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]quinoxaline.
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.
Division of Drug Metabolism, Faculty of Pharmaceutical Sciences, Hokkaido University, Japan.
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Y
Yanagawa
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Sawada
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Deguchi
F J
FJ
Gonzalez
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Kamataki
D000123:Acetyltransferases / Q000378:metabolism*; D016604:Aflatoxin B1 / Q000633:toxicity; D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D019388:Cytochrome P-450 CYP1A2; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D009152:Mutagenicity Tests; D009153:Mutagens / Q000378:metabolism*; D010088:Oxidoreductases / Q000378:metabolism*; D011804:Quinolines / Q000378:metabolism*; D011810:Quinoxalines / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism
D009153:Mutagens; D011804:Quinolines; D011810:Quinoxalines; D012333:RNA, Messenger; C029216:2-amino-3-methylimidazo(4,5-f)quinoline; C036990:2-amino-3,8-dimethylimidazo(4,5-f)quinoxaline; D003577:Cytochrome P-450 Enzyme System; D016604:Aflatoxin B1; D010088:Oxidoreductases; D019388:Cytochrome P-450 CYP1A2; D000123:Acetyltransferases