Acetaldehyde (AA) is the major metabolite of
ethanol and may be responsible for an increased
gastrointestinal cancer risk associated with alcohol beverage consumption. Furthermore, AA is one of the most abundant
carcinogens in tobacco
smoke and induces
tumors of the respiratory tract in laboratory animals. AA binding to
DNA induces
Schiff base adducts at the exocyclic amino group of dG,
N2-ethylidene-dG, which are reversible on the
nucleoside level but can be stabilized by reduction to
N2-ethyl-dG. Mutagenesis studies in the
HPRT reporter gene and in the p53 tumor suppressor gene have revealed the ability of AA to induce G-->A transitions and A-->T transversions, as well as frameshift and splice mutations. AA-induced point mutations are most prominent at 5'-AGG-3' trinucleotides, possibly a result of sequence specific adduct formation, mispairing, and/or repair. However, DNA sequence preferences for the formation of
acetaldehyde adducts have not been previously examined. In the present work, we employed a stable
isotope labeling-HPLC-ESI+-MS/MS approach developed in our laboratory to analyze the distribution of
acetaldehyde-derived
N2-ethyl-dG adducts along double-stranded
oligodeoxynucleotides representing two prominent
lung cancer mutational "hotspots" and their surrounding DNA sequences. 1,7,NH 2-(15)N-2-(13)C-dG was placed at defined positions within
DNA duplexes derived from the K-ras protooncogene and the p53 tumor suppressor gene, followed by AA treatment and NaBH 3CN reduction to convert
N2-ethylidene-dG to
N2-ethyl-dG. Capillary HPLC-ESI+-MS/MS was used to quantify
N2-ethyl-dG adducts originating from the isotopically labeled and unlabeled
guanine nucleobases and to map adduct formation along
DNA duplexes. We found that the formation of
N2-ethyl-dG adducts was only weakly affected by the local sequence context and was slightly increased in the presence of
5-methylcytosine within CG dinucleotides. These results are in contrast with sequence-selective formation of other tobacco
carcinogen-
DNA adducts along K-ras- and p53-derived duplexes and the preferential modification of endogenously methylated CG dinucleotides by
benzo[a]pyrene diol
epoxide and
acrolein.