Multidrug Resistance-associated Protein Gene Overexpression and Reduced Drug Sensitivity of Topoisomerase II in a Human Breast Carcinoma MCF7 Cell Line Selected for Etoposide Resistance

A human breast cancer cell line (MCF7/WT) was selected for resistance to etoposide (VP-16) by stepwise exposure to 2-fold increasing concentrations of this agent. The resulting cell line (MCF7/VP) was 28-, 21-, and 9-fold resistant to VP-16, VM-26, and doxorubicin, respectively. MCF7/VP cells also exhibited low-level cross-resistance to 4'-(9-acridinylamino)methanesulfon-m-anisidide, mitoxantrone, and vincristine and no crossresistance to genistein and camptothecin. Furthermore, these cells were collaterally sensitive to the alkylating agents melphalan and chiorambucil. DNA topoisomerase II levels were similar in both wild-type MCF7/WT and drug-resistant MCF7/VP cells. In contrast, topoisomerase II from MCF7/VP cells appeared to be 7-fold less sensitive to drug-induced cleavable complex formation in whole cells and 3-fold less sensitive in nuclear extracts than topoisomerase II from MCF7/WT cells. Although this suggested that the resistant cells may contain a qualitatively altered topoisomerase II, no mutations were detected in either the ATP-binding nor the putative breakage/resealing regions of either DNA topoisomerase IIot or II~l. In addition, the steady-state intracellular VP-16 concentration was reduced by 2-fold in the resistant cells, in the absence of detectable mdrl/ P-gp expression and without any change in drug efflux. In contrast, expression of the gene encoding the MRP was increased at least 10-fold in resistant MCF7/VP cells as compared to sensitive MCF7/WT cells. These results suggest that resistance to epipodophyllotoxins in MCF7/VP cells is multifactorial, involving a reduction in intracellular drug concentration, possibly as a consequence of MRP overexpression, and an altered DNA topoisomerase II drug sensitivity.