title: Interaction of cyclosporin derivatives with the ATPase activity of human P-glycoprotein.
abstract: 1. P-glycoprotein, a 170-180 kDa membrane glycoprotein that mediates multidrug resistance, hydrolyses ATP to efflux a broad spectrum of hydrophobic agents. In this study, we analysed the effects of three MDR reversing agents, verapamil, cyclosporin A and [3'-keto-Bmt1]-[Val2]-cyclosporin (PSC 833), on the adenosine triphosphatase (ATPase) activity of human P-glycoprotein. 2. P-glycoprotein was immunoprecipitated with a monoclonal antibody (MRK-16) and the P-glycoprotein-MRK-16-Protein A-Sepharose complexes obtained were subjected to a coupled enzyme ATPase assay. 3. While verapamil activated the ATPase, the cyclosporin derivatives inhibited both the substrate-stimulated and the basal P-glycoprotein ATPase. No significant difference was observed between PSC 833 and cyclosporin A on the inhibition of basal P-glycoprotein ATPase activity. PSC 833 was more potent than cyclosporin A for the substrate-stimulated activity. 4. Kinetic analysis indicated a competitive inhibition of verapamil-stimulated ATPase by PSC 833. 5. The binding of 8-azido-[alpha-32P]-ATP to P-glycoprotein was not altered by the cyclosporin derivatives, verapamil, vinblastine and doxorubicin, suggesting that the modulation by these agents of P-glycoprotein ATPase cannot be attributed to an effect on ATP binding to P-glycoprotein. 6. The interaction of the cyclosporin derivatives with ATPase of P-glycoprotein might present an alternative and/or additional mechanism of action for the modulation of P-glycoprotein function.
authors.affiliation: Research Department, Novartis Takarazuka Research Institute, Novartis Pharma, Ltd., Hyougo, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Watanabe
authors.forename: N
authors.initials: N
authors.lastname: Kokubu
authors.forename: S B
authors.initials: SB
authors.lastname: Charnick
authors.forename: M
authors.initials: M
authors.lastname: Naito
authors.forename: T
authors.initials: T
authors.lastname: Tsuruo
authors.forename: D
authors.initials: D
authors.lastname: Cohen
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism*; D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism; D000345:Affinity Labels; D001386:Azides / Q000378:metabolism; D016572:Cyclosporine / Q000494:pharmacology*; D003524:Cyclosporins / Q000494:pharmacology*; D006801:Humans; D014407:Tumor Cells, Cultured
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000345:Affinity Labels; D001386:Azides; D003524:Cyclosporins; C017040:8-azidoadenosine 5'-triphosphate; D016572:Cyclosporine; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; C070272:valspodar