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Two separate sets (Bio. 1.5/H and Bio. 82.73/H) of Syrian hamster lines congenic at the NAT2 gene locus were constructed. In both sets, N-acetylation capacity was NAT2-dependent in vivo and in vitro, with highest levels in homozygous rapid acetylators, intermediate levels in heterozygous acetylators and lowest levels in homozygous slow acetylators. NAT1 and NAT2 acetyltransferase isozymes from liver and colon cytosols activated N-hydroxy-2-aminofluorene (N-OH-AF) to DNA adducts. NAT2-catalyzed metabolic activation of N-OH-AF was acetylator genotype-dependent. NAT1-catalyzed metabolic activation of N-OH-AF was independent of acetylator genotype. Hamster NAT1 and NAT2 were cloned, sequenced, and expressed. Recombinant NAT1 from rapid and slow acetylators activated N-OH-AF at equivalent rates, but rapid acetylator recombinant NAT2 activated N-OH-AF at rates over 750-fold higher than slow acetylator recombinant NAT2. These results provide clear evidence for metabolic activation of arylamine carcinoge...

Arylamine chemicals inflict a number of toxicities including cancer. Metabolic activation (i.e., oxidation) is required in order to elicit the toxic actions. Acetylation is an important step in the metabolic activation and deactivation of arylamines. N-acetylation forms the amide derivative which is often nontoxic. However, O-acetylation of the N-hydroxyarylamine (following oxidation) yields an acetoxy arylamine derivative which breaks down spontaneously to a highly reactive arylnitrenium ion, the ultimate metabolite responsible for mutagenic and carcinogenic lesions. Human capacity to acetylate arylamine chemicals is subject to a genetic polymorphism. Individuals segregate into rapid, intermediate, or slow acetylator phenotypes by Mendelian inheritance regulated by a single gene encoding for a polymorphic acetyltransferase isozyme (NAT2). Individuals homozygous for mutant alleles are deficient in the polymorphic acetyltransferase and are slow acetylators. A second acetyltransferase isozyme (NAT1) is monomorphic and is not regulated by the acetylator genotype. Several human epidemiological studies suggest an association between slow acetylator phenotype and urinary bladder cancer. In contrast, a few studies suggest a relationship between rapid acetylator phenotype and colorectal cancer The basis for this paradox may relate to the relative importance of N- versus O-acetylation in the etiology of these cancers. Conclusions drawn from human epidemiological data are often compromised by uncontrolled environmental and other genetic factors. Our laboratory recently completed construction of homozygous rapid, heterozygous intermediate, and homozygous slow acetylator congenic Syrian hamsters to be homologous in greater than 99.975% of their genomes. The availability of these acetylator congenic lines should eliminate genetic variability in virtually all aspects of arylamine carcinogenesis except at the acetylator gene locus. Ongoing studies in these congenic hamster lines should provide unequivocal information regarding the role of genetic acetylator phenotype in susceptibility to arylamine-related cancers.

Our laboratory recently reported the successful construction of homozygous rapid (Bio. 82.73/H-Patr) and homozygous slow (Bio. 82.73/H-Pat(s)) acetylator congenic Syrian hamsters. These hamsters are isogenic except for the polymorphic acetylator gene locus (Pat) and perhaps other closely linked loci. The purpose of the present investigation was to assess the expression of acetylator genotype both in vivo and in vitro in a variety of hepatic and extrahepatic organ cytosols. Levels of arylamine N-acetyl-transferase were generally high and in the relative order: liver greater than colon greater than kidney greater than pancreas greater than prostate, urinary bladder, and lung. However, an acetylator gene dose-response was clearly expressed in each tissue, with highest levels in homozygous Patr acetylators, intermediate levels in heterozygous Patr/Pat(s) acetylators, and lowest levels in homozygous Pat(s) acetylators. The magnitude of the acetylator genotype-dependent differences in N-acetyltransferase activity were substrate specific, wherein p-aminobenzoic acid showed the largest differences and p-aminophenol the smallest. The N-acetylation of p-aminobenzoic acid in vivo also reflected acetylator genotype in the congenic hamsters. These results further document the successful construction of rapid and slow acetylator congenic hamsters which should prove very valuable in future studies to assess the role of acetylator genotype in the toxicity and carcinogenicity of arylamine chemicals.