Role of P-glycoprotein in cyclosporine cytotoxicity in the cyclosporine-sirolimus interaction.
Cyclosporine nephrotoxicity remains a major side effect in solid organ transplantation, and can be exacerbated by concomitant administration of sirolimus. Cyclosporine and sirolimus are P-glycoprotein (Pgp) substrates. We hypothesized that the Pgp activity level may affect cyclosporine cytotoxicity by interfering with the ability of Pgp to remove cyclosporine from within tubular cells, and that an interaction between cyclosporine and sirolimus on Pgp function may explain the enhancement of cyclosporine nephrotoxicity by sirolimus. Cyclosporine cytotoxicity was evaluated in primary cultures of normal human renal epithelial cells (HRECs) by cell viability and cytotoxicity assays. Verapamil, quinine, PSC833, and PGP-4008 were used as Pgp inhibitors. Rhodamine-123 (R-123), a fluorescent substrate of Pgp, was used to assess Pgp-mediated transport. Cellular cyclosporine concentration was measured by high-performance liquid chromatography coupled to tandem mass spectrometry. Pgp expression and function were confirmed in HRECs and cyclosporine and sirolimus were shown to be Pgp inhibitors in this model. Verapamil-induced inhibition of Pgp led to a significant increase in cellular concentration of cyclosporine (P<0.05). Cyclosporine exerted a concentration-dependent cytotoxic effect on HRECs that was significantly increased by inhibition of Pgp activity. Sirolimus exerted an inhibitory effect on R-123 efflux in HRECs and increased cellular cyclosporine concentrations in a dose-dependent manner. These data demonstrate that Pgp plays a critical role in protecting renal epithelial cells from cyclosporine toxicity. The inhibitory effect of sirolimus on Pgp-mediated efflux and the cellular concentration of cyclosporine could explain the exacerbation of cyclosporine nephrotoxicity observed clinically.
Anglicheau
D
D
INSERM, U775, Université René Descartes, Paris, France. dany.anglicheau@univ-paris5.fr
Pallet
N
N
Rabant
M
M
Marquet
P
P
Cassinat
B
B
Méria
P
P
Beaune
P
P
Legendre
C
C
Thervet
E
E
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000083:Acetanilides / Q000494:pharmacology; D018929:Cell Culture Techniques; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D016572:Cyclosporine / Q000633:toxicity*; D003524:Cyclosporins / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D005456:Fluorescent Dyes; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D007668:Kidney / Q000166:cytology; D011758:Pyrroles / Q000494:pharmacology; D011803:Quinine / Q000494:pharmacology; D011804:Quinolines / Q000494:pharmacology; D012235:Rhodamines; D020123:Sirolimus / Q000494:pharmacology*; D014700:Verapamil / Q000494:pharmacology
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000083:Acetanilides; D003524:Cyclosporins; D005456:Fluorescent Dyes; D007166:Immunosuppressive Agents; C483891:N-1-benzyl-2,3-dihydro-1H-pyrrolo(2,3-b)quinolin-4-yl-2-phenylacetamide; D011758:Pyrroles; D011804:Quinolines; D012235:Rhodamines; D016572:Cyclosporine; D011803:Quinine; D014700:Verapamil; C070272:valspodar; D020123:Sirolimus