MDR1 haplotypes significantly minimize intracellular uptake and transcellular P-gp substrate transport in recombinant LLC-PK1 cells

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Journal of Pharmaceutical Sciences


To date, research on the effect of single nucleotide polymorphisms (SNPs) on P-glycoprotein (P-gp) expression and functionality has rendered inconsistent results. This study systematically evaluates the impact of MDR1 haplotypes (1236/2677, 1236/3435, 2677/3435, 1236/2677/3435) on P-gp functionality compared to individual SNPs (1236, 2677, and 3435) in validated stable recombinant epithelial cells. Recombinant LLC-PK1 cells expressing MDR1wt or its variants were developed and validated for this purpose. Intracellular accumulation and time-dependant efflux of a P-gp substrate, Rhodamine 123 (R123, 5 μM) were evaluated in control and recombinant cells. Additionally, the transepithelial transport of R123 (1 μM) and Vinca alkaloids (5 μM) was evaluated. Except for MDR12677T and MDR11236T/2677T cells expressing MDR1 variants displayed intermediate R123 intracellular accumulation (1.5-2-fold higher) and lower effluxed R123 (10-20% vs. 52%) compared to those expressing MDR1wt. Efflux ratios across MDR1wt expressing cells were significantly larger for R123 (3.95 ± 1.1), Vinblastine (3.75 ± 0.26), and Vincristine (2.8 ± 0.29). Recombinant cells expressing MDR1 variants displayed 0%-22.7% P-gp activity (∼80%-100% efflux loss). Results suggest that MDR1 polymorphisms at the 1236, 2677, and/or 3435 positions significantly minimize P-gp functionality in vitro, the extent of which appears to be substrate dependant. © 2006 Wiley-Liss, Inc.

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