Dataset Information

Effect of rapid human N-acetyltransferase 2 haplotype on DNA damage and mutagenesis induced by 2-amino-3-methylimidazo-[4,5-f]quinoline (IQ) and 2-amino-3,8-dimethylimidazo-[4,5-f]quinoxaline (MeIQx).

ABSTRACT: Heterocyclic amines such as 2-amino-3-methylimidazo-[4,5-f]quinoline (IQ) and 2-amino-3,8-dimethylimidazo-[4,5-f]quinoxaline (MeIQx) are dietary carcinogens generated when meats are cooked well-done. Bioactivation includes N-hydroxylation catalyzed by cytochrome P4501A2 (CYP1A2) followed by O-acetylation catalyzed by N-acetyltransferase 2 (NAT2). Nucleotide excision repair-deficient Chinese hamster ovary (CHO) cells stably transfected with human CYP1A2 and either NAT2*4 (rapid acetylator) or NAT2*5B (slow acetylator) alleles were treated with IQ or MeIQx to examine the effect of NAT2 genetic polymorphism on IQ- or MeIQx-induced DNA adducts and mutagenesis. MeIQx and IQ both induced decreases in cell survival and significantly (p<0.001) greater number of endogenous hypoxanthine phosphoribosyl transferase (hprt) mutants in the CYP1A2/NAT2*4 than the CYP1A2/NAT2*5B cell line. IQ- and MeIQx-induced hprt mutant cDNAs were sequenced and over 85% of the mutations were single-base substitutions with the remainder exon deletions likely caused by splice-site mutations. For the single-base substitutions, over 85% were at G:C base pairs. Deoxyguanosine (dG)-C8-IQ and dG-C8-MeIQx adducts were significantly (p<0.001) greater in the CYP1A2/NAT2*4 than the CYP1A2/NAT2*5B cell line. DNA adduct levels correlated very highly with hprt mutants for both IQ and MeIQx. These results suggest substantially increased risk for IQ- and MeIQx-induced DNA damage and mutagenesis in rapid NAT2 acetylators.

SUBMITTER: Metry KJ

PROVIDER: S-EPMC2820402 | biostudies-literature | 2010 Feb

REPOSITORIES: biostudies-literature

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Effect of rapid human N-acetyltransferase 2 haplotype on DNA damage and mutagenesis induced by 2-amino-3-methylimidazo-[4,5-f]quinoline (IQ) and 2-amino-3,8-dimethylimidazo-[4,5-f]quinoxaline (MeIQx).

Metry Kristin J KJ Neale Jason R JR Doll Mark A MA Howarth Ashley L AL States J Christopher JC McGregor W Glenn WG Pierce William M WM Hein David W DW

Mutation research 20091211 1-2

Heterocyclic amines such as 2-amino-3-methylimidazo-[4,5-f]quinoline (IQ) and 2-amino-3,8-dimethylimidazo-[4,5-f]quinoxaline (MeIQx) are dietary carcinogens generated when meats are cooked well-done. Bioactivation includes N-hydroxylation catalyzed by cytochrome P4501A2 (CYP1A2) followed by O-acetylation catalyzed by N-acetyltransferase 2 (NAT2). Nucleotide excision repair-deficient Chinese hamster ovary (CHO) cells stably transfected with human CYP1A2 and either NAT2*4 (rapid acetylator) or NAT ...[more]

