title: Role of aromatic amine acetyltransferases, NAT1 and NAT2, in carcinogen-DNA adduct formation in the human urinary bladder.
abstract: The metabolic activation and detoxification pathways associated with the carcinogenic aromatic amines provide an extraordinary model of polymorphisms that can modulate human urinary bladder carcinogenesis. In this study, the metabolic N-acetylation of p-aminobenzoic acid (PABA) to N-acetyl-PABA (NAT1 activity) and of sulfamethazine (SMZ) to N-acetyl-SMZ (NAT2 activity), as well as the O-acetylation of N-hydroxy-4-aminobiphenyl (OAT activity; catalyzed by NAT1 and NAT2), were measured in tissue cytosols prepared from 26 different human bladder samples; then DNA was isolated for determination of NAT1 and NAT2 genotype and for analyses of carcinogen-DNA adducts. Both PABA and OAT activities were detected, with mean activities +/- SD of 2.9 +/- 2.3 nmol/min/mg protein and 1.4 +/- 0.7 pmol bound/mg DNA/min/mg protein, respectively. However, SMZ activities were below the assay limits of detection (< 10 pmol/min/mg protein). The levels of putative carcinogen-DNA adducts were quantified by 32P-postlabeling and averaged 2.34 +/- 2.09 adducts/10(8) deoxyribonucleotide phosphate (dNp). Moreover, the DNA adduct levels in these tissues correlated with their NAT1-dependent PABA activities (r = 0.52; P < 0.01) but not with their OAT activities. Statistical and probit analyses indicated that this NAT1 activity was not normally distributed and appeared bimodal. Applying the NAT1:OAT activity ratios (N:O ratio) allowed arbitrary designation of rapid and slow NAT1 phenotypes, with a cutpoint near the median value. Within each of these subgroups, NAT1 correlated with OAT (P < 0.05); DNA adduct levels were elevated 2-fold in individuals with the rapid NAT1 or NAT1/OAT phenotype. Examination of DNA sequence polymorphisms in the NAT1 gene by PCR have demonstrated that an NAT1 polyadenylation polymorphism is associated with differences in tissue NAT1 enzyme activity; accordingly, NAT1 activity in the bladder of individuals with the heterozygous NAT1*10 allele was 2-fold higher than in subjects homozygous for the putative wild-type NAT1*4 allele. Likewise, DNA adduct levels in the mucosa of the urinary bladder were found to be 2-fold (P < 0.05) higher in individuals with the heterozygous NAT1*10 allele (3.5 +/- 2.1 adducts/10(8) dNp) as compared to NAT1*4 homozygous (1.8 +/- 1.9 adducts/10(8) dNp). Thus, these data provide strong support for the hypothesis that NAT1 activity in the urinary bladder mucosa represents a major bioactivation step that converts urinary N-hydroxy arylamines to reactive N-acetoxy esters that form covalent DNA adducts.(ABSTRACT TRUNCATED AT 400 WORDS)
authors.affiliation: Division of Molecular Epidemiology (HFT-100), National Center for Toxicological Research, Jefferson, Arkansas 72079, USA.
authors.forename: A F
authors.initials: AF
authors.lastname: Badawi
authors.forename: A
authors.initials: A
authors.lastname: Hirvonen
authors.forename: D A
authors.initials: DA
authors.lastname: Bell
authors.forename: N P
authors.initials: NP
authors.lastname: Lang
authors.forename: F F
authors.initials: FF
authors.lastname: Kadlubar
mesh_terms: D010129:4-Aminobenzoic Acid / Q000378:metabolism; D000107:Acetylation; D001191:Arylamine N-Acetyltransferase / Q000235:genetics / Q000502:physiology*; D001483:Base Sequence; D002273:Carcinogens / Q000378:metabolism*; D018736:DNA Adducts / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D012907:Smoking / Q000378:metabolism; D013418:Sulfamethazine / Q000378:metabolism; D001743:Urinary Bladder / Q000378:metabolism*
chemical_list: D002273:Carcinogens; D018736:DNA Adducts; D013418:Sulfamethazine; D001191:Arylamine N-Acetyltransferase; C478900:NAT2 protein, human; D010129:4-Aminobenzoic Acid