2003 LADEK ZDRÓJ

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LADEK ZDRÓJ

2003

LASER SPECTROSCOPIC STUDY

OF PHTHALOCYANINE DERIVATIVES SYNTHESIZED FOR PHOTODYNAMIC

THERAPY

András Grofcsik

Budapest University of Technology and Economics

Department of Physical Chemistry

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Photodynamic therapy (PDT)

Hematoporphyrin derivative (HpD) Phthalocyanines (Pc)

Synthesis

Photophysical properties in solutions

in vesicles

5-Amino-levulinic acid (ALA)

Cancer therapies surgery

radiotherapy chemotherapy

Photodynamic therapy (PDT): use of visible light in combination with a photosensitiser

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Number of publications in PDT

0 500 1000 1500 2000 2500 3000

1981-85 1986-90 1991-95 1996-2000

Administration of

Irradiation with visible light Photosensitizer

accumulates in

Tumour is selectively

Steps of photodynamic therapy

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Photodynamic effect:

Cell destruction by

photosensitiser + visible light + O

) (

) (

)

( S

P S

P T

P  





Products O

es Biomolecul

O S

P O

T P











* 2 1

* 2 1 0

2 3

1

) ( )

(

•TYPE II: energy transfer

•TYPE I: electron-transfer: Radicals and radical ions

vessel blood

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

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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

The first sensitiser used in clinical PDT:

Hematoporphyrin derivative (HpD) Photofrin®

It is a mixture of compounds.

Hematoporphyrin

HPLC analysis of HpD

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• 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:

”

Second generation" photosensitisers:

porphyrins chlorins

bacteriochlorins

phthalocyanines (Pc) naphthalocyanines

5-Aminolevulinic acid (ALA)

Phthalocyanine

0 0,2 0,4 0,6 0,8 1 1,2 1,4 1,6 1,8

500 550 600 650 700 750



Absorbance

b

a

Absorpion spectrum of a porphyrin (a) and a phthalocyanine (b) derivative (Ethanol solutions,

c = 1.5*10

mol dm

)

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Synthesis

4 M = 2H M = Zn ROH, DMF, K₂CO₃

60 W, 20 min

3 2

1

60 W, 3 minDBU Zn(OAc)₂ 60 W, 15 min

K₂CO₃

ROH, DMF CN

CN RO

CN CN NO₂

OR

RO

OR

N N

N

N N N

N N

OR M

N N

N M N

N

Phthalocyanine derivatives

I^{: M: Zn}

R: 4-tert-Bu-Ph-

II^{: M: Zn}

R: CH₃O(CH₂)₂O(CH₂)₂-

III^{: M: Zn}

R: (2,6-dimethyl-4- N,N-dimethylamino-

IV^{: M: H}₂

R: CH₃O(CH₂)₂O(CH₂)₂-

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Structure of III

Experimental setup for studying triplet states

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Triplet lifetime and triplet absorption spectrum

I I

lg I A

0 0



 

 

k A

ln A

ln  ₀   ₂  ^{400 450 500 550 600}

0,000 0,002 0,004 0,006 0,008 0,010 0,012 0,014

Ethanol solution of I (7,7*10^--6 mol/dm³)

Absorbance

 [nm]

0 500 1000 1500 2000 2500

-0,03 -0,02 -0,01 0,00 0,01

0,02 Triplet decay

Absorbance

t [ns]

Reaction of triplet Pc with molecular oxygen:

2 1 0

2 3

1

) ( )

( T O Pc S O

Pc   

The rate constant can be determined from the decay curves.

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0,0 0,1 0,2 0,3 0,4 0,5 0,6 0,7 0,8 0

5 10 15 20 25

I 0 [mV]

Laser energy [mJ]

Ref I II III IV

Quantum yield of singlet oxygen formation

ref ref

phtal phtal

m

m  





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

Pc

(solvent) I (Et) I (To) II (Et) II (To) III (Et) IV (To)

 (ns) 306 287 381 263 231 504

k*10^-9

(M^-1s^-1)

1.55 1.67 1.25 1.81 2.09 0.95

 0.47 0.58 0.42 0.44 0.59 0.19

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Photosensitisers in vesicles

Vesicles are simple models of cell membranes.

DPPC (dipalmitoylphosphatidylcholine)

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Temperature dependence of the rate constant (III in DPPC vesicles)



 



  

 RT

exp E A

k ^a Arrhenius plot:

E_a = 60.7 kJ

20 20,4 20,8 21,2 21,6 22

ln k

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Use of 5-amino-levulinic acid (ALA) for PDT

COOH CH₂ CH₂ C O

CH₂ NH₂

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

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)

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Basal cell carcinoma - before treatment

After PDT with ALA

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PARTICIPANTS

Miklós Kubinyi István Bitter

Viktor Csokai Janka Tatai

Klára Szegletes

Éva Bacskay János Brátán Tamás Vidóczy Péter Baranyai

Lajos Csokonai Vitéz

The End