DNA adduct formation, metabolism, and morphological transforming activity of aceanthrylene in C3H10T1/2CL8 cells

Abstract

Aceanthrylene (ACE), a cyclopenta-fused polycyclic aromatic hydrocarbon (CP-PAH) related to anthracene, has been studied for its ability to be metabolized, to form DNA adducts, and to morphologically transform C3H10T1/2CL8 mouse embryo fibroblasts in culture. Although ACE has been previously shown to be a strong mutagen in Salmonella typhimurium strains TA89 and TA100, it did not transform C3H10T1/2 cells (0.4–16 μg/ml) under 2 treatment protocols: treatment (for 24 h) 1 day after seeding the cells; treatment (for 24 h) 5 days after seeding the cells. Both protocols are effective in detecting the morphological transforming activity of PAH and CP-PAH and the latter protocol has been shown to be effective in detecting chemicals which are active in the first protocol only with the additional treatment of the cells with a tumor promoter. ACE is metabolized by C3H10T1/2 cells to ACE-1,2-dihydrodiol (the cyclopenta-ring dihydrodiol) at a rate of 450 pmoles ACE-1,2-dihydrodiol formed/h/10 6 cells. ACE-7,8-dihydrodiol and ACE-9,10-dihydrodiol, identified as major Aroclor-1254-induced rat liver microsomal metabolites from their UV, NMR, and mass spectral data, were not identified in incubations of C3H10T1/2 cells with ACE. ACE-DNA adducts in C3H10T1/2 cells were isolated, separated, identified, and quantitated using the 32P-postlabeling method. ACE forms 4 major adducts and each was identified as an ACE-1,2-oxide/2′-deoxyguanosine adduct. The level of adduction was 2.18 pmoles ACE adducts/mg DNA after a 24-h incubation of ACE (16 μg/ml) with C3H10T1/2 cells. ACE-DNA adduct persistence and repair were evaluated in C3H10T1/2 cells using a hydroxyurea block after ACE treatment. ACE-DNA adducts were not repaired under the conditions used in the morphological transformation studies. Thus, ACE provides an interesting example of a mutagenic PAH which is metabolized by C3H10T1/2 cells to active intermediates, forms relatively stable and persistent 2′-deoxyguanosine adducts in C3H10T1/2 cells, and yet induces no detectable morphological transforming activity under the experimental conditions used.