- Email: [email protected]
The cationic meso-substituted porphyrins: An interesting group of photosensitizers
295
J. Photochem. Photobiol. B: Biol,, 18 (1993) 295-298
News and Views The cationic meso-substituted porphyrins: an interesting group photosensitizers
of
the nucleus was identified as the subcellular site of localization of T,MPyP [9]. Previous studies conducted in our laboratory also demonstrated the same cellular localization in an in viva system
WI.
A. Villanueva Departamento de Biologia, Universidad Autdnoma de Madrid, 28049 Madrid (Spain)
The growing interest in the cationic porphyrin meso-tetra(4Wmethylpyridyl)porphine (T,MPyP) began when it was described as a photosensitizing agent by Diamond et al. [l] in 1977. More recently, Fiel et al. [2] have reported the unexpected capacity of this porphyrin to bind to DNA by intercalation, showing some degree of specificity at GC base pair sites (G, guanine; C, cytosine) [3, 41. The affinity of this and other cationic porphyrins, including some metal derivatives, for DNA has been analysed in recent years. Pasternack et al. [5] have presented evidence to indicate that the nature of the interactions with nucleic acids depends on the occurrence of axial ligand(s) with the metal. Porphyrins with one or two axial ligands (e.g. Fe(III)-, Co(III)- or Zn-T,MPyP) are unable to intercalate between the base pairs of DNA. They bind on the outside of the DNA helix, mainly to the phosphate groups. These workers have also proposed that these axial ligand derivatives may be used as “molecular callipers” for limiting the thickness of an intercalating drug. Thus a knowledge of the interactions of cationic porphyrins with nucleic acids could be useful in understanding the structure of nucleic acids at several organization levels. A detailed description of these approaches and results has been reported by Fiel [6]. T,MPyP also possesses other properties, which make it particularly attractive for the investigation of photobiological processes. It exhibits high quantum yields for formation of singlet oxygen (‘0,) [7], and it remains monomeric within a large concentration range [6, 81. Studies on the cellular localization and photodynamic effects of T,MPyP in cell cultures have been performed in our laboratory. This porphyrin shows good uptake and high phototoxicity in HeLa cells, the extent of cellular damage being dependent on the light dose. Using fluorescence microscopy, loll-1344/93/$6.00
Since the intracellular localization of a sensitizer may be closely related to its mechanism of damage, we have analysed the possible cellular DNA photodamage induced by T,MPyP in HeLa cells. Our results show how this porphyrin is able to induce breaks in the chromosomes of HeLa cells when they are exposed to sublethal fluences of light [9]. Previous experiments with isolated DNA indicated the occurrence of single- and double-strand breaks provoked by T,MPyP in the presence of light. This photodamage seems to be mediated mainly by ‘02, since the active oxygen species can react with DNA producing damage [ll-141. A review of the mutagenic and genotoxic properties of ‘02 was presented by Piette [15]. It is interesting to note that most photosensitizers used in photodynamic therapy (PDT), including phthalocyanines and various porphyrins (e.g. haematoporphyrin (HP) and haematoporphyrin derivative (HPD)), are not able to penetrate into the cell nucleus. However, the production of photoinduced damage in DNA has been described [16-201. Moan and Berg [21] have proposed that this DNA photodamage may be due to damage of the nuclear membrane. It is clear that a more detailed study of the genotoxic effects induced by photosensitizing agents is of fundamental importance in PDT, though DNA does not appear to be the primary cellular target for many drugs. Photogenotoxic effects such as the induction of mutations may be produced. In this case, it will be necessary to fix the lethal dose of treatment to avoid this undesirable effect. T,MPyP is an interesting drug which can be used to compare the effect of DNA-bound and unbound photosensitizers; this is important to determine the degree of ‘02 accessibility to oxidizable targets. Moan [22] has estimated the ‘0, diffusion distance in cells to be about 0.1 pm. Finally, in vitro studies of T,MPyP have provided some information on cellular effects; the extension of these experiments to model systems in vivo is needed to establish a complete picture of its photosensitizing properties. 0
