Pp IX
is higher in cancer
cells due to their
higher metabolic
activity and in some
cases also due to
decreased efficiency of
ferrochelatase and
increased efficiency of
Pp IX synthesis from
ALA
NH
H3C
NH
N
N
NH
N
Photosensitizers of 2. generation:
CH3 H3C
Hämatoporphyrin
(Hp)
H3C
CH3
CH3
R
HO2C
CO2H
CO2H
C
HO C
CO H
- long wavelength
absorption withHO
large
extinction coefficient
R = H, SO
3H,NHCOPh
Dihämatoporphyrinether
Hämatoporphyrin
(Hp)
selective
accumulation
in tumor
M = Zn
(ein aktiver Bestandteil von HpD)
2
2
N
R
N
R
R
N
RR
N
M
N
N
N
R
N
R
N
N
N
N
M
N
N
N
2
R
M
N
N
N
N
Phtalocyanine:
Phthalocyanin (Pc)
maximal ex = 740 nm
N
N
M = Zn, AlIII, SiIV, GeIV
R = H, SO3H
R
N
N
N
R = H, SO3H,NHCOPh
Naphtalocyanine:
M = Zn AlIII, SiIV, GeI
exM== Zn,
820 nm
R = H, SO3H
R
Phthalocyanin (Pc)
R
R
Naphthalocyanin (Nc)
MeO2C
MeO2C
MeO2C
MeO2C
H
H
N
HN
HN
N
NH
NH
N
NH
HO2C
Benzoporphyrin
N
EtO2C
N
HN
N
Porphycen:
ex = 710 nm
NH
HN
N
NHN
NH
NHN
Chlorin e6: N
HN
N
ex = 750
nm
N
CO2H
NH
HO2C
CO2CH3
CO2H
HO2C
Chlorin e6
CO2CH3
Etiopurpurin
N
HO2C
CO2H
Porphycen
Photosensitizers of 3. generation
(selective acculmulation)
• Monoclonal antibodies bind
selectively to an antigen on
cancer cells
• The spacer is either
cyclodextrin or Avidin-Biotinsystem
Antibody
PS
Spacer
„Drug Targeting“
Summary
HO
CH3
CH3
H3C
OH
N
NH
H3C
HN
CH3
N
CH3
HO2C
Limitations:
Hämatoporphyrin (Hp)
Low penetration depth in tissue (ideal
R with endoscopic
for skin cancer or
illumination)
Type
I reaction
REAKTIONEN
TYP I
Freieradicals
Radikale
Free
O2
ISC
Energie
Energy
des
of
Zustandes
states
Fluoreszenz
PDD
T1
O2
SUPEROXID
S1
N
N
Advantages:
N
RLow cost
N
M
N
Relatively low side
effects
N
N
Singulett-Sauerstoff
Singlet
Oxygen
Type II reaction
Absorption
S0
T0
Triplett-Sauerstoff
Triplet Oxygen
CO2H
REAKCE TYP II
REAKTIONEN
Oxidationen
Cycloadditionen an:
R
N
M = Zn, AlIII, SiIV, G
Goals:
R = H, SO H
High selectivity for cancer cells 3
R
Optimal illumination
dose
Phthalocyanin (Pc)
Methodological outlook:
Multiphoton excitation
• High intensity of ps- or fs-lasers
• Excitation by light of double or
triple wavelength compared to
single photon excitation
• Light of longer wavelength
penetrates deeper to the tissue
Longwave excitation of a PS with
shortwave absorption
Acknowledgement
The course was inspired by courses of:
Prof. David M. Jameson, Ph.D.
Prof. RNDr. Jaromír Plášek, Csc.
Prof. William Reusch
Financial support from the grant:
FRVŠ 33/119970