Antitumor Agent Merbarone and Intracellular Inhibition of Topoisomerase II by the In Vitro

Merbarone has previously been shown to have antitumor activity of unknown mechanism in P38S and 1,1210 tumor models (A. D. Brewer et a/., Biochem. Pharmacol., 34:2047-2050,1985) and is currently undergo ing Phase I clinical trials. Here we report that merbarone is an inhibitor of topoisomerase II. Merbarone inhibited purified mammalian topoisomerase II with a 50% inhibitory concentration of 20 MM.as assessed by ATP-dependent unknotting of P4 phage DNA or relaxation of supercoiled pBR322 plasmid. In contrast to the type II enzyme, inhibition of catalytic activity of topoisomerase I required about 10-fold higher concentrations of merbarone, with a 50% inhibitory concentration of approximately 200 MM.Unlike epipodophyllotoxin analogues and certain DNA intercalative agents which stabilize the topoisomerase II-DNA "cleavable complex," merbarone did not cause detectable topoisomerase II-induced DNA cleav age. Furthermore, merbarone inhibited the production by amsacrine or teniposide of topoisomerase Il-associated DNA strand breaks; under identical conditions novobiocin did not decrease these breaks, setting merbarone apart from a novobiocin-like class of topoisomerase II inhib itor. In LI 210 cells, merbarone produced only small numbers of proteinassociated DNA strand breaks, and only at very high concentrations. Merbarone reduced in a concentration-dependent manner the number of amsacrineor teniposide-stimulated protein-associated DNA strand breaks in LI 210 cells or their isolated nuclei. The data suggest that merbarone represents a novel type of topoisomerase II inhibitor.