- Email: [email protected]
In vitro and in vivo photosensitization
391
In vitro and in vivo photosensitization G. Jori Department
of Biology,
University
of Padova
@al~_l
Photosensitized processes are attracting the interest of a large number of photobiologlsts as a consequence of recently developed novel applications in important fields, such as photomedicine and photoecology. The attention of investigators is mainly focused on photosensitization in heterogeneous media, which often mimic specific situations occurring in vivo. Attempts to correlate the photophysical parameters (quantum yield of photogeneration of the photosensitizer electronically excited states; lifetime and reactivity of these excited states as well as of transients - activated oxygen species, substrate radicals, etc.) with the physico-chemical properties and the optical characteristics of the system are made. Thus, the competition between type I (hydrogen or electron transfer between the photoexcited sensitizer and the substrate) and type II (energy transfer from the photoexcited sensitizer to a suitable substrate) can be modulated by controlling the partial oxygen pressure ln a given cellular or tissular compartment or by taking advantage of the changes in the reactivity of intermediate species in different mlcroenvironments (e.g. across lipid/water interphases, hydrophobic/hydrophilic domains of a macromolecule, etc.) Moreover, the extent of photosensitized effects in vivo can be modulated by the wavelength-dependence of light penetration into tissues. This factor is controlled by the light-scattering and light-absorption properties of the tissue, e.g. the size and geometry of the subcellular or endotissular components, the degree of pigmentation, etc. In the case of mammahan tissues, it has been experimentally observed that the depth of light penetration usually increases with increasing wavelength and reaches maximal values in the 700-1000 run spectral interval. On the other hand, light penetration into tissues is . . nnmmal in the UV region as a result of large scattering of the incident light and strong absorption by endogenous chromophores, such as proteins, nucleic acids, urocanic acid and melanin. Light penetration decreases again in the IR mainly because of the water absorption bands. Therefore, UV or blue light wavelengths should be chosen when the photoeffects are to be confined within a narrow spatial range from the site where light interacts with the tissue, while the use of red light allows the uniform illumination of relatively large tissular volumes. A further enhancement of the selectivity of in vivo photosensitization is dependent on the competition between light absorption by endogenous cell constituents and externally added photosensitlzers. The situation is especially favourable with photosensitizers absorbing above 600 nm, where light absorption by endogenous chromophores is generally minimal. Hence, those photosensitizers which exhibit strong absorption bands in the red (porphyrins, chlorlns, phthalocyanines, naphthalocyanines, etc.) appear to be most suitable for in vivo photosensitization. Moreover, we can often obtain a spatial selectivity by introducing the photosensitizer into the system in association with a carrier directed toward a specific cell organelle or tissue district. The approaches developed so far include the use of photosensitizers selectively accumulated by mitochondria or bound to monoclonal antibodies directed against antigens which are present at the surface of specific cell populations. 1 G. Moreno, R. H. Pottier and T. G. Truscott (eds.), Photosensitization: Molecular, CeUular and Medical Aspects, Springer, Berlin, 1988. 2 G. Bock and S. Harnett (eds.), Photosensitizing Compounds: their Chemistry, Biolom and Clinical Use, CIBA Foundation Symposium 146, Wiley, Chichester, 1989.
392
3 J. A. Parrish, M. L. Kripke and W. L. Morison (eds.), Photoimmunology, Plenum, New York, 1983. 4 G. Jori, Photosensitized processes in vivo: proposed phototherapeutic applications, Photochem. Photobiol., 52 (1990) 439-443.
