ACTIVE EFFLUX OF THE FREE ACID FORM OF THE FLUORESCENT DYE 2',7'-BIS(2-CARBOXYETHYL)-5(6)-CARBOXYFLUORESCEIN IN MULTIDRUG-RESISTANCE-PROTEIN-OVEREXPRESSING MURINE AND HUMAN LEUKEMIA-CELLS

Murine and human cell lines overexpressing the multidrug-resistance pr otein (MRP) showed a marked decreased accumulation of the fluorescent dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF). In cont rast, less altered accumulation was seen in the P-glycoprotein(P-gp)-o verexpressing cell lines. The decreased drug accumulation was reversed by the energy inhibitors sodium azide/2-deoxyglucose and by the vinca alkaloid, vincristine, but not by the chemotherapeutic agents, etopos ide and adriamycin. Decreased accumulation was linked to active efflux of the hydrophilic free acid form of BCECF from the MRP-overexpressin g cell lines, indicating that dye extrusion occurs after the dye ester has been converted to the free acid form in the cytoplasm. The findin g suggests that MRP mediates removal of substrates from a cytoplasmic location. Buthionine sulfoximine (BSO), an inhibitor of glutathione sy nthesis, decreased the vincristine and etoposide resistance displayed by the MRP-expressing murine cell lines, but did not affect the accumu lation of BCECF. Thus, while glutathione may be involved in MRP-mediat ed resistance to some chemotherapeutic agents, it is not necessary for efflux of substrates such as BCECF.