Medical Hypothesfs (1990) 32.1%123 CT,Innmman Grnun IIK Ltd 19W
Reactive Oxygen Production Against Malaria Potential Cancer Risk Factor M. 0.
EZE,*+, D. J. HUNTING*
A
and A. U. OGANt
*MRC Group in the Radiation Sciences, QuBbec, Canada J7H $N4. fDepartment (Correspondence to MOE)
Facult6 de medecine, UniversiM de Sherbrooke, of Biochemistry, University of Nigeria, Nsukka,
Sherbrooke, Nigeria
Abstract - In response to malaria infection, phagocytes, such as macro-phages and neutrophils, produce superoxide and thence the other reactive oxygen species (ROS) with which to kill the parasites. Excess ROS is normally eliminated by the body’s natural scavenger molecules; however, in the event of a vast excess of ROS, as may be the case in acute as well as chronic malaria patients, the natural scavengers may be overwhelmed. We hypothesize that unscavenged ROS in malaria patients causes DNA damage in normal host cells which, if unrepaired or incorrectly repaired, could result in oncogene activation and eventually lead to cancer. An epidemiologic study may be warranted in malaria-endemic regions to investigate the possible relationship between malaria infection and cancer risk.
Introduction It is evident that a number of common human cancers are caused by exposure to agents in our environment (1, 2). These agents include chemicals and radiation (2). At present, the most credible hypothesis to account for the environmental etiology of many cancers is based on the fact that most, if not all, carcinogenic agents ultimately result in DNA damage (3). If this damage occurs in certain oncogenes, incorrect repair or lack of repair can result in altered gene
Date Date
received received
29 May 1989 1X August 1989
products or levels of gene expression, cell transformation and, ultimately, cancer (2-4). An illustration of the apparent relationship between DNA damage/repair and cancer is given by the human hereditary disease, Xeroderma Pigmentosum (2, 5). Ultraviolet radiation causes skin cancers in subjects suffering from this condition presumably because their cells are unable to repair the DNA damage caused by the radiation. Oxygen-dependent phagocytes
microbicidal action of
In response to malaria and other infections, phagocytic cells such as polymorphonuclear leucocytes (PMNL, neutrophils) (6, 7), as well as macrophages (6, 8-12), engage in the 121
122
MEDICAL HYPOTHESES
‘Respiratory Burst’ as a host cell-mediated immune (CMI) reaction. Superoxide (02), the first product of this respiratory burst, is formed by a one-electron reduction of molecular oxygen, catalyzed by the phagocyte’s membrane NADPH oxidase. The superoxide free radical further reacts to yield the other reactive oxygen species (ROS) (hydrogen peroxide, hydroxyl radical and hypochlorite) which function in the microbicidal event (6, 7, 13). Natural defenses against the toxicity of reactive oxygen species: scavengers
Cells contain several enzymes (superoxide dismutase, catalase, glutathione reductase/ glutathione peroxidase) and other molecules (antioxidants like glutathione, ascorbate and a-tocopherol) that scavenge reactive oxygen species (3, 6, 13); however, in the event of excessively high ROS production, these natural measures may be inadequate. There is evidence that excess ROS can lead to inflammation at the point of production (in the same manner as in the joints of rheumatoid arthritis patients (13)), and probably contribute to the immunopathology of the malarial infection (9, 12). The implication of this is that there is likely to be an optimal level of production of these reactive oxygen species which is compatible with the normal microbicidal function of the phagocytes without causing deleterous side effects on the host. Chemicals/drugs
and reactive oxygen productiot‘
Certain drugs like the trypanocidal /3-lapachone (14), and the antimalarial, primaquine (15,16), as well as the antitumor glycopeptide, bleomycin (17), and a host of others, bring about their curative effects by leading to ROS production against their respective target pathogens. Some of the active principles in the medicinal herbs used by herbalists in the developing countries in the preparation of traditional drug concoctions, including antimalarial remedies widely taken in the malaria-endemic regions, may also produce ROS in the course of manifesting their curative effects. These reactive oxygen species could be produced either directly, or by triggering phagocytes. A very important example of a ROS-yielding plant extract is phorbolmyristate acetate (PMA), one of the phorbol esters extracted from the seed oil of Croton tiglium L. (18). PMA has a high potency for triggering the respiratory burst in phagocytes, thus leading to the production of large amounts of OZ. By virtue of this, phorbol
esters are tumor DNA damage.
