A human lymphocyte based ex vivo assay to study the effect of drugs on P-glycoprotein (P-gp) function.
OBJECTIVE
The effect of drugs on P-glycoprotein (P-gp) is normally studied in transfected or overexpressing cell lines derived from tumor cells or animal 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, vinblastine) and of other transporters (indomethacin, probenecid, sulfinpyrazone) on intracellular accumulation of rhodamine 123 was evaluated by flow cytometry.


RESULTS
The efflux of rhodamine 123 was inhibited by P-gp inhibitors in a saturable, concentration-dependent manner. The potency of inhibition of P-gp was cyclosporine > tacrolimus > quinidine > verapamil > vinblastine. Vinblastine inhibited P-gp at lower concentrations, whereas at high concentrations, there was an activation of rhodamine 123 efflux from lymphocytes. The multidrug resistance associated protein (MRP) inhibitors, sulfinpyrazone and probenecid, did not have any significant effect on intracellular accumulation of rhodamine 123, but indomethacin caused a concentration-dependent increase in retention of rhodamine 123, indicating the involvement of other uncharacterized transporters.


CONCLUSIONS
Lymphocytes can serve as a model tissue for studying modulation of P-gp activity by drugs. Both inhibitors and inducers of P-gp activity can be evaluated.
Parasrampuria
D A
DA
Department of Biopharmaceutical Sciences, School of Pharmacy, University of California, San Francisco 94143-0446, USA.
Lantz
M V
MV
Benet
L Z
LZ
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D002469:Cell Separation; D016572:Cyclosporine / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005456:Fluorescent Dyes / Q000378:metabolism; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D008214:Lymphocytes / Q000187:drug effects* / Q000378:metabolism; D011802:Quinidine / Q000494:pharmacology; D020112:Rhodamine 123 / Q000378:metabolism; D016559:Tacrolimus / Q000494:pharmacology; D014700:Verapamil / Q000494:pharmacology
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D002121:Calcium Channel Blockers; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D007166:Immunosuppressive Agents; D020112:Rhodamine 123; D016572:Cyclosporine; D014700:Verapamil; D011802:Quinidine; D016559:Tacrolimus