Identification of Stable Benzo[a]pyrene-7,8-dione-DNA Adducts in Human Lung Cells

Metabolic activation of the proximate carcinogen benzo­[a]­pyrene-7,8-trans-dihydrodiol (B­[a]­P-7,8-trans-dihydrodiol) by aldo-keto reductases (AKRs) leads to B­[a]­P-7,8-dione that is both electrophilic and redox-active. B­[a]­P-7,8-dione generates reactive oxygen species resulting in oxidative DNA damage in human lung cells. However, information on the formation of stable B­[a]­P-7,8-dione-DNA adducts in these cells is lacking. We studied stable DNA adduct formation of B­[a]­P-7,8-dione in human lung adenocarcinoma A549 cells, human bronchoalveolar H358 cells, and immortalized human bronchial epithelial HBEC-KT cells. After treatment with 2 μM B­[a]­P-7,8-dione, the cellular DNA was extracted from the cell pellets subjected to enzyme hydrolysis and subsequent analysis by LC-MS/MS. Several stable DNA adducts of B­[a]­P-7,8-dione were only detected in A549 and HBEC-KT cells. In A549 cells, the structures of stable B­[a]­P-7,8-dione-DNA adducts were identified as hydrated-B­[a]­P-7,8-dione-N²-2′-deoxyguanosine and hydrated-B­[a]­P-7,8-dione-N1-2′-deoxyguanosine. In HBEC-KT cells, the structures of stable B­[a]­P-7,8-dione-DNA adducts were identified as hydrated-B­[a]­P-7,8-dione-2′-deoxyadenosine, hydrated-B­[a]­P-7,8-dione-N1- or N3-2′-deoxyadenosine, and B­[a]­P-7,8-dione-N1- or N3-2′-deoxyadenosine. In each case, adduct structures were characterized by MSⁿ spectra. Adduct structures were also compared to those synthesized from reactions of B­[a]­P-7,8-dione with either deoxyribonucleosides or salmon testis DNA in vitro but were found to be different.