Benzene is a known leukemia-inducing agents in humans and multi-site carcinogen in rodents. It is present in tobacco smoker and also a major environmental pollutant. About 60% of smoking-related acute myeloid leukemia mortality is attributed to benzene. The molecular mechanisms responsible for benzene-induced toxicity and carcinogenicity are not well defined; however, metabolism is a pre-requisite for its biological effects. The cytochrome P4502E1 activation of benzene is considered the dominant pathway in productive active metabolites. The CYP2E1 pathways alone is insufficient to explain the mechanism of benzene toxicity. Thus, other cellular processes participate in the biological activation of benzene. There is mounting evidence to suggest that bone marrow depression is associated with increased production of reactive oxygen species and nitric oxide (.NO) by bone marrow. On the basis of our preliminary studies we hypothesize that (a) superoxide generated in redox cycling of ring-hydroxylated metabolites reacts with . NO (induced by benzene metabolites) to form peroxynitrite. Peroxynitrite can damage DNA directly, or modify benzene and its primarily metabolites non- enzymatically to more toxic hydroxylated and nitrated products. Therefore, peroxynitrite, may in part, be responsible for the toxicity of benzene. (b) Further activation of primary may also contribute to benzene toxicity. These hypotheses will be tested by formulating the following Specific Aims. (1) Demonstrate that peroxynitrite is the primary causative agent in DNA strand breakage induced by the synergistic interaction of ring-hydroxylated benzene metabolites with an NO-donor compound in plasmid DNA in vitro. (2) Investigate catalytic role of myeloperoxidase in generating reactive benzene metabolites and study the involvement of these metabolites in DNA damage and cell toxicity in vitro. 3 (I) Extend information obtained from Aims 1 and 2 to in vivo B6C3F1 mice. Determine dose-dependent effects of benzene and its metabolites in formation and persistence of oxidized DNA bases and co-valent DNA adducts as well as production of 3-nitro tyrosine and etheno-base adducts and their relationships with cytotoxicity. 3 (II) Investigate whether C57BI/6AiTAC (KO)Nos2 mice (deficient in iNOS) are protected against benzene induced bone marrow suppression. The results of this study will provide detailed information on the mechanisms and the nature of the DNA lesions that are responsible for benzene toxicity; such knowledge is of paramount importance for risk assessment.
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