Metabolism of 2-acetylaminofluorene in the Chinese hamster ovary cell mutation assay.
Chinese hamster ovary (CHO) cells were exposed to 2-acetylaminofluorene (2-AAF) and 2-aminofluorene (2-AF), and several of their N-oxidized metabolites in order to study the mechanisms by which arylamides and arylamines produce mutations in mammalian cells. The number of mutations induced at the hypoxanthine-guanine phosphoribosyl transferase locus by each compound (mutants/10(6) CHO cells/nmol compound/ml) was estimated to be: N-acetoxy-2-AAF, 310; N-hydroxy-2-AF, 3; N-hydroxy-2-AAF (with and without hepatic S9 activation), 0.7; 2-AAF (with S9), 0.1; and 2-AF (with S9), 0.09. With each compound, DNA adducts were also identified and quantified, and in all cases the major adduct was N-(deoxyguanosin-8-yl)-2-AF. 2-AAF and N-hydroxy-2-AAF also formed minor amounts of N-(deoxyguanosin-8-yl)-2-AAF and 3-(deoxyguanosin-N2-yl)-2-AAF. The relationship between mutation induction and adduct formation for each of the derivatives was similar to that previously reported for N-hydroxy-2-AF. Inclusion of the deacetylase inhibitor, paraoxon, reduced the mutagenicity of 2-AAF, N-hydroxy-2-AAF and N-acetoxy-2-AAF, and the DNA adducts produced by N-acetoxy-2-AAF to background levels. Acetyl coenzyme A increased the mutations and CHO cytosol-mediated DNA binding of N-hydroxy-2-AAF, but did not substantially increase these responses from N-hydroxy-2-AF. N-Hydroxy-2-AAF was not detectably metabolized by CHO cells. Taken together, these data indicate that CHO cells metabolized N-acetoxy-2-AAF to a reactive derivative by N-deacetylation to N-acetoxy-2-AF, while N-hydroxy-2-AF reacted directly with DNA. The major pathway of N-hydroxy-2-AAF activation appeared to be an initial O-acetylation to N-acetoxy-2-AAF and this occurred to only a limited extent in the CHO cells. N-Hydroxy-2-AAF also seemed to form an additional unknown ester intermediate that gave rise to acetylated DNA adducts. The initial step in the activation of 2-AAF and 2-AF was an N-oxidation to N-hydroxy-2-AAF and N-hydroxy-2-AF, respectively. The limited O-acetylase activity in CHO cells appeared to contribute to the low sensitivity of these cells toward mutation induction by arylamines and arylamides.
Heflich
R H
RH
Department of Health and Human Services, Food and Drug Administration, Jefferson, AR 72079.
Djurić
Z
Z
Zhuo
Z
Z
Fullerton
N F
NF
Casciano
D A
DA
Beland
F A
FA
D015073:2-Acetylaminofluorene / Q000378:metabolism* / Q000494:pharmacology; D000099:Acetoxyacetylaminofluorene / Q000378:metabolism / Q000494:pharmacology; D000105:Acetyl Coenzyme A / Q000494:pharmacology; D000107:Acetylation; D000818:Animals; D001711:Biotransformation; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D003600:Cytosol / Q000378:metabolism; D004249:DNA Damage; D005260:Female; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism*; D005449:Fluorenes / Q000378:metabolism / Q000494:pharmacology; D006881:Hydroxyacetylaminofluorene / Q000378:metabolism / Q000494:pharmacology; D009152:Mutagenicity Tests; D010053:Ovary; D010084:Oxidation-Reduction; D010261:Paraoxon / Q000494:pharmacology
D005449:Fluorenes; C012177:2-aminofluorene; D006881:Hydroxyacetylaminofluorene; D000099:Acetoxyacetylaminofluorene; C001368:N-hydroxy-2-aminofluorene; D000105:Acetyl Coenzyme A; D015073:2-Acetylaminofluorene; D010261:Paraoxon