Polycyclic aromatic hydrocarbons (PAHs) are a widespread class of environmental pollutants many of which exhibit high carcinogenic potency. It is now generally believed that PAHs undergo metabolic activation to reactive diolepoxide intermediates which bind covalently to nucleic acids. The long range objective of our research program is to elucidate the mechanism of carcinogenesis of PAHs at the molecular-genetic level.
The specific aims of this proposal are: (1) to devise methods for the efficient syntheses on preparative scale of adducts of carcinogenic diol epoxide metabolites of PAHs with nucleosides and nucleosides covalently bound at specific base sites; (2) to incorporate these adducts into specific oligonucleotide sequences, both single and double stranded; and (3) to determine the complete structures of these PAH oligonucleotide adducts by NMR, UV and linear dichroism spectroscopy, and X-ray crystal analysis. These investigations will provide convenient access to the pure adducts of PAH diol epoxides covalently bound to predetermined base sites of nucleosides and nucleotides in gram scale, a significant advance over direct alkylation which generally provides milligram amounts of mixtures of adducts and is unsuitable for the preparation of minor adducts. The availability of adducts of precisely defined molecular structures will make possible the synthesis of oligonucleotide sequences containing PAH residues covalently bound at specific base sites in sufficient quantities for determination of their molecular structures by physical methods. The information obtained will provide direct experimental evidence on the structures of such adducts which may be expected to settle current controversies on this topic and provide insight into the question of whether carcinogenic PAH diol epoxides afford adducts of uniquely distinctive structure. The techniques developed will allow custom synthesis of specific PAH-alkylated oligomers which model the sites of preferential binding of PAHs to DNA as determined by """"""""footprinting"""""""" techniques, as well as the synthesis of PAH-alkylated oligomers in which the base sequences are varied to determine the effects of nearest neighbors on structure and function. The PAH oligonucleotide adducts will be incorporated into DNA and RNA for biological studies to determine the effects of adduct structure on replication and repair. The information derived from these studies will be utilized to relate PAH-nucleic acid adduct structure to the molecular biological consequences of adduct formation. The synthetic PAH nucleoside conjugates will also be furnished to other investigators as authentic standards for the identification of the major and minor adducts formed with DNA by the metabolism of hydrocarbons in vivo.
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