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

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.