title: 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 32P-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.
authors.affiliation: Environmental Carcinogenesis Division, National Health and Environmental Effects Research Laboratory, Office of Research and Development, US Environmental Protection Agency, MD-68, Research Triangle Park, NC 27711, USA. nesnow.stephen@epa.gov
authors.forename: Stephen
authors.initials: S
authors.lastname: Nesnow
authors.forename: Christine
authors.initials: C
authors.lastname: Davis
authors.forename: Garret B
authors.initials: GB
authors.lastname: Nelson
authors.forename: Guy
authors.initials: G
authors.lastname: Lambert
authors.forename: William
authors.initials: W
authors.lastname: Padgett
authors.forename: Maria
authors.initials: M
authors.lastname: Pimentel
authors.forename: Alan H
authors.initials: AH
authors.lastname: Tennant
authors.forename: Andrew D
authors.initials: AD
authors.lastname: Kligerman
authors.forename: Jeffrey A
authors.initials: JA
authors.lastname: Ross
mesh_terms: D000818:Animals; D001564:Benzo(a)pyrene / Q000737:chemistry / Q000633:toxicity*; D002471:Cell Transformation, Neoplastic / Q000187:drug effects*; D002478:Cells, Cultured; D020552:Comet Assay; D018736:DNA Adducts / Q000737:chemistry / Q000187:drug effects*; D004249:DNA Damage / Q000187:drug effects* / Q000235:genetics; D004101:Dihydroxydihydrobenzopyrenes / Q000737:chemistry* / Q000633:toxicity*; D004622:Embryo, Mammalian / Q000166:cytology / Q000187:drug effects; D005347:Fibroblasts / Q000187:drug effects / Q000473:pathology; D007553:Isotope Labeling; D008322:Mammals; D051379:Mice; D009152:Mutagenicity Tests / Q000379:methods; D010761:Phosphorus Radioisotopes
chemical_list: D018736:DNA Adducts; D004101:Dihydroxydihydrobenzopyrenes; D010761:Phosphorus Radioisotopes; C015813:4,5-dihydroxy-4,5-dihydrobenzo(a)pyrene; D001564:Benzo(a)pyrene