PMID: 20004212

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Project description:2-Amino-1,7-dimethylimidazo[4,5-g]quinoxaline (MeIgQx) is a recently discovered heterocyclic aromatic amine (HAA) that is formed during the cooking of meats. MeIgQx is an isomer of 2-amino-3,8-dimethylmidazo[4,5-f]quinoxaline (MeIQx), a rodent carcinogen and possible human carcinogen that also occurs in cooked meats. MeIgQx is a bacterial mutagen, but knowledge about its metabolism and carcinogenic potential is lacking. Metabolism studies on MeIgQx and MeIQx were conducted with human and mouse liver microsomes, and recombinant human P450s. DNA binding studies were also investigated in mice to ascertain the genotoxic potential of MeIgQx in comparison to MeIQx. Both HAAs underwent comparable rates of N-oxidation to form genotoxic N-hydroxylated metabolites with mouse liver microsomes (0.2-0.3 nmol/min/mg protein). The rate of N-oxidation of MeIQx was 4-fold greater than the rate of N-oxidation of MeIgQx with human liver microsomes (1.7 vs 0.4 nmol/min/mg protein). The rate of N-oxidation, by recombinant human P450 1A2, was comparable for both substrates (6 pmol/min/pmol P450 1A2). MeIgQx also underwent N-oxidation by human P450s 1A1 and 1B1 at appreciable rates, whereas MeIQx was poorly metabolized by these P450s. The potential of MeIgQx and MeIQx to form DNA adducts was assessed in female C57BL/6 mice given [(14)C]-MeIgQx (10 μCi, 9.68 mg/kg body wt) or [(14)C]-MeIQx (10 μCi, 2.13 mg/kg body wt). DNA adduct formation in the liver, pancreas, and colorectum was measured by accelerator mass spectrometry at 4, 24, or 48 h post-treatment. Variable levels of adducts were detected in all organs. The adduct levels were similar for both HAAs, when adjusted for dose, and ranged from 1 to 600 adducts per 10(7) nucleotides per mg/kg dose. Thus, MeIgQx undergoes metabolic activation and binds to DNA at levels that are comparable to MeIQx. Given the high amounts of MeIgQx formed in cooked meats, further investigations are warranted to assess the carcinogenic potential of this HAA.

Project description:A targeted liquid chromatography/tandem mass spectrometry-based metabolomics type approach, employing a triple stage quadrupole mass spectrometer in the product ion scan and selected reaction monitoring modes, was established to profile 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), and their principal metabolites in the urine of omnivores. A mixed-mode reverse phase cation exchange resin enrichment procedure was employed to isolate MeIQx and its oxidized metabolites, 2-amino-8-(hydroxymethyl)-3-methylimidazo[4,5-f]quinoxaline (8-CH(2)OH-IQx) and 2-amino-3-methylimidazo[4,5-f]quinoxaline-8-carboxylic acid (IQx-8-COOH), which are produced by cytochrome P450 1A2 (P450 1A2). The phase II conjugates N(2)-(beta-1-glucosiduronyl)-2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline and N(2)-(3,8-dimethylimidazo[4,5-f]quinoxalin-2-yl)-sulfamic acid were measured indirectly, following acid hydrolysis to form MeIQx. The enrichment procedure permitted the simultaneous analysis of PhIP, N(2)-(beta-1-glucosidurony1)-2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine, N3-(beta-1-glucosidurony1)-2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine, 2-amino-1-methyl-6-(4'-hydroxy)-phenylimidazo[4,5-b]pyridine (4'-HO-PhIP), and the isomeric N(2)- and N3-glucuronide conjugates of the carcinogenic metabolite, 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine (HONH-PhIP), which is formed by P450 1A2. The limit of quantification (LOQ) for MeIQx, PhIP, and 4'-HO-PhIP was approximately 5 pg/mL; the LOQ values for 8-CH(2)OH-IQx and IQx-8-COOH were, respectively, <15 and <25 pg/mL, and the LOQ values for the glucuronide conjugates of PhIP and HONH-PhIP were 50 pg/mL. The metabolism was extensive; less than 9% of the dose was eliminated in urine as unaltered MeIQx, and <1% was eliminated as unaltered PhIP. Phase II conjugates of the parent amines accounted for up to 12% of the dose of MeIQx and up to 2% of the dose of PhIP. 8-CH(2)OH-IQx and IQx-8-COOH accounted for up to 76% of the dose of MeIQx, and the isomeric glucuronide conjugates of HONH-PhIP accounted for up to 33% of the dose of PhIP that were eliminated in urine within 10 h of meat consumption. P450 1A2 significantly contributes to the metabolism of both HAAs but with marked differences in substrate specificity. P450 1A2 primarily catalyzes the detoxification of MeIQx by oxidation of the 8-methyl group, whereas it catalyzes the bioactivation of PhIP by oxidation of the exocyclic amine group.

Dataset Information

Effect of rapid human N-acetyltransferase 2 haplotype on DNA damage and mutagenesis induced by 2-amino-3-methylimidazo-[4,5-f]quinoline (IQ) and 2-amino-3,8-dimethylimidazo-[4,5-f]quinoxaline (MeIQx).

Publications

Effect of rapid human N-acetyltransferase 2 haplotype on DNA damage and mutagenesis induced by 2-amino-3-methylimidazo-[4,5-f]quinoline (IQ) and 2-amino-3,8-dimethylimidazo-[4,5-f]quinoxaline (MeIQx).

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