promoters,
Mutagenic and carcinogenic ROS
probably
causing
potentials of excess
There is evidence from in vitro experiments which supports the hypothesis that ROS generated by phagocytes may play a role in the genesis of cancer. For example, activated phagocytes can generate DNA damage and mutations in bacteria (6, 19), and in cultured mammalian cells (6, 20) and activated human neutrophils can cause malignant transformation of mouse lOT1/2 cells in culture. These effects can be blocked by scavengers of activated oxygen species, thus implicating ROS (3, 4). Although there is no direct in vivo evidence for the role of activated phagocytes in carcinogenesis, the well known association of chronic inflammation and cancer (6) is consistent with this possibility. Consequentiy, ROS produced in great excess in the host would confound the scavenging capacity of the natural scavengers in the. cell or tissue in question. The unscavenged oxygen species then, in addition to causing the immunopathology of malaria (9, 12), would also cause DNA damage which if unrepaired, or if incorrectly repaired, could ‘lead to cancer subsequently as discussed above. The cause of ROS production could be the malaria infection or the therapeutic agents consumed. Conclusion Phagocytes abound in the peripheral blood, the spleen and the liver, and the cells of these tissues are active in the fight against the various stages of the malaria parasite. In the course of this fight, these cells engage in the respiratory burst producing ROS, some of which may escape the natural scavengers. We hypothesize, therefore, that there may be a relationship between the endemicity of malaria and the incidence of certain cancers, such as leukemia, and human liver and spleen cancers in the malaria-endemic regions, due to the possibility of unrepaired (or incorrectly repaired) ROS-mediated DNA damage. Indeed, a recent survey (21) has revealed that certain lung diseases like tuberculosis are strong risk factors for lung cancer. ROS produced by lung phagocytes in reaction to the primary lung disease are likely to be important here. In the same survey, increasing dietary intake of pcarotene decreased the cancer risk. This effect of
REACTIVE
OXYGEN
PRODUCTION
AGAINST
MALARIA
@-carotene suggests an advantageous role for its oxygen radical scavenging potential. Given that an estimated 100 million people are afflicted with malaria yearly (22, 23), the carcinogenic potential of malaria merits further investigation. The hypothesis that malarial infections and/or treatments increase the incidence of could be tested in an cancers certain epidemiologic study. Mechanistic studies would be necessary to determine the role of ROS, if any, in cancer induction. Acknowledgements This work was supported by a research grant from the Medical Research Council of Canada to DJH. in whose laboratory MOE is a visiting scientist.
References I. Cairns, J. Mutation selection and the natural history of cancer. Nature 255: 197. 1975. 2. Cleaver, J. E. DNA repair processes and their impairment in some human diseases. P 29 in Progress in Genetic Toxicology. (D. Scott, B. A. bridges ani F. H. Sobels, eds) Elsevier/North-Holland Biomedical Press, New York, 1977. 3. Cerutti, P. A. Prooxidant states and tumor promotion. Science 227: 375, 1985. 4. Weitzman, S. A., Wietzberg, A. B., Clark, E. P., Stossell, T. P. Phagocytes as carcinogens: Malignant transformation produced by human neutrophils, Science 227: 1231, 1985. 5. Illingworth. J. E. Internal ultraviolet luminescence - Internal carcinomas. Medical Hypothesis 27: 313, 1988. 6. Babior. B. M. Oxidants from phagocytes: Agents of defense and destruction. Blood 64: 959, 1984. 7. Forman, H. J., Thomas, M. J. Oxidant production and bactericidal activity of phagocytes. Ann. Rev. Physiol. 48: 669, 1986. 8. Allison, A. C., Eugui, E. M. The role of cell-mediated immune responses in resistance to malaria, with special reference to oxidant stress. Ann. Rev. Immunol. 1: 361, 1983. 9. Clark, I. A.. Hunt. N. M. Evidence for reactive oxygen intermediates causing hemolysis and parasite death in malaria. Infect. Immun. 39: 1. 1983.