1993 - Elsevier Sequoia. All rights reserved
296
News and V&TVS
1 I. Diamond, S. G. Granelli and A. F. McDonagh, Photochemotherapy and photodynamic toxicity: simple methods for identifying potentially active agents, B&hem. Med., 17 (1977) 121-127. 2 R. J. Fiel, J. C. Howard, E. H. Mark and N. Datta-Gupta, Interaction of DNA with a porphyrin ligand: evidence for intercalation, Nucleic Acids Rex, 6 (1979) 3093-3118. 3 M. J. Carvlin and R. J. Fiel, Intercalative and nonintercalative binding of large cationic porphyrin ligands to calf thymus DNA, Nucleic Acids Res., II (1983) 6121-6139. 4 D. L. Banville, L. G. Marzilli and J. A. Strickland, Comparison of the effects of cationic porphyrins on DNA properties: influence of GC content of native and synthetic polymers, Biopolymers, 25 (1986) 1837-1858. 5 R. F. Pasternack, E. J. Gibbs and J. J. Villafranca, Interactions of porphyrins with nucleic acids, B&hem&y, 22 (1983) 2406-2414. 6 R. J. Fiel, Porphyrin-nucleic acid interactions: a review, J. Biomol. Struct. Dynamics, 6 (1989) 1259-1274. 7 J. B. Verlhac, A. Gaudemer and I. Kraljic, Water-soluble porphyrins and metalloporphyrins as photosensitizers in aerated aqueous solutions. I. Detection and determination of quantum yield of formation of singlet oxygen, Nouv. J. Chim., 8 (1984) 401-406. 8 R. F. Pastemack, E. J. Gibbs, A. Gaudemer, A. Antebi, S. Bassner, L. De Poy, D. H. Turner, A. Williams, F. Laplace. M. H. Lansard, C. Merienne and M. PerrCe-Fauvet, Molecular complexes of nucleosides and nucleotides with a monomeric cationic porphyrin and some of its metal derivatives, J. Am. Chem. Sot., IO7 (1985) 8179-8186. 9 A. Villanueva, A. Juarranz, V. Diaz, J. G6mez and M. Catiete, Photodynamic effects of a cationic mesosubstituted porphyrin in cell cultures, Anti-Cancer Drug Design, 7 (1992) 297-303. 10 A. Villanueva, M. Cafiete and M. J. Hazen, Uptake and DNA photodamage induced in plant cells “in viva” by two cationic porphyrins, Mutagenesis, 4 (1989) 157-159. 11 R. 3. Fiel, N. Datta-Gupta, E. H. Mark and J. C. Howard, Induction of DNA damage by porphyrin photosensitizers, Cancer Res., 41 (1981) 3543-3545. 12 J. M. Kelly and M. J. Murphy, A comparative study of the 5,10,15,20-tetrakis(N-methylpyridinium-4interaction of yl)porphyrin and its zinc complex with DNA using fluorescence spectroscopy and topoisomerisation, Nucleic Acids Res., 13 (1985) 167-184. 13 T. Le Doan, L. Perrouault, M. Rougee, R. V. Bensasson and C. Htlbne, Photosensitized reactions on DNA by meso substituted cationic prophyrins, Photo&em. Photobiol., 43s (1986) 48s. 14 D. Praseuth, A. Gaudemer, J. B. Verlhac, I. Kraljic, I. Sissoeff and E. GuillC, Photocleavage of DNA in the presence of synthetic water-soluble porphyrins, Photochem. Photobiol., 44 (1986) 717-724. 15 J. Piette, Mutagenic and genotoxic properties of singlet oxygen, I. Pholochem. Photobiol. B: Biol., 4 (1990) 335-342. 16 J. Moan, H. Waksvik and T. Christensen, DNA single-strand breaks and sister chromatid exchanges induced by treatment with hematoporphyrin and light or by X-rays in human NHIK 3025 cells, Cancer Res., 40 (1980) 2915-2918. 17 C. J. Gomer, N. Rucker, A. Banerjee and W. F. Benedict, Comparison of mutagenicity and induction of sister chromatid exchange in Chinese hamster cells exposed to hematoporphyrin derivative photoradiation, ionizing radiation or ultraviolet radiation, Cancer Res., 43 (1983) 2622-2627.
