ARYLAMINE N-ACETYLTRANSFERASE ACTIVITY IN HUMAN CULTURED-CELL LINES

Many arylamine and hydrazine drugs and xenobiotics are acetylated by l iver N-acetyltransferase (NAT; EC 2.3.1.5). Two loci, mnat and pnat, e ncode the enzymes designated monomorphic and polymorphic NAT (mNAT and pNAT) respectively. These isoenzymes have different substrate specifi cities. In addition, at the polymorphic locus a diversity of alleles i s found, which differ by specific point mutations that may or may not result in amino acid substitutions. These point mutations result in th e 'slow' acetylation of substrates of pNAT. The substrates for NAT inc lude carcinogenic arylamines. Susceptibility to bladder cancer has bee n related to slow acetylation. NAT has been characterized in immortali zed human cell lines to assess their use in studies of the metabolism or arylamines in vitro. A monocytic cell line (U937) and two hepatoma cell lines of parenchymal lineage (HepG2 and Hep3B) have been shown to catalyse acetylation of substrates of mNAT but do not acetylate sulph amethazine, a substrate specific for pNAT. Using PCR to amplify the al leles of pNAT, followed by restriction-enzyme digestion of the product , the cell lines have been genotyped: U937 cells are homozygous slow a cetylators (S1a/S1a) and HepG2 cells are heterozygous slow acetylators (S1a/S2). Transcription of pnat was confirmed in the hepatoma cell li nes, by amplification of cDNA generated from these cells. In addition, splicing of mRNA specific for pNAT has been demonstrated by using a p rimer which anneals to a region in the 5' promoter region. Unlike the hepatoma cell lines, in U937 cells the pNAT gene is not transcribed. H owever, transcription of mnat was shown to occur in all three cell lin es.