• Aromatic DNA adducts and polymorphisms in metabolic genes in healthy adults: findings from the EPIC-Spain cohort.

      Agudo, Antonio; Peluso, Marco; Sala, Nuria; Capella, Gabriel; Munnia, Armelle; Piro, Sara; Marin, Fatima; Ibanez, Raquel; Amiano, Pilar; Tormo, M. Jose; et al. (2009-06)
      Aromatic compounds such as polycyclic aromatic hydrocarbons, arylamines and heterocyclic amines require metabolic activation to form metabolites able to bind to DNA, a process mediated by polymorphic enzymes. We measured aromatic DNA adducts in white blood cells by the (32)P-post-labelling assay in a sample of 296 healthy adults (147 men and 149 women) from five regions of Spain. We also analyzed functional polymorphisms in the metabolic genes CYP1A1, CYP1A2, EPHX1, GSTM1, GSTT1, NAT2 and SULT1A1. A significant increased level of DNA aromatic adducts was found related to the fast oxidation-hydrolysis phenotype defined by the polymorphism I462V in CYP1A1, the allele A in IVS1-154C>A of CYP1A2 and the combination Tyrosine-Arginine for Y113H and H139R of EPHX1. Geometric means (adducts per 10(-9) normal nucleotides) were 2.17, 4.04 and 6.30 for slow, normal and fast phenotypes, respectively (P-trend = 0.01). Slow acetylation by NAT2 was associated with a significant decrease in adduct level; subjects with slow alleles *5A and *7A/B had in average 1.56 x 10(-9)adducts, as compared with 5.60 for those with normal NAT2 activity (P-value = 0.01). No association was seen with polymorphisms of other metabolic genes such as GSTM1, GSTT1 or SULT1A1. We concluded that the metabolic pathways of oxidation, hydrolysis and acetylation are relevant to the formation of bulky DNA adducts. This could suggest a potential involvement of aromatic compounds in the formation of such adducts; however, given lack of specificity of the post-labeling assay, a firm conclusion cannot be drawn.
    • Concordance between the deduced acetylation status generated by high-speed: real-time PCR based NAT2 genotyping of seven single nucleotide polymorphisms and human NAT2 phenotypes determined by a caffeine assay.

      Rihs, Hans-Peter; John, Andrea; Scherenberg, Michael; Seidel, Albrecht; Bruning, Thomas (2007-02)
      BACKGROUND: The utility of typing single nucleotide polymorphisms (SNPs) for the determination of the N-acetyltransferase 2 (NAT2) acetylation status is a matter of debate. AIMS OF THE STUDY: Evaluation of the concordance between deduced genotype results of seven human NAT2 SNPs generated by Real-time PCR analysis and human NAT2 phenotypes. METHODS: NAT2 phenotypes of 38 Caucasian workers were determined using a suitable caffeine test method. Genomic DNA aliquots were used for the determination of seven human NAT2-specific SNPs (G191A, C282T, T341C, C481T, G590A, A803G, G857A). RESULTS AND CONCLUSIONS: Phenotypic results based on the molar ratio of 5-acetylamino-6-formylamino-3-methyluracil (AFMU)/(AFMU+1-methlyuric acid (1U)+1-methylxanthine (1X)) calculated from excreted caffeine metabolite levels in urine samples with 0.3 as a cut-off point between slow (<0.3) and rapid acetylators (>or=0.3). Twenty-seven samples belonged to the slow (mean 0.13; range: 0.03-0.25), 11 to the rapid (mean: 0.41; range: 0.34-0.48) acetylators. LightCycler analyses revealed 11 different NAT2 variant combinations, whereby *5B/*5B and *5B/*6A or *5A/*6C (each 21%), were the most frequent. The deduced acetylation status of the seven NAT2 SNPs matched perfectly with the 38 results determined by phenotyping. This study showed a 100% concordance between NAT2 phenotypes and the deduced NAT2 genotypes and the suitability of the high-speed NAT2-specific LightCycler analysis in a Caucasian population.