Photodynamic
therapy:
basic and pre-clinical
studies
G. Jori Department
of Biology,
University
of Padova
(Italy)
Photodynamic therapy (PDT) is a recently developed modality for treatment of tumours although its possible extension to the treatment of atheromas, psoriasis and viral or microbial infections is presently under investigation in several centres. PDT is based on the property of some “photodynamlc-type” photosensitizers (i.e. photosensitlzers requiring oxygen ln order to exert their action) to be accumulated in signiilcant amounts and retained for prolonged periods of time by tumour tissues. In several cases, the concentration of the photosensitlzer in the tumour is larger than in peritumoral tissues, so that the photodamage preferentially involves the neoplastic areas. The selectivity of the phototherapeutic process and the extent of the photodamage can be enhanced through the use of photosensitizers absorbing in the wavelength region above 600 run. At present, most clinical applications of PDT are carried out using a chemical derivative of hematoporphyrin, Photofrln II. At the doses used for clinical PDT (1.5-3 mg kg-’ body weight) Photofrin II localizes in tumours in concentrations around 1 pg g-’ of tissue and, upon excitation with 630 nm light, photosensitizes tumour necrosis mainly via b-reversible damage of tumour vasculature. Many different types of solid tumours have been found to respond to PDT with Photofrln II; one exception being pigmented melanoma, the poor response of which to PDT is likely to be a result of the high melanin content, which intercepts the incident light. Moreover, PDT can be used for both eradication of relatively small tumours and palliation of inoperable or obstructing tumours. A major limitation of PDT is represented by the composite nature of Photofrin II and its low extinction coefficient at wavelengths longer than 600 run. Thus, several new “second generation” photosensitizers are being tested as possible candidates to replace Photofrin II in PDT. Preference is given to hydrophobic-type photosensitizers, which have been shown to yield optimal tumour localization. This may be (at least in part) because of the transport of these photosensitizers via serum lipoproteins, since one lipoprotein class (low density llpoprotelns, LDL) gives a preferential interaction with tumour cells through receptor-mediated endocytosis. For those photosensitizers which are waterinsoluble, in vivo administration can be performed after incorporation into liposomes, oil emulsions or inclusion complexes. Among the most promising new photosensitizers, one can list some purpurins, phthalocyanines, naphthalocyanines, and benzoporphyrin derivative. The hydrophobic character of such photosensitlzers favours the occurrence of direct photodamage of malignant cells ln the tumour as compared with vascular damage. Thus, the possibility exists to control the mechanism of PDT-action on tumours through a suitable choice of the physico-chemical properties of the photosensitlzer. In particular, sparing the vascular system allows the supply of oxygen to the tumour tissue for some hours after the end of PDT, which should increase the efficiency of cell damage and tumour response to the phototreatment. 1 J. T. Dougherty (ed.), Photodynamic therapy: mechanisms,SPZE Proc., (1989) 1065. 2 D. Kessel (ed.), Photodynamic Therapy of Neoplastic Disease, Vols. I and II, CRC Press, Boca Raton, FL, 1990.
In vitro and in vivo photosensitization G. Jori Department
of Biology,
University
of Padova
@al~_l
Photosensitized processes are attracting the interest of a large number of photobiologlsts as a consequence of recently developed novel applications in important fields, such as photomedicine and photoecology. The attention of investigators is mainly focused on photosensitization in heterogeneous media, which often mimic specific situations occurring in vivo. Attempts to correlate the photophysical parameters (quantum yield of photogeneration of the photosensitizer electronically excited states; lifetime and reactivity of these excited states as well as of transients - activated oxygen species, substrate radicals, etc.) with the physico-chemical properties and the optical characteristics of the system are made. Thus, the competition between type I (hydrogen or electron transfer between the photoexcited sensitizer and the substrate) and type II (energy transfer from the photoexcited sensitizer to a suitable substrate) can be modulated by controlling the partial oxygen pressure ln a given cellular or tissular compartment or by taking advantage of the changes in the reactivity of intermediate species in different mlcroenvironments (e.g. across lipid/water interphases, hydrophobic/hydrophilic domains of a macromolecule, etc.) Moreover, the extent of photosensitized effects in vivo can be modulated by the wavelength-dependence of light penetration into tissues. This factor is controlled by the light-scattering and light-absorption properties of the tissue, e.g. the size and geometry of the subcellular or endotissular components, the degree of pigmentation, etc. In the case of mammahan tissues, it has been experimentally observed that the depth of light penetration usually increases with increasing wavelength and reaches maximal values in the 700-1000 run spectral interval. On the other hand, light penetration into tissues is . . nnmmal in the UV region as a result of large scattering of the incident light and strong absorption by endogenous chromophores, such as proteins, nucleic acids, urocanic acid and melanin. Light penetration decreases again in the IR mainly because of the water absorption bands. Therefore, UV or blue light wavelengths should be chosen when the photoeffects are to be confined within a narrow spatial range from the site where light interacts with the tissue, while the use of red light allows the uniform illumination of relatively large tissular volumes. A further enhancement of the selectivity of in vivo photosensitization is dependent on the competition between light absorption by endogenous cell constituents and externally added photosensitlzers. The situation is especially favourable with photosensitizers absorbing above 600 nm, where light absorption by endogenous chromophores is generally minimal. Hence, those photosensitizers which exhibit strong absorption bands in the red (porphyrins, chlorlns, phthalocyanines, naphthalocyanines, etc.) appear to be most suitable for in vivo photosensitization. Moreover, we can often obtain a spatial selectivity by introducing the photosensitizer into the system in association with a carrier directed toward a specific cell organelle or tissue district. The approaches developed so far include the use of photosensitizers selectively accumulated by mitochondria or bound to monoclonal antibodies directed against antigens which are present at the surface of specific cell populations. 1 G. Moreno, R. H. Pottier and T. G. Truscott (eds.), Photosensitization: Molecular, CeUular and Medical Aspects, Springer, Berlin, 1988. 2 G. Bock and S. Harnett (eds.), Photosensitizing Compounds: their Chemistry, Biolom and Clinical Use, CIBA Foundation Symposium 146, Wiley, Chichester, 1989.