18 E. Ben-Hur, T. Fujihara, F. Suzuki and M. M. Elkind, Genetic toxicology of the photosensitization of Chinese hamster cells by phthalocyanines, Photochem. Photobiol., 45 (1987) 227-230. 19 N. Ramakrishnan, M. E. Clay, L. Y. Xue, H. H. Evans, A. R. Antunez and N. L. Oleinic, Induction of DNA-protein cross-links in Chinese hamster cells by the photodynamic action of chloroaluminium phthalocyanine and visible light, Phorochem. Photobiol., 48 (1988) 297-303. 20 H. H. Evans, R. M. Rerko, J. Mencl, M. E. Clay, A. R. Antunez and N. L. Oleinic, Cytotoxic and mutagenic effects of the photodynamic action of chloroaluminium phthalocyanine and visible light in L5178Y cells, Photochem. Photobiol., 49 (1989) 43-48. of cancer: ex21 J. Moan and K. Berg, Photochemotherapy perimental research, Photochem. Photobiol., 55 (1992) 931-948. 22 J. Moan, On the ditision length of singlet oxygen in cells and tissues,J. Photochem. Photobiol. B: Biol., 6 (1990) 343-344.
Reflections damage Michael
on type I photodynamic
A. J. Rodgers
Center for Photochemical Sciences, Department of ChemCtry, Bowling Green State University Bowling Green, OH 43403 WW
1. Introduction Recently, views have been expressed in this journal [l] and in Photochemistry and Photobiology [2] that have drawn attention to the two possible mechanisms (types I and II) by which photodynamic damage is thought to occur. Notwithstanding the contrasting viewpoints of Foote [2] and Vidbczy [l], the basic difference between the mechanisms seems to be in the nature of the primary dark reaction of the photosensitizer excited state. This is either an electron transfer (LT) process or an energy transfer (NT) process. The energy recipient in the latter is fairly safely identified as 02, it being the only biological molecule having an electronic state (‘A,) low enough in energy to behave as an acceptor when the donors are T1 states of porphyrins, xanthenes and phthalocyanines. Coreactants in electron transfer reactions are less readily identified, and probably for this reason the focus of photodynamic research has been on singlet oxygen pathways. However, given some reasonable requisite assumptions, electron transfer targets can be identified, and useful experimental strategies can be formulated. In the following, the principal targets for LT-induced photodynamic damage are identified as the amino acids cysteine (Cys), methionine (Met), tryptophan (Trp) and tyrosine
J. Photochem. Photobiol. B: Biol,, 18 (1993) 295-298
News and Views The cationic meso-substituted porphyrins: an interesting group photosensitizers
of
the nucleus was identified as the subcellular site of localization of T,MPyP [9]. Previous studies conducted in our laboratory also demonstrated the same cellular localization in an in viva system
WI.
A. Villanueva Departamento de Biologia, Universidad Autdnoma de Madrid, 28049 Madrid (Spain)
The growing interest in the cationic porphyrin meso-tetra(4Wmethylpyridyl)porphine (T,MPyP) began when it was described as a photosensitizing agent by Diamond et al. [l] in 1977. More recently, Fiel et al. [2] have reported the unexpected capacity of this porphyrin to bind to DNA by intercalation, showing some degree of specificity at GC base pair sites (G, guanine; C, cytosine) [3, 41. The affinity of this and other cationic porphyrins, including some metal derivatives, for DNA has been analysed in recent years. Pasternack et al. [5] have presented evidence to indicate that the nature of the interactions with nucleic acids depends on the occurrence of axial ligand(s) with the metal. Porphyrins with one or two axial ligands (e.g. Fe(III)-, Co(III)- or Zn-T,MPyP) are unable to intercalate between the base pairs of DNA. They bind on the outside of the DNA helix, mainly to the phosphate groups. These workers have also proposed that these axial ligand derivatives may be used as “molecular callipers” for limiting the thickness of an intercalating drug. Thus a knowledge of the interactions of cationic porphyrins with nucleic acids could be useful in understanding the structure of nucleic acids at several organization levels. A detailed description of these approaches and results has been reported by Fiel [6]. T,MPyP also possesses other properties, which make it particularly attractive for the investigation of photobiological processes. It exhibits high quantum yields for formation of singlet oxygen (‘0,) [7], and it remains monomeric within a large concentration range [6, 81. Studies on the cellular localization and photodynamic effects of T,MPyP in cell cultures have been performed in our laboratory. This porphyrin shows good uptake and high phototoxicity in HeLa cells, the extent of cellular damage being dependent on the light dose. Using fluorescence microscopy, loll-1344/93/$6.00
