Comparison of the genotoxic activities of the K-region dihydrodiol of benzo[a]pyrene with benzo[a]pyrene in mammalian cells: morphological cell transformation; DNA damage; and stable covalent DNA adducts.

Abstract Benzo[ a ]pyrene (B[ a ]P) is the most thoroughly studied polycyclic aromatic hydrocarbon (PAH). Many mechanisms have been suggested to explain its carcinogenic activity, yet many questions still remain. K-region dihydrodiols of PAHs are metabolic intermediates depending on the specific cytochrome P450 and had been thought to be detoxification products. However, K-region dihydrodiols of several PAHs have recently been shown to morphologically transform mouse embryo C3H10T1/2CL8 cells (C3H10T1/2 cells). Because K-region dihydrodiols are not metabolically formed from PAHs by C3H10T1/2 cells, these cells provide a useful tool to independently study the mechanisms of action of PAHs and their K-region dihydrodiols. Here, we compare the morphological cell transforming, DNA damaging, and DNA adducting activities of the K-region dihydrodiol of B[ a ]P, trans- B[ a ]P-4,5-diol with B[ a ]P. Both trans -B[ a ]P-4,5-diol and B[ a ]P morphologically transformed C3H10T1/2 cells by producing both Types II and III transformed foci. The morphological cell transforming and cytotoxicity dose response curves for trans -B[ a ]P-4,5-diol and B[ a ]P were indistinguishable. Since morphological cell transformation is strongly associated with mutation and/or larger scale DNA damage in C3H10T1/2 cells, the identification of DNA damage induced in these cells by trans -B[ a ]P-4,5-diol was sought. Both trans -B[ a ]P-4,5-diol and B[ a ]P exhibited significant DNA damaging activity without significant concurrent cytotoxicity using the comet assay, but with different dose responses and comet tail distributions. DNA adduct patterns from C3H10T1/2 cells were examined after trans -B[ a ]P-4,5-diol or B[ a ]P treatment using 32 P -postlabeling techniques and improved TLC elution systems designed to separate polar DNA adducts. While B[ a ]P treatment produced one major DNA adduct identified as anti-trans -B[ a ]P-7,8-diol-9,10-epoxide-deoxyguanosine, no stable covalent DNA adducts were detected in the DNA of trans -B[ a ]P-4,5-diol-treated cells. In summary, this study provides evidence for the DNA damaging and morphological cell transforming activities of the K-region dihydrodiol of B[ a ]P, in the absence of covalent stable DNA adducts. While trans -B[ a ]P-4,5-diol and B[ a ]P both induce morphological cell transformation, their activities as DNA damaging agents differ, both qualitatively and quantitatively. In concert with the morphological cell transformation activities of other K-region dihydrodiols of PAHs, these data suggest a new mechanism/pathway for the morphological cell transforming activities of B[ a ]P and its metabolites.