Characterization of a unique tomaymycin-d(CICGAATTCICG) sub 2 adduct containing two drug molecules per duplex by NMR, fluorescence, and molecular modeling studies

Tomaymycin is a member of the pyrrolo(1,4)benzodiazepine (P(1,4)B) antitumor antibiotic group. This antibiotic is proposed to react with the exocyclic 2-amino group (N2) of guanine to form a covalent adduct that lies snugly within the minor groove of DNA. While DNA-footprinting experiments using methidiumpropyl-EDTA have revealed the favored bonding sequences for tomaymycin and related drugs on DNA, the stereochemistry at the covalent bonding site (C-11) and orientation in the minor groove were not established by these experiments. In previous studies using a combined fluorescence, high-field NMR, and molecular modeling approach, the authors have shown that for tomaymycin there are two diastereomeric species (11R and 11S) on both calf thymus DNA and d(ATGCAT){sub 2}. Although they were able to infer the identify of the two species on d(ATGCAT){sub 2}, definitive experimental evidence was lacking. They have designed and synthesized a self-complementary 12-mer (d(CICGAATTCICG){sub 2}) based on the Dickerson dodecamer (d(CGCGAATTCGCG){sub 2}) that bonds identically two tomaymycin molecules, each having a defined orientation and stereochemistry. The results presented in this study together with previous investigations show that the orientation of the drug molecule in the minor groove, and stereochemistry at the covalent linkage site, is dependent upon both the flanking sequence and drug structure. This conclusion mandates caution be used in rationalizing the biochemical and and biological effects of P(1,4)B bonding to DNA until precise structural information is established.