title: Competitive inhibition by verapamil of ATP-dependent high affinity vincristine binding to the plasma membrane of multidrug-resistant K562 cells without calcium ion involvement.
abstract: Verapamil, a calcium channel blocker, can inhibit the efflux of antitumor agents from multidrug-resistant cells and reverse drug resistance. We have recently reported that the plasma membrane prepared from an adriamycin (ADM)-resistant variant (K562/ADM) of human myelogenous leukemia K562 cells showed ATP/Mg2+-dependent high affinity binding of vincristine (VCR), which is closely related to the drug transport mechanism in this cell line. To clarify how calcium channel blockers inhibit the transport of antitumor agents from the resistant cells, we analyzed the effect of calcium channel blockers and Ca2+ ion on the VCR binding to K562/ADM plasma membrane. The ATP-dependent VCR binding was inhibited by calcium channel blockers (verapamil, nicardipine, and diltiazem), which are known to inhibit drug efflux from the resistant cells. Addition of [ethylenebis(oxyethylenenitrilo)]tetraacetic acid or high concentration of Ca2+ decreased the amount of VCR binding to some extent; however, a substantial amount of VCR still could bind to K562/ADM plasma membrane. The inhibitory effect of verapamil on the VCR binding was observed regardless of the Ca2+ concentration. Klotz plot analysis revealed that the inhibition of the VCR binding to K562/ADM plasma membrane by verapamil was competitive. Dissociation constant (Kd) of VCR and apparent inhibitory constant (Kiapp) of verapamil were calculated to be 0.1 +/- 0.1 microM (SD) and 1 +/- 1 microM, respectively. These results indicate that Ca2+ ion is not required for the VCR binding and that verapamil competitively inhibits the VCR binding without concerning Ca2+ ion. Antitumor agents (vinblastine, actinomycin D, ADM, and colchicine) and other agents known to reverse multidrug resistance (nicardipine, diltiazem, cyclosporin A, quinidine, and trifluoperazine) also inhibited the VCR binding competitively.
authors.affiliation: Cancer Chemotherapy Center, Japanese Foundation for Cancer Research, Tokyo.
authors.forename: M
authors.initials: M
authors.lastname: Naito
authors.forename: T
authors.initials: T
authors.lastname: Tsuruo
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology*; D001667:Binding, Competitive; D002118:Calcium / Q000502:physiology*; D002462:Cell Membrane / Q000378:metabolism; D004317:Doxorubicin / Q000494:pharmacology; D004351:Drug Resistance; D007951:Leukemia, Myeloid / Q000378:metabolism; D014700:Verapamil / Q000378:metabolism / Q000494:pharmacology*; D014750:Vincristine / Q000378:metabolism*
chemical_list: D014750:Vincristine; D004317:Doxorubicin; D000255:Adenosine Triphosphate; D014700:Verapamil; D002118:Calcium