title: Optimizing chemotherapy by measuring reversal of P-glycoprotein activity in plasma membrane vesicles.
abstract: The appearance of multidrug resistance (MDR) of cancer cells is a major obstacle to successful chemotherapy. Several proteins have been identified that pump chemotherapeutic drugs out of cells, thus bringing about MDR. One representative pump is the P-glycoprotein, whose function can be inhibited by blockers (also known as reversers, modulators or chemosensitizers). In clinical application, many of these blockers are often not effective because they become bound to the plasma of the patients. The extent of plasma binding of the blocker varies in different persons and we have developed a 96-well kit to assay such inter-person differences. The assay uses membrane vesicles isolated from a human lymphoblastoid cell line (CEM Col1000). Uptake of rhodamine into the vesicles was measured with different concentrations of the blockers verapamil and XR9576 in presence of human plasma. The reverser XR9576 is nearly 30 times more effective than the classical blocker verapamil, the relevant K(m) values ranging from 2.66 to 45 nM for XR 9576 and 0.7 to 5.5 microM for verapamil. An even greater difference between these two drugs, nearly 1,000-fold, could be shown also in intact cells by calcein AM uptake experiments.
authors.affiliation: Department of Biological Chemistry, Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 91904, Israel. wdstein@vms.huji.ac.il
authors.forename: Sabine
authors.initials: S
authors.lastname: Köhler
authors.forename: Wilfred D
authors.initials: WD
authors.lastname: Stein
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000187:drug effects / Q000378:metabolism*; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000970:Antineoplastic Agents / Q000494:pharmacology*; D001667:Binding, Competitive; D001681:Biological Assay / Q000295:instrumentation / Q000379:methods; D001692:Biological Transport / Q000187:drug effects; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects; D018432:Drug Resistance, Multiple; D019008:Drug Resistance, Neoplasm; D004358:Drug Therapy / Q000295:instrumentation / Q000379:methods*; D004867:Equipment Design; D006801:Humans; D015458:Leukemia, T-Cell / Q000188:drug therapy / Q000378:metabolism*; D008565:Membrane Proteins; D011786:Quality Control; D015203:Reproducibility of Results; D012235:Rhodamines / Q000493:pharmacokinetics; D012680:Sensitivity and Specificity; D022161:Transport Vesicles / Q000187:drug effects / Q000378:metabolism; D014407:Tumor Cells, Cultured
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000970:Antineoplastic Agents; D008565:Membrane Proteins; D012235:Rhodamines; D000251:Adenosine Triphosphatases