Journal Article

Differential metabolism of 2-amino-1-methyl-6-phenylimidazo[4,5-<i>b</i>]pyridine in rat and human hepatocytes

Sophie Langouët, Axel Paehler, Dieter H. Welti, Nathalie Kerriguy, André Guillouzo and Robert J. Turesky

in Carcinogenesis

Volume 23, issue 1, pages 115-122
Published in print January 2002 | ISSN: 0143-3334
Published online January 2002 | e-ISSN: 1460-2180 | DOI:
Differential metabolism of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine in rat and human hepatocytes

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Metabolism of the carcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) has been compared in human and rat hepatocytes. The identities of seven metabolites were confirmed by UV and mass spectroscopy and by co-elution with reference standards using HPLC. In human hepatocytes, the major biotransformation pathway of PhIP was cytochrome P4501A2 (CYP1A2)-mediated N-oxidation to form the genotoxic metabolite 2-(hydroxyamino)-1-methyl-6-phenylimidazo[4,5-b]pyridine (HONH-PhIP), which underwent glucuronidation at the N2 and N3 positions of PhIP to form stable conjugates. These products combined accounted for as much as 60% of the added PhIP. Direct glucuronidation of PhIP at the N2 and N3 positions also occurred, accounting for up to 20% of the amount added. Glucuronide and sulfate conjugates of 2-amino-4′-hydroxy-1-methyl-6-phenylimidazo[4,5-b]pyridine (4′-HO-PhIP) were also detected, comprising 5 and 12% of the products, respectively. The CYP1A2 inhibitor furafylline diminished the formation of both HONH-PhIP glucuronide conjugates in a concentration-dependent manner, however, levels of 4′-HO-PhIP were unchanged, indicating that CYP1A2 does not significantly contribute to 4′-hydroxylation of PhIP. Hepatocytes of male rats, both untreated and pretreated with the CYP1A2 inducer 3-methylcholanthrene (3-MC) transformed PhIP into 4′-HO-PhIP as the prominent product. Unconjugated and conjugated 4′-HO-PhIP metabolites combined accounted for 18 and 46% of the PhIP products in untreated and in 3-MC-pretreated rat hepatocytes, respectively. The isomeric glucuronide conjugates of HONH-PhIP combined accounted for 11 and 26% of the PhIP, respectively, in untreated and 3-MC-pretreated hepatocytes. The regioselectivity of glucuronidation of PhIP was different in human and rat hepatocytes. Human liver UDP-glucuronosyltransferases favored conjugation to the N2 positions of PhIP and HONH-PhIP, while the N3 atom was the preferred site of conjugation for the rat enzymes. Thus, important differences exist between human and rat enzymes in catalytic activity and regioselectivity of PhIP metabolism. Some human hepatocyte populations are more active at transforming PhIP to a genotoxic species than rat hepatocytes pretreated with the potent CYP1A2 inducer 3-MC.

Keywords: CID, collision-induced dissociation; CYP, cytochrome P450; EROD, ethoxyresorufin O-deethylation; FCS, fetal calf serum; HAA, heterocyclic aromatic amine; LC-ESI-MS, liquid chromatography electrospray ionization mass spectrometry; 3-MC, 3-methylcholanthrene; MeIQx, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline; NOE, nuclear Overhauser enhancement; PhIP, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine; 4'-HO-PhIP, 2-amino-4'-hydroxyl-1-methyl-6-phenylimidazo[4,5-b]pyridine; HONH-PhIP, 2-(hydroxyamino)-1-methyl-6-phenylimidazo[4,5-b]pyridine5-HO-PhIP, 2-amino-1-methyl-6-(5-hydroxy)phenylimidazo[4,5-b]pyridine; PhIP-4'-OSO3H, 4-(2-amino-1-methylimidazo[4,5-b]pyridin-6-yl)phenyl sulfate; HON-PhIP-N2-Gl, N2-β-d-glucosiduronyl-2-(hydroxyamino)-1-methyl-6-phenylimidazo[4,5-b]pyridine; PhIP-N2-Gl, N2-β-d-glucosiduronyl-2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine; HON-PhIP-N3-Gl, N3-β-d-glucosiduronyl-2-(hydroxyamino)-1-methyl-6-phenylimidazo[4,5-b]pyridine; PhIP-N3-Gl, N3-β-d-glucosiduronyl-2-amino-1-methyl-6-phenylimidazo[4,5-b] pyridine; PhIP-4'-O-Gl; 4'-β-d-glucosiduronyloxy-2-amino-4'-hydroxyl-1-methyl-6-phenylimidazo[4,5-b]pyridine.

Journal Article.  5454 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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