ROLE OF THE VACUOLAR H-ATPASE IN DAUNORUBICIN DISTRIBUTION IN ETOPOSIDE-RESISTANT MCF7 CELLS OVEREXPRESSING THE MULTIDRUG-RESISTANCE ASSOCIATED PROTEIN()

Some multidrug-resistant cell lines efflux anticancer drugs but do not overexpress the well-known P-glycoprotein pump or PEP A 190 kDa or mu ltidrug-resistant associated protein (MRP) has been identified and des cribed as an MDR mediator, Many studies on cells overexpressing MRP an d Pgp, show a concentration of the drug inside cytoplasmic vesicles fo llowed by an exocytotic process. We studied daunorubicin (DNR) subcell ular distribution in the presence of an H+-ATPase pump inhibitor 7-chl oro-4-nitrobenz-2-oxa-1,3-diazole (NBD) and verapamil (VPL) in two hum an breast adenocarcinoma MCF7 etoposide-resistant and adriamycin-resis tant cell lines, overexpressing respectively MRP (MCF7/VP) and Pgp (MC F7/ADR). Nucleo-cytoplasmic distribution of daunorubicin was carried o ut using scanning confocal microspectrofluorometry. This technique all ows the determination of nuclear accumulation of anthracyclines. Our r esults show that NBD was able to increase the nuclear accumulation of DNR in MCF7/VP but not in MCF7/ADR cells. Similarly, NBD could reverse DNR resistance in MCF7/VP cells but had no effect on DNR cytotoxicity in MCF7/ADR cells. VPL caused a significant increase in nuclear accum ulation of DNR in MCF7/VP and MCF7/ADR cells. Incubation of MCF7/VP an d MCF7/ADR cells with VPL, increased the sensitivity of these cells. T hese data demonstrate clearly that even if vesicular sequestration can happen in cells overexpressing MRP and Pgp proteins, only the MRP pro tein is able to extrude the drug through intracellular vesicles and ef flux. In cells overexpressing Pgp, drug efflux probably takes place di rectly at the membrane level.