INHIBITION OF DNA TOPOISOMERASES I AND II BY DMP840

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DMP 840 is one of a new class of antitumor agents, bis-naphthalimides, in Phase I clinical trials. In preclinical studies DMP 840 has produced complete regressions at nontoxic concentrations in 5/6 rhabdomyosarcomas grown as xenografts in immune-deprived mice. Our long range goal is to determine the efficacy of DMP 840 in treating pediatric patients with solid tumors. Preliminary biochemical data indicate that DMP 840 is a novel compound in that it inhibits DNA topoisomerases I and II by a mechanisms different from inhibitors of topoisomerases now used clinically. Because the mechanisms of inhibition of topoisomerases by DMP 840 at the molecular level is likely to be a factor in determining the tumor types against which DMP 840 is active and the drugs with which it can be combined effectively, we propose to determine the mechanism of inhibition of topoisomerases I and II by DMP 840. Based on preliminary data, our hypothesis is that DMP 840 prevents the binding of topoisomerases I and II to DNA. If this hypothesis is correct, the cytotoxicity of DMP 840 may be cell cycle-independent, and important factor in treating solid tumors which frequently have low growth fractions. We will use cell lines established from solid tumors having different sensitivities to DMP 840 to examine the effect of this compound on the function of topoisomerases in intact cells. We will also attempt to correlate drug potency with topoisomerase I/II levels, with quantitation on a single cell basis by fluorescence digital imaging microscopy. Isolated enzymes will then be used to determine which step in the catalytic cycle of topoisomerases is inhibited by DMP 840 and effect of DMP 840 on the physical interaction of the compound with DNA, enzyme, or DNA/enzyme complex. These data will then be used for molecular modeling predictions of DNA/drug/enzyme interaction. Additionally, DMP 840 appears to be relatively nontoxic in animal models. However, topoisomerase inhibitors are known to be mutagenic. Therefore, the mutagenic potential of DMP 840 will be evaluated. Our working hypothesis is that because DMP 840 has more than one cellular target, simultaneous mutations would be required in each of these targets to permit cell survival; therefore, DMP 840 may be less mutagenic than topoisomerase inhibitors currently approved for clinical use. Throughout these experiments, we will compare DMP with agents that inhibit either topoisomerase I or II to establish the relative efficacy and toxicity of the three classes of compounds.