392
3 J. A. Parrish, M. L. Kripke and W. L. Morison (eds.), Photoimmunology, Plenum, New York, 1983. 4 G. Jori, Photosensitized processes in vivo: proposed phototherapeutic applications, Photochem. Photobiol., 52 (1990) 439-443.
Photodynamic
therapy:
basic and pre-clinical
studies
G. Jori Department
of Biology,
University
of Padova
(Italy)
Photodynamic therapy (PDT) is a recently developed modality for treatment of tumours although its possible extension to the treatment of atheromas, psoriasis and viral or microbial infections is presently under investigation in several centres. PDT is based on the property of some “photodynamlc-type” photosensitizers (i.e. photosensitlzers requiring oxygen ln order to exert their action) to be accumulated in signiilcant amounts and retained for prolonged periods of time by tumour tissues. In several cases, the concentration of the photosensitlzer in the tumour is larger than in peritumoral tissues, so that the photodamage preferentially involves the neoplastic areas. The selectivity of the phototherapeutic process and the extent of the photodamage can be enhanced through the use of photosensitizers absorbing in the wavelength region above 600 run. At present, most clinical applications of PDT are carried out using a chemical derivative of hematoporphyrin, Photofrln II. At the doses used for clinical PDT (1.5-3 mg kg-’ body weight) Photofrin II localizes in tumours in concentrations around 1 pg g-’ of tissue and, upon excitation with 630 nm light, photosensitizes tumour necrosis mainly via b-reversible damage of tumour vasculature. Many different types of solid tumours have been found to respond to PDT with Photofrln II; one exception being pigmented melanoma, the poor response of which to PDT is likely to be a result of the high melanin content, which intercepts the incident light. Moreover, PDT can be used for both eradication of relatively small tumours and palliation of inoperable or obstructing tumours. A major limitation of PDT is represented by the composite nature of Photofrin II and its low extinction coefficient at wavelengths longer than 600 run. Thus, several new “second generation” photosensitizers are being tested as possible candidates to replace Photofrin II in PDT. Preference is given to hydrophobic-type photosensitizers, which have been shown to yield optimal tumour localization. This may be (at least in part) because of the transport of these photosensitizers via serum lipoproteins, since one lipoprotein class (low density llpoprotelns, LDL) gives a preferential interaction with tumour cells through receptor-mediated endocytosis. For those photosensitizers which are waterinsoluble, in vivo administration can be performed after incorporation into liposomes, oil emulsions or inclusion complexes. Among the most promising new photosensitizers, one can list some purpurins, phthalocyanines, naphthalocyanines, and benzoporphyrin derivative. The hydrophobic character of such photosensitlzers favours the occurrence of direct photodamage of malignant cells ln the tumour as compared with vascular damage. Thus, the possibility exists to control the mechanism of PDT-action on tumours through a suitable choice of the physico-chemical properties of the photosensitlzer. In particular, sparing the vascular system allows the supply of oxygen to the tumour tissue for some hours after the end of PDT, which should increase the efficiency of cell damage and tumour response to the phototreatment. 1 J. T. Dougherty (ed.), Photodynamic therapy: mechanisms,SPZE Proc., (1989) 1065. 2 D. Kessel (ed.), Photodynamic Therapy of Neoplastic Disease, Vols. I and II, CRC Press, Boca Raton, FL, 1990.