HUMAN DNA TOPOISOMERASE II-ALPHA-DEPENDENT DNA CLEAVAGE AND YEAST CELL-KILLING BY ANTHRACYCLINE ANALOGS

Anthracyclines are among the most clinically useful topoisomerase II p oisons. A complete understanding of their molecular mechanism is thus fundamental for a rational design of novel agents. We evaluated four a nthracycline analogues with respect to human topoisomerase II alpha-de pendent DNA cleaving activity, efficiency in killing yeast cells, and uptake and retention in yeast and compared the yeast system to tumor c ell line models. The least JN394top2-4 strain was used because it has a topoisomerase II ts gene mutation: enzyme activity is much less at 3 0 degrees C than at 25 degrees C and is completely lost at 35 degrees C. Untransformed JN394top2-4 cells were 33-fold more sensitive to idar ubicin at 25 degrees C than at 30 degrees C, showing that topoisomeras e II is the primary drug target. Overexpression of human topoisomerase II alpha was toxic to yeast cells when the yeast enzyme was inactivat ed. Drug-dependent killing of yeast cells expressing low levels of the human alpha isoenzyme at 35 degrees C showed that the analogues spann ed a 3-log range of cytotoxic potency in yeast, as they did in tumor c ells. However, the compounds were much less active against the yeast s train than mammalian tumor cell lines. Drug uptake was determined and found to be altered in yeast with respect to tumor cells. Although DNA cleavage stimulated by anthracyclines roughly correlated with cytotox icity, the cleavage level:cytotoxicity ratios were different for the s tudied drugs. Thus, the results suggest that other drug-dependent mole cular factors contribute to drug activity in addition to the cellular content of topoisomerase II alpha and drug uptake.