The Analysis of Pyridyloxobutyl RNA Adducts in Rats Treated With 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone

Abstract

Tobacco-specific nitrosamines are potent carcinogens formed from parent compounds such as nicotine during the curing process of tobacco. The cytochrome P450 enzyme family metabolically activates these nitrosamines, which then attach pyridyloxobutyl (POB) groups to DNA bases. The formation of POB-DNA adducts in tissues of rats treated with tobacco-specific nitrosamines have been demonstrated in previous studies. These adducts can potentially lead to mutations in DNA which promote the formation of tumors. However there exists no data on the formation of POB-RNA adducts. RNA adducts could be important in carcinogenesis, and could potentially be a more reliable biomarker than DNA adducts for tobacco-specific nitrosamine exposure. The objective of our project was to chemically characterize and quantify POB-RNA adducts in the tissue of rats treated with a nitrosamine compound, specifically 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). RNA was isolated from rat liver treated with NNK over a course of 20 weeks and from untreated liver. Combined liquid chromatography-mass spectrometry and high performance liquid chromatography were used to identify RNA adducts. The results indicate the presence of POB-RNA adducts in the treated livers. Further work will involve synthesizing standards by reacting NNK with nucleosides and confirming the structure of adducts with NMR. The results of this study will confirm the presence of RNA adducts due to nitrosamine exposure and provide insight into the utility of RNA adducts as a biomarker used for chemopreventative strategies.