Since the intracellular localization of a sensitizer may be closely related to its mechanism of damage, we have analysed the possible cellular DNA photodamage induced by T,MPyP in HeLa cells. Our results show how this porphyrin is able to induce breaks in the chromosomes of HeLa cells when they are exposed to sublethal fluences of light [9]. Previous experiments with isolated DNA indicated the occurrence of single- and double-strand breaks provoked by T,MPyP in the presence of light. This photodamage seems to be mediated mainly by ‘02, since the active oxygen species can react with DNA producing damage [ll-141. A review of the mutagenic and genotoxic properties of ‘02 was presented by Piette [15]. It is interesting to note that most photosensitizers used in photodynamic therapy (PDT), including phthalocyanines and various porphyrins (e.g. haematoporphyrin (HP) and haematoporphyrin derivative (HPD)), are not able to penetrate into the cell nucleus. However, the production of photoinduced damage in DNA has been described [16-201. Moan and Berg [21] have proposed that this DNA photodamage may be due to damage of the nuclear membrane. It is clear that a more detailed study of the genotoxic effects induced by photosensitizing agents is of fundamental importance in PDT, though DNA does not appear to be the primary cellular target for many drugs. Photogenotoxic effects such as the induction of mutations may be produced. In this case, it will be necessary to fix the lethal dose of treatment to avoid this undesirable effect. T,MPyP is an interesting drug which can be used to compare the effect of DNA-bound and unbound photosensitizers; this is important to determine the degree of ‘02 accessibility to oxidizable targets. Moan [22] has estimated the ‘0, diffusion distance in cells to be about 0.1 pm. Finally, in vitro studies of T,MPyP have provided some information on cellular effects; the extension of these experiments to model systems in vivo is needed to establish a complete picture of its photosensitizing properties. 0
1993 - Elsevier Sequoia. All rights reserved
296
News and V&TVS
1 I. Diamond, S. G. Granelli and A. F. McDonagh, Photochemotherapy and photodynamic toxicity: simple methods for identifying potentially active agents, B&hem. Med., 17 (1977) 121-127. 2 R. J. Fiel, J. C. Howard, E. H. Mark and N. Datta-Gupta, Interaction of DNA with a porphyrin ligand: evidence for intercalation, Nucleic Acids Rex, 6 (1979) 3093-3118. 3 M. J. Carvlin and R. J. Fiel, Intercalative and nonintercalative binding of large cationic porphyrin ligands to calf thymus DNA, Nucleic Acids Res., II (1983) 6121-6139. 4 D. L. Banville, L. G. Marzilli and J. A. Strickland, Comparison of the effects of cationic porphyrins on DNA properties: influence of GC content of native and synthetic polymers, Biopolymers, 25 (1986) 1837-1858. 5 R. F. Pasternack, E. J. Gibbs and J. J. Villafranca, Interactions of porphyrins with nucleic acids, B&hem&y, 22 (1983) 2406-2414. 6 R. J. Fiel, Porphyrin-nucleic acid interactions: a review, J. Biomol. Struct. Dynamics, 6 (1989) 1259-1274. 7 J. B. Verlhac, A. Gaudemer and I. Kraljic, Water-soluble porphyrins and metalloporphyrins as photosensitizers in aerated aqueous solutions. I. Detection and determination of quantum yield of formation of singlet oxygen, Nouv. J. Chim., 8 (1984) 401-406. 8 R. F. Pastemack, E. J. Gibbs, A. Gaudemer, A. Antebi, S. Bassner, L. De Poy, D. H. Turner, A. Williams, F. Laplace. M. H. Lansard, C. Merienne and M. PerrCe-Fauvet, Molecular complexes of nucleosides and nucleotides with a monomeric cationic porphyrin and some of its metal derivatives, J. Am. Chem. Sot., IO7 (1985) 8179-8186. 