High Sensitivity and Fragmentation Specificity in the Analysis of Drug−DNA Adducts by Electrospray Tandem Mass Spectrometry

Hedamycin is a naturally-occurring antitumor antibiotic that binds to double-stranded DNA by both intercalation and alkylation. DNA sequencing shows that alkylation occurs exclusively at guanine bases, but with a distinct preference for guanines located in 5'-C\GT-3' and 5'-C\GG-3' sequences. NMR-derived solution structures of hedamycin covalently bound to double-stranded oligonucleotides show that the anthrapyrantrione chromophore of the ligand threads the double helix, placing the two amino sugars in the minor groove and the alkylating bis-epoxide side chain in the major groove. Our previous studies of hedamycin--DNA binding by ESI-MS have confirmed that hedamycin significantly increases duplex stability and provided strong evidence for the formation of specifically, base-paired duplexes in the gas phase. We demonstrate here that ESI-MS/MS can be used to determine unequivocally the sequence selectivity of base alkylation by a DNA-binding antitumor drug, using hedamycin as a model.