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Stable Expression of Human CYP1A2 and N-Acetyltransferases in Chinese Hamster CHL Cells: Mutagenic Activation of 2-Amino-3-methylimidazo[4,5-f]quinoline and 2-Amino-3,8-dimethylimidazo[4,5-f]quinoxaline¹

Division of Drug Metabolism, Faculty of Pharmaceutical Sciences, Hokkaido University, N12 W6, Sapporo, Hokkaido 060, Japan [Y. Y., M. S., T. K.]; Department of Molecular Neurobiology, Tokyo Metropolitan Institute for Neurosciences, 2-6 Musashidai, Fuchu, Tokyo 183, Japan [T. D.]; and Laboratory of Molecular Carcinogenesis, National Institutes of Health, Bethesda, Maryland 20892 [F. J. G.]

In order to investigate the metabolic activation pathway of food-derived heterocyclic amines, 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), cultured cell lines which stably expressed human cytochrome P4501A2 (CYP1A2) and N-acetyltransferases (NATs) were developed by the method of complementary DNA (cDNA) transfection. First, a cell line expressing CYP1A2, designated A2R-5, was established from the cell line CR-68, which was previously established by introducing NADPH-cytochrome P-450 reductase cDNA into Chinese hamster CHL cells. The expression of CYP1A2 in the transfected cells was confirmed by determining sensitivity to aflatoxin B₁. As the next step, the A2R-5 as well as CR-68 cells were further transfected with human monomorphic NAT (NAT1) or polymorphic NAT (NAT2) cDNAs. The expression of NAT in the transfected cells was confirmed using p-aminobenzoic acid and sulfamethazine as substrates, while no activity was seen in parental CR-68 and A2R-5 cells. The cell line, ANP-25, which expressed both CYP1A2 and NAT2, was approximately 370- and 100-fold more sensitive to IQ and MeIQx, respectively, than parental CR-68 cells in cytotoxicity assays. There were no clear differences in sensitivity to both compounds among CR-68, A2R-5, and the cell lines which expressed NAT1 alone, NAT2 alone, and CYP1A2 plus NAT1. Mutagenicity of IQ and MeIQx at the hypoxanthine-guanine phosphoribosyltransferase locus was also detectable only in ANP-25 cells but not in A2R-5 or the cell line expressing CYP1A2 plus NAT1. From these results, it is proposed that both CYP1A2 and NAT2 (but not NAT1) are required for mutagenic activation of these compounds, implying that acetylator polymorphism may be an important risk factor in the carcinogenicity of these compounds.

¹ This work was supported partly by a grant-in-aid for scientific research from the Ministry of Education, Science and Culture of Japan and by the Japan Health Sciences Foundation.

² Present address: Division of Environmental Hygiene, Hokkaido Institute of Pharmaceutical Sciences, 7-1 Katsuraoka-cho, Otaru, Hokkaido 047-02, Japan.

³ Present address: Research and Development Division, BML, Inc., 1361-1 Matoba, Kawagoe, Saitama 350, Japan.

⁴ To whom requests for reprints should be addressed.

Received 12/ 6/93. Accepted 5/ 3/94.

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