ALTERED DNA TOPOISOMERASE-II IN MULTIDRUG-RESISTANCE

The characteristic feature of multidrug resistance (MDR) associated wi th drugs that interact with DNA topoisomerase II (topo II) is alterati ons in topo II activity or amount (at-MDR). We have characterized the at-MDR phenotype in human leukemic CEM cells selected for resistance t o the topo II inhibitor, VM-26. Compared to drug-sensitive cells, the key findings are that at-MDR cells exhibit (i) decreased topo II activ ity; (ii) decreased drug sensitivity, activity and amount of nuclear m atrix topo II; (iii) increased ATP requirement of topo II; (iv) a sing le base mutation in topo II resulting in a change of Arg to Gln at pos ition 449, at the start of the motif B/nucleotide binding site; and (v ) decreased topo II phosphorylation, suggesting decreased kinase or in creased phosphatase activities. Recent results using single-stranded c onformational polymorphism analysis reveals the presence of a mutation in the motif B/nucleotide binding site of the topo IIalpha gene in CE M at-MDR cells and in another leukemic cell line selected for resistan ce to m-AMSA. Finally, we have observed marked changes in the nuclear distribution of topo II in cells treated with anti-topo II drugs and h ave also found these changes to be attenuated in drug-resistant cells. We postulate that traditional inhibitors of topo II alter the equilib rium of the strand-passing reaction such that the number of enzyme-DNA covalent complexes increases. We further suggest that when the enzyme is bound to DNA it is protected from proteolysis, thus allowing more topo II molecules to be detected. We propose that MDR associated with alterations in topo II may have clinical consequences, and our current efforts involve exploiting these biochemical and molecular observatio ns in the development of probes that may be useful to identify such dr ug resistant cells in the tumors of patients.