A human lymphocyte based ex vivo assay to study the effect of drugs on P-glycoprotein (P-gp) function

Purpose. The effect of drugs on P-glycoprotein (P-gp) is normally studied i n transfected or overexpressing cell lines derived from tumor cells or anim al tissue. We wanted to develop an assay using normal healthy human tissue to study and characterize the drug-transporter interaction. Methods. Lymphocytes were isolated from healthy human blood. The effect of inhibitors of P-gp (cyclosporine, tacrolimus, verapamil, quinidine, vinblas tine) and of other transporters (indomethacin, probenecid, sulfinpyrazone) on intracellular accumulation of rhodamine 123 was evaluated by flow cytome try. Results. The efflux of rhodamine 123 was inhibited by P-gp inhibitors in a saturable, concentration-dependent manner. The potency of inhibition of P-g p was cyclosporine > tacrolimus > quinidine > verapamil > vinblastine. Vinb lastine inhibited P-gp at lower concentrations, whereas at high concentrati ons, there was an activation of rhodamine 123 efflux from lymphocytes. The multidrug resistance associated protein (MRP) inhibitors, sulfinpyrazone an d probenecid, did not have any significant effect on intracellular accumula tion of rhodamine 123, but indomethacin caused a concentration-dependent in crease in retention of rhodamine 123, indicating the involvement of other u ncharacterized transporters. Conclusions. Lymphocytes can serve as a model tissue for studying modulatio n of P-gp activity by drugs. Both inhibitors and inducers of P-gp activity can be evaluated.