C1027 CHROMOPHORE, A POTENT NEW ENEDIYNE ANTITUMOR ANTIBIOTIC, INDUCES SEQUENCE-SPECIFIC DOUBLE-STRAND DNA CLEAVAGE

C1027, a new macromolecular antitumor antibiotic produced by a Streptomyces strain, shows highly potent cytotoxicity to cultured cancer cells and marked DNA cleaving ability. The structure of its chromophore, responsible for most of the biological activities of the antibiotic, was recently determined and found to contain a nine-membered enediyne. In contrast to other enediyne antibiotics, such as neocarzinostatin, calicheamicin, esperamicin, and recently found kedarcidin, C1027 damages duplex DNA even in the absence of thiols. The DNA damage caused by C1027 includes double-strand breaks, single-strand breaks, and abasic sites. Experiments with plasmid DNA and P-32-end-labeled restriction fragments demonstrated that the chromophore, extracted from the protein-containing holoantibiotic, interacts in the DNA minor groove and cleaves double-helical DNA with a remarkable sequence-selectivity causing direct double-strand breaks. The double-strand cleavage sites, occurring predominantly at CTT (T) under bar T/AAA (A) under bar G, ATA (A) under bar T/ATT (A) under bar T CTT (T) under bar A/TAA (A) under bar G, CTC (T) under bar T/AAG (A) under bar G, and especially GTT (A) under bar T/ATA (A) under bar C, consist of five nucleotide sequences with a two-nucleotide 3'-stagger of the cleaved residues (cutting sites are underlined). The chemical structures of the damaged residues at the GTT (A) under bar T/ATA (A) under bar C cleavage site suggest a model in which a C1027-induced double-strand break results from abstraction, by a single molecule of the diradical form of the chromophore, of a C4' hydrogen atom from the A residue of GTT (A) under bar T and a C5' hydrogen atom from the A of ATA (A) under bar C on the opposite strand. Single-strand breaks, which are mainly produced at adenylate and thymidylate residues, appear to be separate events presumably resulting from different binding modes of the drug to DNA. The highly sequence-selective double-strand breaks induced by C1027 chromophore and their resistance to repair may account for the potent cytotoxicity of the antibiotic.