N-terminal and core-domain random mutations in human topoisomerase II alpha conferring bisdioxopiperazine resistance

Random mutagenesis of human topoisomerase II a cDNA followed by functional expression in yeast cells lacking endogenous topoisomerase II activity in t he presence of ICRF-187, identified five functional mutations conferring ce llular bisdioxopiperazine resistance, The mutations L169F, G551S, P592L, D6 45N, and T996L confer >37, 37, 18, 14, and 19 fold resistance towards ICRF- 187 in a 24 h clonogenic assay, respectively. Purified recombinant L169F pr otein is highly resistant towards catalytic inhibition by ICRF-187 in vitro while G551S, D645N, and T996L proteins are not. This demonstrates that cel lular bisdioxopiperazine resistance can result from at least two classes of mutations in topoisomerase II; one class renders the protein non-responsiv e to bisdioxopiperazine compounds, while an other class does not appear to affect the catalytic sensitivity towards these drugs. In addition, our resu lts indicate that different protein domains are involved in mediating the e ffect of bisdioxopiperazine compounds. (C) 2000 Federation of European Bioc hemical Societies. Published by Elsevier Science B.V. All rights reserved.