Interaction of cyclosporin derivatives with the ATPase activity of human P‐glycoprotein

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‐Bmt]‐[Val*]‐cyclosporin (PSC 833), on the adenosine triphosphatase (ATPase) activity of human P‐glycoprotein. 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. 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. Kinetic analysis indicated a competitive inhibition of verapamil‐stimulated ATPase by PSC 833. The binding of 8‐azido‐[α‐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. 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. British Journal of Pharmacology (1997) 122, 241–248; doi:10.1038/sj.bjp.0701377