ABCC2 (1249G > A) polymorphism implicates altered transport activity for sorafenib.

1. Multidrug resistance-associated protein 2 (MRP2), encoded by the ABCC2 gene, is an efflux transporter of several endogenous substrates and xenobiotics. Here, we investigated whether the 1249G > A (rs2273697) polymorphism in ABCC2 affects the ability of MRP2 to pump the multi-tumor drug sorafenib out of cells. 2. Human embryonic kidney 293 (HEK 293) cell lines transfected with ABCC2-1249G and ABCC2-1249A were used to assess the sensitivity and accumulation to sorafenib. The isolated MRP2 were applied to estimate the ATPase activity. 3. The HEK293 cell line overexpressing the ABCC2 1249A allele showed a significantly higher 50% inhibitory concentration (IC50) than a cell line overexpressing ABCC2-1249G or a non-overexpressing control cell line. Intracellular accumulation of sorafenib was much lower in ABCC2-1249A cells than in ABCC2-1249G cells expressing comparable levels of MRP2. Isolated ABCC2-1249A protein showed higher ATPase activity than ABCC2-1249G protein. 4. Our results suggest that the ABCC2 polymorphism 1249G > A increases the ATPase activity of MRP2, leading to greater efflux of sorafenib.

Impacts of ABCB1 (G1199A) polymorphism on resistance, uptake, and efflux to steroid drugs.

1. P-glycoprotein (P-gp) substrates, including steroid drugs, involve in the inter-individual differences in resistant phenotype. This study was performed to evaluate whether G1199A polymorphism in ABCB1 gene can alter the sensitivity, accumulation, and transepithelial efflux to steroids in LLC-PK1 cells. 2. The stable recombinant LLC-PK1 cell lines transfected with ABCB1 1199G and ABCB1 1199A were used to assess the sensitivity, accumulation, and transepithelial permeability to steroids. 3. The cells transfected with 1199A allele displayed stronger resistance to aldosterone, dexamethasone, and cortisol (2.5-, 2.0-, and 1.6-fold, respectively) than cells overexpressing 1199G allele, while the two types of recombinant cells showed a similar resistance to corticosterone. The accumulation of aldosterone, dexamethasone, and cortisol in recombinant 1199A cells were significantly decreased when compared to 1199G cells (2.9-, 4.4-, and 3.9-fold, respectively). The net efflux ratios of P-gp-mediated aldosterone, dexamethasone, and cortisol in cells expressing 1199A allele were apparently greater than cells transfected with 1199G allele (3.3-, 3.5-, and 4.0-fold, respectively). 4. The impacts of ABCB1 (G1199A) single nucleotide polymorphism on the efflux of P-gp substrates presented as drug-specific. Overall, the transport ability of P-gp-dependent steroid drugs in recombinant model overexpressing variant 1199A allele is stronger in comparison to cells overexpressing wild-type 1199G allele. Therefore, the ABCB1 (G1199A) polymorphism may affect effective steroids concentration in target cells by regulating the drug transport and distribution.

Effects of the ABCB1 (1199G > A) Polymorphism on Steroid Sex Hormone-Induced P-Glycoprotein Expression, ATPase Activity, and Hormone Efflux.

This study examined how the 1199G > A polymorphism in the ABCB1 gene encoding P-glycoprotein (P-gp) affects the protein's expression, ATPase activity, and ability to pump female steroid sex hormones out of LLC-PK1 cells. The ABCB1 (1199G) or ABCB1 (1199A) allele was transfected into cells, which were incubated for 48 h with various hormone concentrations, then analyzed by Western blotting to examine expression of P-gp protein and by reverse transcription-polymerase chain reaction (RT-PCR) to examine expression of mRNA. Cells were also compared in terms of their transepithelial permeability to steroid sex hormones in the presence and absence of the specific P-gp inhibitor GF120918. P-gp ATPase activity induced by steroid sex hormones was also assayed. Estriol and ethynyl estradiol up-regulated levels of ABCB1 mRNA in a concentration-dependent manner, with ABCB1 (1199A) mRNA showing greater up-regulation than ABCB1 (1199G) mRNA. Estrone, estriol, and ethynyl estradiol were substrates of both types of P-gp in transepithelial permeability assays, and the ABCB1 (1199A) protein showed a significantly higher net efflux ratio for estrone (13.4 vs. 7.4, p < 0.005), estriol (5.6 vs. 3.3, p < 0.05), and ethynyl estradiol (12.7 vs. 5.3, p < 0.005). Induction of P-gp ATPase activity by ethynyl estradiol and progesterone increased with increasing hormone concentration, and the magnitude of stimulation was greater for ABCB1 (1199A) P-gp than for ABCB1 (1199G) P-gp. These results indicate that the ABCB1 (1199G > A) polymorphism influences steroid sex hormone-induced expression and function of P-gp, which may help to explain inter-patient differences in P-gp-mediated chemotherapy resistance in vivo.