Effects of MDR1 1236C > T-2677G > T-3435C > T polymorphisms on the intracellular accumulation of tacrolimus, cyclosporine A, sirolimus and everolimus

Abstract

1. Overexpression of P-glycoprotein (P-gp, encoded by MDR1) mediates resistance to multiple immunosuppressors. Several common MDR1 variants (1236C > T, 2677G > T, 3435C > T) impact the efflux activity of P-gp-mediated substrates. We assessed the effect of these polymorphisms on the sensitivity, intracellular accumulation, and efflux of tacrolimus, cyclosporine A, sirolimus and everolimus in transfected LLC-PK1 cells. 2. LLC-PK1 cell lines were transfected with empty vector (pcDNA3.1) and recombinant MDR1_T-T-T, MDR1_C-T-T, MDR1_C-G-T and MDR1_C-G-C vectors, respectively and further screened in the presence of puromycin. The IC₅₀ values, intracellular accumulation, and apparent permeability ratios of tacrolimus, cyclosporine A, sirolimus and everolimus were evaluated. 3. MDR1 overexpression increased the resistance of LLC-PK1 cells to tacrolimus, cyclosporine A, sirolimus and everolimus. The resistance of cells expressing MDR1_C-G-C wild-type haplotypes to tacrolimus were increased compared to MDR1_T-T-T, MDR1_C-T-T, MDR1_C-G-T variant haplotypes. The efflux ability of P-gp-mediated tacrolimus in cells transfected with MDR1_C-G-C was higher than cells overexpressing MDR1_T-T-T, MDR1_C-T-T, MDR1_C-G-T variant haplotypes. In addition, the resistance of cells expressing MDR1_C-G-C wild-type haplotypes to sirolimus were increased compared to MDR1_C-T-T, MDR1_C-G-T variant haplotypes. The efflux ability of P-gp-mediated sirolimus in cells overexpressing MDR1_C-G-C was higher than cells transfected with MDR1_C-T-T, MDR1_C-G-T variant haplotypes. 4. These findings indicate that wild-type MDR1 exports tacrolimus and sirolimus more efficiency than the MDR1_T-T-T, MDR1_C-T-T, MDR1_C-G-T variant protein. This observation indicates that 1236C > T, 2677G > T, 3435C > T variant haplotypes drastically decrease the efflux ability of P-gp-mediated tacrolimus and sirolimus in a substrate-specific manner.

Keywords: MDR1 polymorphisms; accumulation; efflux; immunosuppressor; resistance.