9 A. Villanueva, A. Juarranz, V. Diaz, J. G6mez and M. Catiete, Photodynamic effects of a cationic mesosubstituted porphyrin in cell cultures, Anti-Cancer Drug Design, 7 (1992) 297-303. 10 A. Villanueva, M. Cafiete and M. J. Hazen, Uptake and DNA photodamage induced in plant cells “in viva” by two cationic porphyrins, Mutagenesis, 4 (1989) 157-159. 11 R. 3. Fiel, N. Datta-Gupta, E. H. Mark and J. C. Howard, Induction of DNA damage by porphyrin photosensitizers, Cancer Res., 41 (1981) 3543-3545. 12 J. M. Kelly and M. J. Murphy, A comparative study of the 5,10,15,20-tetrakis(N-methylpyridinium-4interaction of yl)porphyrin and its zinc complex with DNA using fluorescence spectroscopy and topoisomerisation, Nucleic Acids Res., 13 (1985) 167-184. 13 T. Le Doan, L. Perrouault, M. Rougee, R. V. Bensasson and C. Htlbne, Photosensitized reactions on DNA by meso substituted cationic prophyrins, Photo&em. Photobiol., 43s (1986) 48s. 14 D. Praseuth, A. Gaudemer, J. B. Verlhac, I. Kraljic, I. Sissoeff and E. GuillC, Photocleavage of DNA in the presence of synthetic water-soluble porphyrins, Photochem. Photobiol., 44 (1986) 717-724. 15 J. Piette, Mutagenic and genotoxic properties of singlet oxygen, I. Pholochem. Photobiol. B: Biol., 4 (1990) 335-342. 16 J. Moan, H. Waksvik and T. Christensen, DNA single-strand breaks and sister chromatid exchanges induced by treatment with hematoporphyrin and light or by X-rays in human NHIK 3025 cells, Cancer Res., 40 (1980) 2915-2918. 17 C. J. Gomer, N. Rucker, A. Banerjee and W. F. Benedict, Comparison of mutagenicity and induction of sister chromatid exchange in Chinese hamster cells exposed to hematoporphyrin derivative photoradiation, ionizing radiation or ultraviolet radiation, Cancer Res., 43 (1983) 2622-2627.
18 E. Ben-Hur, T. Fujihara, F. Suzuki and M. M. Elkind, Genetic toxicology of the photosensitization of Chinese hamster cells by phthalocyanines, Photochem. Photobiol., 45 (1987) 227-230. 19 N. Ramakrishnan, M. E. Clay, L. Y. Xue, H. H. Evans, A. R. Antunez and N. L. Oleinic, Induction of DNA-protein cross-links in Chinese hamster cells by the photodynamic action of chloroaluminium phthalocyanine and visible light, Phorochem. Photobiol., 48 (1988) 297-303. 20 H. H. Evans, R. M. Rerko, J. Mencl, M. E. Clay, A. R. Antunez and N. L. Oleinic, Cytotoxic and mutagenic effects of the photodynamic action of chloroaluminium phthalocyanine and visible light in L5178Y cells, Photochem. Photobiol., 49 (1989) 43-48. of cancer: ex21 J. Moan and K. Berg, Photochemotherapy perimental research, Photochem. Photobiol., 55 (1992) 931-948. 22 J. Moan, On the ditision length of singlet oxygen in cells and tissues,J. Photochem. Photobiol. B: Biol., 6 (1990) 343-344.
Reflections damage Michael
on type I photodynamic
A. J. Rodgers
Center for Photochemical Sciences, Department of ChemCtry, Bowling Green State University Bowling Green, OH 43403 WW
1. Introduction Recently, views have been expressed in this journal [l] and in Photochemistry and Photobiology [2] that have drawn attention to the two possible mechanisms (types I and II) by which photodynamic damage is thought to occur. Notwithstanding the contrasting viewpoints of Foote [2] and Vidbczy [l], the basic difference between the mechanisms seems to be in the nature of the primary dark reaction of the photosensitizer excited state. This is either an electron transfer (LT) process or an energy transfer (NT) process. The energy recipient in the latter is fairly safely identified as 02, it being the only biological molecule having an electronic state (‘A,) low enough in energy to behave as an acceptor when the donors are T1 states of porphyrins, xanthenes and phthalocyanines. Coreactants in electron transfer reactions are less readily identified, and probably for this reason the focus of photodynamic research has been on singlet oxygen pathways. However, given some reasonable requisite assumptions, electron transfer targets can be identified, and useful experimental strategies can be formulated. In the following, the principal targets for LT-induced photodynamic damage are identified as the amino acids cysteine (Cys), methionine (Met), tryptophan (Trp) and tyrosine