123 10. Taverne. J., Rahman, D., Dockrell, H. M.. Alavi, A., Leveton, C., Playfair. J. H. L. Activation of liver macrophages in murine malaria is enhanced by vaccination. Clin. Exotl. Immunol. 70: 508. 1987. 11. Eze. M.‘O. Requirement for lipopolysaccharide for enhanced 6~ vitro superoxide producing competence in macrophages from normal and malaria (Plasmodium chabaudi) infected mouse spleen. Free Radical Res. Comms. In Press 1989. 12. Kharazmi, A., Jepsen, S., Anderson, B. J. Generation of reactive oxygen radicals by human phagocytic cells activated by Plasmodium falciparum. Stand. J. Immunol. 25: 335, 1987. 13. Halliwell, B. Oxidants and human disease: Some new concepts. FASEB J. 1: 358. 1987. 14. Boveris, A., Docampo, R., Turrens, J. F., Stoppani, A. 0. M. Effects of P-lapachome on superoxide anion and hydrogen peroxide production in Trypanosoma cruzi. Biochem. J. 175: 431, 1978. 15. Summerfield. M., Tudhope, G. R. Studies with primaquine in vitro: Superoxide radical formation and oxidation of hemoglobin. Br. J. Clin. Pharmacol. 6: 319, 1978. 16. Kelman. S. N., Sullivan, S. G., Stern, A. Primaquinemediated oxidative metabolism in the human red cell: Lack of dependence on oxyhemoglobin, H& formation, or glutathione turnover. Biochem. Pharmacol. 31: 2409, 1982. 17 Goldstein, S.. Czapski, G. Mechanism of reduction of bleomycin-Cu(I1) by Oz and oxidation of bleomycin-Cu(I) by H20Z in the absence and presence of DNA. Int. J. Radiat. Biol. 51: 693, 1987. 18. Merck Index. 9th ed. Phorbol. Item. No. 7143, 1976. 19. Aronovich, J.. Samuni. A., Godinger. D., Czapski, G. In vivo degradation of bacterial DNA by HzO1 and o-phenanthroline. p 346 in Superoxide and Superoxide Dismutase .in Chemistry Biology and Medicine (G. Rotilio, ed) Elsevier Science Publishers B.Y. (Biomedical Division), New York, 1986. 20. Weitzmann, S. A., Lee, R. M., Ouellette. A. J. Alterations in c-abl gene methylation in cells transformed by phagocyte-generated oxidants. Biochem. Biophys. Res. Comms. 158: 24. 1989. 21. Wu, A. H., Yu, M. C., Thomas, D. C., Pike, M. C., Henderson. B. E. Personal and family history of lung disease as risk factors for adenocarcinoma of the lung. Cancer Res. 48: 7279. 1988. 22. Wyler. D. 1. Malaria: host-pathogen biology. Rev. Infect. Diseases 4: 785. 1982. 23. Wyler, D. J. Malaria: resurgence, resistance and research. New Engl. J. Med. 308: 875 and 934, 1983.
Reactive Oxygen Production Against Malaria Potential Cancer Risk Factor M. 0.
EZE,*+, D. J. HUNTING*
A
and A. U. OGANt
*MRC Group in the Radiation Sciences, QuBbec, Canada J7H $N4. fDepartment (Correspondence to MOE)
Facult6 de medecine, UniversiM de Sherbrooke, of Biochemistry, University of Nigeria, Nsukka,
Sherbrooke, Nigeria
Abstract - In response to malaria infection, phagocytes, such as macro-phages and neutrophils, produce superoxide and thence the other reactive oxygen species (ROS) with which to kill the parasites. Excess ROS is normally eliminated by the body’s natural scavenger molecules; however, in the event of a vast excess of ROS, as may be the case in acute as well as chronic malaria patients, the natural scavengers may be overwhelmed. We hypothesize that unscavenged ROS in malaria patients causes DNA damage in normal host cells which, if unrepaired or incorrectly repaired, could result in oncogene activation and eventually lead to cancer. An epidemiologic study may be warranted in malaria-endemic regions to investigate the possible relationship between malaria infection and cancer risk.
