1 LADEK ZDRÓJ 2003

2 LASER SPECTROSCOPIC STUDY OF PHTHALOCYANINE DERIVATIVES SYNTHESIZED FOR PHOTODYNAMIC THERAPY András Grofcsik Budapest University of Technology and Economics Department of Physical Chemistry

3 Photodynamic therapy (PDT) Hematoporphyrin derivative (HpD) Phthalocyanines (Pc) Synthesis Photophysical properties in solutions in vesicles 5-Amino-levulinic acid (ALA)

4 Cancer therapies surgery radiotherapy chemotherapy Photodynamic therapy (PDT): use of visible light in combination with a photosensitiser

5 Number of publications in PDT

6 Administration of photosensitizer Irradiation with visible light Photosensitizer accumulates in the tumour. Tumour is selectively destroyed Steps of photodynamic therapy

7 Photodynamic effect: Cell destruction by photosensitiser + visible light + O 2 TYPE II: energy transfer TYPE I: electron-transfer:Radicals and radical ions

8 The wavelength dependence of depth of penetration of light into soft tissue

9 Requirements for the photosensitiser: Selective accumulation in malignant tissues High absorbance between 600 and 800 nm Chemical homogenity Long triplet lifetime and sufficient triplet energy (>94 kJ/mol) Chemical, biological and photochemical stability Little or no dark toxicity Simple and cheap syntesis

10 The first sensitiser used in clinical PDT: Hematoporphyrin derivative (HpD) Photofrin® It is a mixture of compounds.

11 Hematoporphyrin

12 HPLC analysis of HpD

13 Selective accumulation in malignant tissues High absorbance between 600 and 800 nm Chemical homogenity Long triplet lifetime and sufficient triplet energy (>94 kJ/mol) Chemical, biological and photochemical stability Little or no dark toxicity Simple and cheap syntesis Requirements for the photosensitiser:

14 ” Second generation" photosensitisers: porphyrins chlorins bacteriochlorins phthalocyanines (Pc) naphthalocyanines 5-Aminolevulinic acid (ALA)

15 Phthalocyanine

16 Absorpion spectrum of a porphyrin (a) and a phthalocyanine (b) derivative (Ethanol solutions, c = 1.5*10 -5 mol dm -3 )

17 Synthesis

18 Phthalocyanine derivatives I : M: Zn R: 4-tert-Bu-Ph- II : M: Zn R: CH 3 O(CH 2 ) 2 O(CH 2 ) 2 - III : M: Zn R: (2,6-dimethyl-4- N,N-dimethylamino- methylen)-phenyl- IV : M: H 2 R: CH 3 O(CH 2 ) 2 O(CH 2 ) 2 -

19 Structure of III

20 Experimental setup for studying triplet states

21 Triplet lifetime and triplet absorption spectrum ,000 0,002 0,004 0,006 0,008 0,010 0,012 0,014 Ethanol solution of I (7,7* mol/dm 3 ) Absorbance [nm] ,03 -0,02 -0,01 0,00 0,01 0,02 Triplet decay Absorbance t [ns]

22 Reaction of triplet Pc with molecular oxygen: The rate constant can be determined from the decay curves.

23 0,00,10,20,30,40,50,60,70, I 0 [mV] Laser energy [mJ] Ref I II III IV Quantum yield of singlet oxygen formation

24 Triplet lifetimes (  ), second order rate constants and quantum yields of singlet oxygen formation

25 Photosensitisers in vesicles Vesicles are simple models of cell membranes. DPPC (dipalmitoylphosphatidylcholine)

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28 Temperature dependence of the rate constant (III in DPPC vesicles) Arrhenius plot: E a = 60.7 kJ 20 20,4 20,8 21,2 21,6 22 0,003000,003050,003100,003150,003200,003250,003300,003350,003400, /T [1/K] ln k

29 Use of 5-amino-levulinic acid (ALA) for PDT ALA stimulates the cellular synthesis of an endogenous photosensitiser: Protoporphyrin IX Administration of exogenious ALA causes the build-up of phototoxic levels of Protoporphyrin IX

30 Advantages (over HpD): Treatment follows 2-4 hours after administration Systemic clearence of photosensitiser within 24 hours Treatment can be repeated within two days ALA can be administered topically The method was approved by FDA in the 90s) In Hungary clinical trials started in 2001 (National Medical Center)

31 Basal cell carcinoma - before treatment

32 After PDT with ALA

33 PARTICIPANTS Miklós Kubinyi István Bitter Viktor Csokai Janka Tatai Klára Szegletes Éva Bacskay János Brátán Tamás Vidóczy Péter Baranyai Lajos Csokonai Vitéz

34 The End