EVIDENCE FOR CYTOPLASMIC P-GLYCOPROTEIN LOCATION ASSOCIATED WITH INCREASED MULTIDRUG-RESISTANCE AND RESISTANCE TO CHEMOSENSITIZERS

A new human myeloma cell line, 8226/MDR(10)V, was selected from a P-gl ycoprotein-positive cell line, 8226/Dox(40), in the continuous presenc e of doxorubicin and verapamil. MDR(10)V cells are 13-fold more resist ant to doxorubicin and 4-fold more resistant to vincristine than the p arent cell line, Dox(40). Chemosensitizers are also less effective in reversing resistance in the MDR(10)V compared to the Dox(40) cells. De spite higher resistance to cytotoxic agents, MDR(10)V expresses 40% le ss P-glycoprotein in the plasma membrane compared to Dox(40); however, total cellular P-glycoprotein is the same in both cell lines. Confoca l immunofluorescence microscopy shows 2.5-fold more P-glycoprotein in the cytoplasm of MDR(10)V cells as compared to Dox(40) cells. The cyto plasmic location of P-glycoprotein in the MDR(10)V cells is associated with a redistribution of doxorubicin. In Dox(40) cells, doxorubicin i s concentrated in the nucleus, whereas in MDR(10)V cells, 90% of doxor ubicin is found in the cytoplasm. In the presence of equivalent intrac ellular doxorubicin, there was a decrease in DNA-protein crosslinks in the MDR(10)V cell line compared to the Dox(40) cell line. This findin g is in agreement with the intracellular doxorubicin fluorescence stud ies showing less doxorubicin in the nuclei of MDR(10)V cells compared to Dox(40) cells. Verapamil is less effective in increasing doxorubici n accumulation in the nuclei of MDR(10)V cells compared to Dox(40) cel ls. Processing of P-glycoprotein from the endoplasmic reticulum to the medial Golgi was identical between the two cell lines as determined b y endoglycosidase H sensitivity of newly sensitized P-glycoprotein. No mutations were found in MDR1 cDNA from MDR(10)V cells compared to Dox (40) cells. These results suggest that resistance to chemosensitizing agents plus cytotoxic drugs is associated with a redistribution of P-g lycoprotein from the plasma membrane to the cytoplasm, which in turn r educes the amount of cytotoxic drug reaching the nucleus.