4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and N'-nitrosonornicotine (NNN) are two of the most important carcinogens in tobacco smoke and are the only carcinogens known to occur in relatively high levels in unburned tobacco. As such, these two nitrosamines are likely to be causative factors in human cancer. Our studies, supported by this grant, have established metabolic pathways of these carcinogens leading to DNA modification. A major advance in the past grant period has been the demonstration that NNK methylates target tissue DNA of F344 rats, with formation of the promutagenic lesion 06-methylguanine. In addition, we have shown that NNK is more carcinogenic than dimethylnitrosamine (DMN) in F344 rats. We are proposing to continue our studies on the metabolic activation of NNK and NNN as follows: (1) characterize and quantify the adducts formed by pyridyloxobutylation of DNA upon Alpha-hydroxylation of NNK and NNN by continuing our studies on the reactions of model compounds with deoxyribonucleosides and DNA, by continuing our investigations of the in vivo modification of DNA by high specific activity tritium labelled NNN and NNK, and by developing new postlabelling methods for detection of the DNA adducts; (2) Compare the kinetics of formation and persistence of specific methylated DNA adducts in rat target tissues upon treatment with either NNK, its metabolite NNAl, or DMN at doses equivalent to those used in our bioassay, and determine the effects of NNK and its metabolites on DNA repair enzymes; (3) Test our hypothesis that the unique tumorigenicity of NNK is due to combined methylation and pryidyloxobutylation of DNA by carrying out a comparative bioassay in A/J mice of NNK and the appropriate model compounds, and; (4) Assess the role of Beta- and Gamma-hydroxylation in the metabolic activation of NNK. These studies will lead to a better understanding of the role of these nitrosamines in cancer induction by tobacco.