Introduction It is evident that a number of common human cancers are caused by exposure to agents in our environment (1, 2). These agents include chemicals and radiation (2). At present, the most credible hypothesis to account for the environmental etiology of many cancers is based on the fact that most, if not all, carcinogenic agents ultimately result in DNA damage (3). If this damage occurs in certain oncogenes, incorrect repair or lack of repair can result in altered gene
Date Date
received received
29 May 1989 1X August 1989
products or levels of gene expression, cell transformation and, ultimately, cancer (2-4). An illustration of the apparent relationship between DNA damage/repair and cancer is given by the human hereditary disease, Xeroderma Pigmentosum (2, 5). Ultraviolet radiation causes skin cancers in subjects suffering from this condition presumably because their cells are unable to repair the DNA damage caused by the radiation. Oxygen-dependent phagocytes
microbicidal action of
In response to malaria and other infections, phagocytic cells such as polymorphonuclear leucocytes (PMNL, neutrophils) (6, 7), as well as macrophages (6, 8-12), engage in the 121
122
MEDICAL HYPOTHESES
‘Respiratory Burst’ as a host cell-mediated immune (CMI) reaction. Superoxide (02), the first product of this respiratory burst, is formed by a one-electron reduction of molecular oxygen, catalyzed by the phagocyte’s membrane NADPH oxidase. The superoxide free radical further reacts to yield the other reactive oxygen species (ROS) (hydrogen peroxide, hydroxyl radical and hypochlorite) which function in the microbicidal event (6, 7, 13). Natural defenses against the toxicity of reactive oxygen species: scavengers
Cells contain several enzymes (superoxide dismutase, catalase, glutathione reductase/ glutathione peroxidase) and other molecules (antioxidants like glutathione, ascorbate and a-tocopherol) that scavenge reactive oxygen species (3, 6, 13); however, in the event of excessively high ROS production, these natural measures may be inadequate. There is evidence that excess ROS can lead to inflammation at the point of production (in the same manner as in the joints of rheumatoid arthritis patients (13)), and probably contribute to the immunopathology of the malarial infection (9, 12). The implication of this is that there is likely to be an optimal level of production of these reactive oxygen species which is compatible with the normal microbicidal function of the phagocytes without causing deleterous side effects on the host. Chemicals/drugs
and reactive oxygen productiot‘
Certain drugs like the trypanocidal /3-lapachone (14), and the antimalarial, primaquine (15,16), as well as the antitumor glycopeptide, bleomycin (17), and a host of others, bring about their curative effects by leading to ROS production against their respective target pathogens. Some of the active principles in the medicinal herbs used by herbalists in the developing countries in the preparation of traditional drug concoctions, including antimalarial remedies widely taken in the malaria-endemic regions, may also produce ROS in the course of manifesting their curative effects. These reactive oxygen species could be produced either directly, or by triggering phagocytes. A very important example of a ROS-yielding plant extract is phorbolmyristate acetate (PMA), one of the phorbol esters extracted from the seed oil of Croton tiglium L. (18). PMA has a high potency for triggering the respiratory burst in phagocytes, thus leading to the production of large amounts of OZ. By virtue of this, phorbol
esters are tumor DNA damage.
promoters,
Mutagenic and carcinogenic ROS
probably
causing
potentials of excess
There is evidence from in vitro experiments which supports the hypothesis that ROS generated by phagocytes may play a role in the genesis of cancer. For example, activated phagocytes can generate DNA damage and mutations in bacteria (6, 19), and in cultured mammalian cells (6, 20) and activated human neutrophils can cause malignant transformation of mouse lOT1/2 cells in culture. These effects can be blocked by scavengers of activated oxygen species, thus implicating ROS (3, 4). Although there is no direct in vivo evidence for the role of activated phagocytes in carcinogenesis, the well known association of chronic inflammation and cancer (6) is consistent with this possibility. Consequentiy, ROS produced in great excess in the host would confound the scavenging capacity of the natural scavengers in the. cell or tissue in question. The unscavenged oxygen species then, in addition to causing the immunopathology of malaria (9, 12), would also cause DNA damage which if unrepaired, or if incorrectly repaired, could ‘lead to cancer subsequently as discussed above. The cause of ROS production could be the malaria infection or the therapeutic agents consumed. Conclusion Phagocytes abound in the peripheral blood, the spleen and the liver, and the cells of these tissues are active in the fight against the various stages of the malaria parasite. In the course of this fight, these cells engage in the respiratory burst producing ROS, some of which may escape the natural scavengers. We hypothesize, therefore, that there may be a relationship between the endemicity of malaria and the incidence of certain cancers, such as leukemia, and human liver and spleen cancers in the malaria-endemic regions, due to the possibility of unrepaired (or incorrectly repaired) ROS-mediated DNA damage. Indeed, a recent survey (21) has revealed that certain lung diseases like tuberculosis are strong risk factors for lung cancer. ROS produced by lung phagocytes in reaction to the primary lung disease are likely to be important here. In the same survey, increasing dietary intake of pcarotene decreased the cancer risk. This effect of
REACTIVE
OXYGEN
PRODUCTION
AGAINST
MALARIA
@-carotene suggests an advantageous role for its oxygen radical scavenging potential. Given that an estimated 100 million people are afflicted with malaria yearly (22, 23), the carcinogenic potential of malaria merits further investigation. The hypothesis that malarial infections and/or treatments increase the incidence of could be tested in an cancers certain epidemiologic study. Mechanistic studies would be necessary to determine the role of ROS, if any, in cancer induction. Acknowledgements This work was supported by a research grant from the Medical Research Council of Canada to DJH. in whose laboratory MOE is a visiting scientist.
