| Abstract
| - Despite the prevalence of topoisomerase II-targeted drugs in cancer chemotherapy and theimpact of drug resistance on the efficacy of treatment, interactions between these agents and topoisomeraseII are not well understood. Therefore, to further define interactions between anticancer drugs and the typeII enzyme, a nitrocellulose filter assay was used to characterize the binding of etoposide to yeasttopoisomerase II. Results indicate that etoposide binds to the enzyme in the absence of DNA. The apparentKd value for the interaction was ∼5 μM drug. Etoposide also bound to ytop2H1012Y, a mutant yeasttype II enzyme that is ∼3−4-fold resistant to etoposide. However, the apparent Kd value for the drug(∼16 μM) was ∼3 times higher than that determined for wild-type topoisomerase II. Although it hasbeen widely speculated that resistance to topoisomerase II-targeted anticancer agents results from adecreased drug−enzyme binding affinity, these data provide the first direct evidence in support of thishypothesis. Finally, the ability of yeast topoisomerase II to bind etoposide was dependent on the presenceof the hydroxyl moiety of Tyr783, suggesting specific interactions between etoposide and the active siteresidue that is involved in DNA scission.
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