Verapamil stimulates glutathione transport by the 190-kDa multidrug resistance protein 1 (MRP1).

Abstract

Multidrug resistance in tumor cells is often associated with reduced drug accumulation resulting from increased expression of the 190-kDa multidrug resistance protein 1 (MRP1) or the 170-kDa P-glycoprotein. However, unlike P-glycoprotein, MRP1 is a primary active transporter of many conjugated organic anions, including the cysteinyl leukotriene LTC(4). Moreover, agents such as verapamil that reverse P-glycoprotein-mediated resistance are often poorly, or not at all, effective in MRP1-overexpressing cells. In the present study, we investigated the effects of verapamil on MRP1-mediated transport processes. We found that verapamil inhibited LTC(4) transport into inside-out membrane vesicles prepared from MRP1-transfected cells in a competitive manner, but only in the presence of reduced glutathione (GSH) or its nonreducing S-methyl derivative. In the presence of 1 mM GSH, the apparent K(i) for verapamil was 1.2 microM, and in the presence of 100 microM verapamil, the apparent K(i) for GSH was 77 microM. Verapamil itself was not transported by MRP1 in either intact cells or membrane vesicles. However, verapamil strongly stimulated MRP1-mediated GSH uptake by membrane vesicles in a concentration-dependent and osmotically sensitive manner that was inhibitable by MRP1-specific monoclonal antibodies. In the presence of 100 microM verapamil, the apparent K(m) and V(max) for GSH uptake were 83 microM and 55 pmol mg(-1) min(-1), respectively. It is proposed that the variable ability of verapamil to modulate MRP1-mediated resistance in different cell lines may be more closely linked to its effect on the GSH status of the cells than on its ability to inhibit the MRP1 transporter itself.