Acetaldehyde is the major metabolite of ethanol and has recently been classified by IARC as a Group-1 human carcinogen. Despite a clear association, the underlying mechanisms of ethanol-induced carcinogenesis remain unclear. Acetaldehyde is a DNA-modifying compound. If not eliminated or repaired, these modifications can result in miscoding events that lead to mutations, and ultimately, cancer initiation. To gain new advances on ethanol and acetaldehyde-induced DNA damage and devise preventive measures, an innovative mass spectrometry method able to simultaneously detect all acetaldehyde-derived DNA adducts was developed. Our method is based on a gas phase fractionation-adductomic approach, which monitors the constant neutral loss of features common to all DNA adducts: the 2’-deoxyribose and/or one of the four nucleobases. By exposing DNA samples to 12C2- and 13C2-acetaldehyde and performing the data analysis through in-house software, we were able to identify 20 new DNA adducts, as well as all previously observed DNA adducts associated with acetaldehyde exposure. The reduced product of Schiff bases/imines were found in samples reduced with NaBH3CN, meanwhile DNA crosslinks and Mannich products were observed in non-reduced samples. N6-ethyl-dAdo and N4-ethyl-dCyd were identified as novel adducts. Because of the biological relevance of DNA methylation and their structural similarity to methyl-dAdo and methyl-dCyd, characterization of N6-ethyl-dAdo and N4-ethyl-dCyd was achieved by synthetizing isotopically-labeled standards. N6-Ethyl-dAdo, N4-Ethyl-dCyd, D5-N6-Ethyl-dAdo and D5-N4-Ethyl-dCyd were synthesized, purified, and structurally characterized by one- and two-dimensional NMR and HRMS analyses. A dose-response relationship was observed between the three primary DNA adducts and acetaldehyde exposure. Finally, the presence of these adducts was confirmed in oral DNA samples from humans exposed to alcohol. Ultimately, we confirmed that our approach can characterize ethanol- and acetaldehyde-induced DNA damage and that both dAdo and dCyd adducts are potential markers of these exposures.

Molecular characterization of alcohol-induced DNA damage for cancer prevention

256th National Meeting (2018)

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