Metabolism and DNA adduct formation of 2-acetylaminofluorene by bladder explants from human, dog, monkey, hamster and rat

Abstract

Cultured bladder explants from human, dog, monkey, hamster and rat were used to study the metabolism, and DNA-adduct formation of [3H]2-acetylaminofluorene (AAF). After 24 h of incubation with 1 microM [3H]AAF, the organic-soluble and glucuronide fractions represented 29.0-58.4% and 0.7-10.8%, respectively, of the total amount of radioactive material in the medium. In the organic-soluble fraction of all species, the presence of both ring-hydroxylated (at the 7-, 9-, 5-, 3- and 1-carbons) metabolites and N-hydroxy-AAF could be demonstrated, with 7-hydroxy-AAF and 9-hydroxy-AAF being the most ubiquitous metabolic products. In addition, 2-aminofluorene could be detected in all cases with the highest amounts formed by dog bladder explants. Species susceptible to aromatic amine-induced bladder carcinogenesis (human and dog) showed widely varying DNA binding (0.23 +/- 0.20 and 1.95 +/- 1.2 mumol AAF/mol deoxyribonucleotide, respectively), and in explants of the less susceptible species (monkey, hamster and rat) DNA binding was comparable (0.46 +/- 0.21, 0.37 +/- 0.07 and 0.34 +/- 0.27, respectively). Enzymatic hydrolysis of the [3H]AAF--DNA to the nucleoside level revealed that N-(deoxyguanosine-8-yl)-2-aminofluorene, together with small amounts (5.9-15.4% of the total) of its imidazole ring-opened derivative, was the most ubiquitous adduct formed (17.8-47.2% of the total) in all species, except the dog. In dog bladder DNA, N-(deoxyguanosine-8-yl)-2-acetylaminofluorene was the principal DNA adduct (39.0-46.5% of the total) with smaller amounts (15.5-22.2% of the total) in the other species. In addition, small amounts (3.5-8.6% of the total) of another adduct, 3-(deoxygaunosin-N2-yl)-2-acetylaminofluorene, could be detected in all cases. It is concluded that bladder explants of the human, dog, monkey, hamster and rat metabolize AAF mainly to ring-hydroxylated products, but also form small amounts of the proximate carcinogenic metabolite N-hydroxy-AAF. Neither the overall binding of AAF to bladder DNA, nor the formation of specific AAF--DNA adducts is correlated with the relative susceptibilities of these five species to aromatic amine-induced urinary bladder carcinogenesis.