High-Resolution Mass Spectrometric Analysis of Selected DNA Adducts in Smokeless Tobacco Users’ Oral Cells

Abstract

Smokeless tobacco, used by more than 360 million people worldwide, is an accepted cause of cancers of the oral cavity, esophagus, and pancreas. DNA adducts are considered as critical intermediates in the carcinogenic process, but there are no published studies that have used structure-specific methods to identify and quantify oral cell DNA adducts in people who use smokeless tobacco. In the study presented here, we used liquid chromatography-nanoelectrospray ionization-high resolution tandem mass spectrometry to identify and quantify DNA adducts in oral cells of 24 individuals who use smokeless tobacco and 25 individuals not using any tobacco or nicotine product. We quantified the following DNA adducts: (8R/S)-3-(2′-deoxyribos-1′-yl)-5,6,7,8-tetrahydro-8-hydroxypyrimido[1,2-a]purine-10(3H)-one (γ-OH-Acr-dGuo) from acrolein; (6S,8S and 6R,8R)-3-(2′-deoxyribos-1′-yl)-5,6,7,8-tetrahydro-8-hydroxy-6-methylpyrimido[1,2-a]purine-10(3H)-one [(S,R)γ-OH-Cro-dGuo] from crotonaldehyde and/or acetaldehyde; 3-methyladenine (3-Me-Ade); 7-ethylguanine (7-Et-Gua); and 8-oxo-deoxyguanosine (8-oxo-dGuo). Levels [median (min, max)] fmol/μmol dGuo or Gua, in samples in which analytes were detected, were: γ-OH-Acr-dGuo [350 (169, 2217) vs 114 (14, 278)], (6S,8S and 6R,8R)γ-OH-Cro-dGuo [1058 (222, 2569) vs not detected], and 7-Et-Gua [2445 (1052, 5389) vs 102 (13, 627)], which were significantly greater in the oral cell DNA of individuals who use smokeless tobacco than in those not using any tobacco or nicotine product (all adjusted Ps < 0.01), while amounts of 3-Me-Ade [3739 (93, 11495) vs 1447 (318, 3596) fmol/μmol Gua, adjusted P = 0.052] and 8-oxo-dGuo [1520 (513, 5232) vs 2092 (433, 3325) pmol/μmol dGuo, adjusted P = 0.13] were not significantly different between the two groups. The sizable and significant elevations of 7-Et-Gua and (6S,8S and 6R,8R)γ-OH-Cro-dGuo in oral cell DNA when smokeless tobacco was used versus controls were particularly notable and unexpected. The results of this study provide some potentially important new data regarding DNA adduct levels in oral cells of people who use smokeless tobacco.