Metabolism and macromolecular interaction of the tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone in cultured explants and epithelial cells of human buccal mucosa.

Metabolism and macromolecular interaction of the tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) were studied in human buccal mucosa in vitro. Microautoradiographic analysis of [5-3H]NNK-exposed explant cultures demonstrated a uniform distribution of bound radioactivity in the mucosal epithelium, without significant binding in the underlying connective tissue. The metabolism of [5-3H]NNK at concentrations of both 6 and 100 microM resulted in seven identified metabolites in both explant and epithelial cell cultures. Formation of 4-(methylnitrosamino)-1-(3-pyridyl)butan-1-ol by carbonyl reduction of NNK accounted for almost 95% of the total metabolism, whereas the proportions of other metabolites obtained by alpha-carbon hydroxylation and pyridine N-oxidation reactions varied from undetectable levels to approximately 2% in both experimental systems. A positive correlation between concentration and the metabolic route associated with the formation of DNA methylating intermediates (alpha-hydroxylation at the N-methylene carbon) was found, i.e. when the concentration of NNK was raised from 6 to 100 microM, keto-acid formation which in part reflects DNA methylation was increased preferentially over ketoalcohol production, an index of DNA pyridyloxobutylation. Both the total rate of NNK metabolism and the amount of protein adducts were higher in cells from primary cultures up to the third passage than in explants cultured for 1 day. Between 10(-9) to 10(-4) M, neither NNK nor its precursor alkaloid nicotine affected the colony forming efficiency of normal and tumorous buccal epithelial cells, although at 10(-3) M each agent inhibited this function. Taken together, the results demonstrate the capability of human buccal mucosal epithelium to metabolize NNK by three major pathways, including those involved in the formation of adducts with cellular macromolecules.