The cytotoxicity of (−)-lomaiviticin A arises from induction of double-strand breaks in DNA

Abstract

The metabolite (–)-lomaiviticin A, which contains two diazotetrahydrobenzo[b]fluorene (diazofluorene) functional groups, inhibits the growth of cultured human cancer cells at nanomolar–picomolar concentrations; however, the mechanism responsible for the potent cytotoxicity of this natural product is not known. Here we report that (–)-lomaiviticin A nicks and cleaves plasmid DNA by an ROS- and iron-independent pathway and that the potent cytotoxicity of (–)-lomaiviticin A arises from induction of DNA double-strand breaks (dsbs). In a plasmid cleavage assay, the ratio of single-strand breaks (ssbs) to dsbs is 5.3±0.6:1. Labeling studies suggest this cleavage occurs via a radical pathway. The structurally related isolates (–)-lomaiviticin C and (–)-kinamycin C, which contain one diazofluorene, are demonstrated to be much less effective DNA cleavage agents, thereby providing an explanation for the enhanced cytotoxicity of (–)-lomaiviticin A compared to other members of this family.

Structural basis for DNA cleavage by the potent antiproliferative agent (–)-lomaiviticin A

“…Prior cell free and tissue culture studies of (-)-lomaiviticin A (1) have established that its potent cytotoxic effects derive from the induction of DSBs in DNA (18,19). Deuterium-labeling experiments are consistent with a mechanism involving the generation of vinyl radical intermediates at each diazo carbon of 1, followed by hydrogen atom abstraction from DNA (18).…”

“…The molecular mechanism of DNA DSB induction by (-)-lomaiviticin A (1) has been studied (18). Cell-free deuteriumlabeling experiments are consistent with a pathway comprising reductive activation of one diazofluorene (Fig.…”

“…The RMSD between the annealed complex and the molecular dynamics-minimized structure, with the distance constraints removed, is 3.1 Å and contains changes to areas with large degrees of freedom, such as the oleandrose residues and the nucleotides that have been twisted out of the duplex. A single-point calculation reveals the bound conformation of 1 as 0.40 kcal/mol higher in energy than that of the minimized structure in solution (18).…”

“…(-)-Lomaiviticin C (3) and (-)-kinamycin C (4) are several orders of magnitude less potent, whereas (-)-lomaiviticin B (2) is ∼10-100-fold less potent. The cytotoxicity of 1 derives from the induction of double-strand breaks (DSBs) in DNA (18,19). K562 cells exposed to 5 nM of 1 for 30 min accumulated DSB levels that were comparable to 40 Gy of ionizing radiation.…”

Properties and reactivity of nucleic acids relevant to epigenomics, transcriptomics, and therapeutics

“…86,87 The most prominent of these is the kinamycin and lomaiviticin family of natural products. These, however, seem to operate by radical-induced strand-breaking 88 and hence will not be discussed here. The diazopeptides, exemplified in Fig.…”