References I. Cairns, J. Mutation selection and the natural history of cancer. Nature 255: 197. 1975. 2. Cleaver, J. E. DNA repair processes and their impairment in some human diseases. P 29 in Progress in Genetic Toxicology. (D. Scott, B. A. bridges ani F. H. Sobels, eds) Elsevier/North-Holland Biomedical Press, New York, 1977. 3. Cerutti, P. A. Prooxidant states and tumor promotion. Science 227: 375, 1985. 4. Weitzman, S. A., Wietzberg, A. B., Clark, E. P., Stossell, T. P. Phagocytes as carcinogens: Malignant transformation produced by human neutrophils, Science 227: 1231, 1985. 5. Illingworth. J. E. Internal ultraviolet luminescence - Internal carcinomas. Medical Hypothesis 27: 313, 1988. 6. Babior. B. M. Oxidants from phagocytes: Agents of defense and destruction. Blood 64: 959, 1984. 7. Forman, H. J., Thomas, M. J. Oxidant production and bactericidal activity of phagocytes. Ann. Rev. Physiol. 48: 669, 1986. 8. Allison, A. C., Eugui, E. M. The role of cell-mediated immune responses in resistance to malaria, with special reference to oxidant stress. Ann. Rev. Immunol. 1: 361, 1983. 9. Clark, I. A.. Hunt. N. M. Evidence for reactive oxygen intermediates causing hemolysis and parasite death in malaria. Infect. Immun. 39: 1. 1983.
123 10. Taverne. J., Rahman, D., Dockrell, H. M.. Alavi, A., Leveton, C., Playfair. J. H. L. Activation of liver macrophages in murine malaria is enhanced by vaccination. Clin. Exotl. Immunol. 70: 508. 1987. 11. Eze. M.‘O. Requirement for lipopolysaccharide for enhanced 6~ vitro superoxide producing competence in macrophages from normal and malaria (Plasmodium chabaudi) infected mouse spleen. Free Radical Res. Comms. In Press 1989. 12. Kharazmi, A., Jepsen, S., Anderson, B. J. Generation of reactive oxygen radicals by human phagocytic cells activated by Plasmodium falciparum. Stand. J. Immunol. 25: 335, 1987. 13. Halliwell, B. Oxidants and human disease: Some new concepts. FASEB J. 1: 358. 1987. 14. Boveris, A., Docampo, R., Turrens, J. F., Stoppani, A. 0. M. Effects of P-lapachome on superoxide anion and hydrogen peroxide production in Trypanosoma cruzi. Biochem. J. 175: 431, 1978. 15. Summerfield. M., Tudhope, G. R. Studies with primaquine in vitro: Superoxide radical formation and oxidation of hemoglobin. Br. J. Clin. Pharmacol. 6: 319, 1978. 16. Kelman. S. N., Sullivan, S. G., Stern, A. Primaquinemediated oxidative metabolism in the human red cell: Lack of dependence on oxyhemoglobin, H& formation, or glutathione turnover. Biochem. Pharmacol. 31: 2409, 1982. 17 Goldstein, S.. Czapski, G. Mechanism of reduction of bleomycin-Cu(I1) by Oz and oxidation of bleomycin-Cu(I) by H20Z in the absence and presence of DNA. Int. J. Radiat. Biol. 51: 693, 1987. 18. Merck Index. 9th ed. Phorbol. Item. No. 7143, 1976. 19. Aronovich, J.. Samuni. A., Godinger. D., Czapski, G. In vivo degradation of bacterial DNA by HzO1 and o-phenanthroline. p 346 in Superoxide and Superoxide Dismutase .in Chemistry Biology and Medicine (G. Rotilio, ed) Elsevier Science Publishers B.Y. (Biomedical Division), New York, 1986. 20. Weitzmann, S. A., Lee, R. M., Ouellette. A. J. Alterations in c-abl gene methylation in cells transformed by phagocyte-generated oxidants. Biochem. Biophys. Res. Comms. 158: 24. 1989. 21. Wu, A. H., Yu, M. C., Thomas, D. C., Pike, M. C., Henderson. B. E. Personal and family history of lung disease as risk factors for adenocarcinoma of the lung. Cancer Res. 48: 7279. 1988. 22. Wyler. D. 1. Malaria: host-pathogen biology. Rev. Infect. Diseases 4: 785. 1982. 23. Wyler, D. J. Malaria: resurgence, resistance and research. New Engl. J. Med. 308: 875 and 934, 1983.
