Arsenic-resistant cells (R15), derived from a human lung adenocarcinoma cell line (CL3), were 10-fold more resistant to sodium arsenite (As(III)). Because R15 cells accumulated less arsenic than parental CL3 cells, this arsenic resistance may be due to higher efflux and/or lower uptake of As(III). We therefore compared expression of the multidrug resistance-associated proteins MRP1, MRP2, and MRP3 in these two cell lines. MRP2 expression was 5-fold higher in R15 cells than in CL3 cells, whereas MRP1 and MRP3 expression levels were similar. Furthermore, verapamil and cyclosporin A, inhibitors of multidrug resistance transporters, significantly reduced the efflux of arsenic from R15. Thus, increased arsenic extrusion by MRP2 may contribute to arsenic resistance in R15 cells. We also examined the expression of several aquaglyceroporins (AQPs), which mediate As(III) uptake by cells. Little AQP7 or AQP9 mRNA was detected by reverse transcription-PCR in either cell line, whereas AQP3 mRNA expression was 2-fold lower in R15 cells than in CL3 cells. When AQP3 expression in CL3 cells was knocked down by RNA interference, CL3 cells accumulated less arsenic and became more resistant to As(III). Conversely, overexpression of AQP3 in human embryonic kidney 293T cells increased arsenic accumulation, and the cells were more susceptible to As(III) than 293T cells transfected with vector alone. These results suggest that AQP3 is involved in As(III) accumulation. Taken together, our results suggest that enhanced expression of MRP2 and lower expression of AQP3 are responsible for lower arsenic accumulation in arsenic-resistant R15 cells.

SummaryArsenic is an established human carcinogen. The role of aquaglyroporins (AQPs) in arsenic disposition was recently identified. In order to examine whether organic anion transporting polypeptide-C (OATP-C) also plays a role in arsenic transport, OATP-C cDNA was transfected into cells of a human embryonic kidney cell line (HEK-293). Transfection increased uptake of the model OATP-C substrate, estradiol-17b-D-glucuronide, by 10-fold. In addition, we measured uptake and cytotoxicity of arsenate, arsenite, monomethylarsonate(MMA V ), and dimethylarsinate (DMA V ). Transfection of OATP-C increased uptake and cytotoxicity of arsenate and arsenite, but not of MMA V or DMA V . Rifampin and taurocholic acid (a substrate of OATP-C) reversed the increased toxicity of arsenate and arsenite seen in OATP-C-transfected cells. The increase in uptake of inorganic arsenic was not as great as that of estradiol-17b-D-glucuronide. Our results suggest that OATP-C can transport inorganic arsenic in a (GSH)-dependent manner. However, this may not be the major pathway for arsenic transport.

Ticlopidine is sometimes coadministered with ergoloid mesylates or ginkgo biloba in clinical situations. Our objective was to examine the effect of ergoloid mesylates and ginkgo biloba on ticlopidine pharmacokinetics. Ticlopidine, ergoloid mesylates, and ginkgo biloba significantly inhibited the organic anion transporting polypeptide (OATP-B)-mediated uptake of [(3)H]-estrone-3-sulfate in a concentration-dependent manner. When ergoloid mesylates was coadministered with ticlopidine, the ticlopidine area under the plasma drug concentration-time profile (AUC) from 0 to 12 hours was decreased 30% and the peak plasma drug concentration (C(max)) was decreased 29%, compared with ticlopidine administration alone. There were no significant changes in the pharmacokinetic parameters of ticlopidine when it was coadministered with ginkgo biloba. In summary, ergoloid mesylates is a more potent inhibitor of OATP-B than is ginkgo biloba, and it can reduce the oral bioavailability of drugs transported by OATP-B. Ergoloid mesylates markedly decreased the AUC and C(max) of ticlopidine, probably by inhibiting the OATP-B-mediated uptake of ticlopidine during the intestinal absorption phase. The results support a new model of intestinal drug-drug interaction.

reported that erythromycin significantly increased the area under the curve per kilogram of bromocriptine by 286%, and a 4.6-fold greater maximum plasma concentration than that observed with bromocriptine alone was found. In their report the caffeine-bromocriptine interaction was also studied, and no significant interaction was found. The interaction between erythromycin and bromocriptine is clinically significant. However, the mechanism is not known.Erythromycin is a well-known substrate and inhibitor of cytochrome P450 (CYP) 3A4. Most drug-drug interactions involving erythromycin have been attributed to CYP3A4 inhibition. Peyronneau et al 2 indicated that bromocriptine is also a substrate of CYP3A4. We therefore used a human microsome assay to study whether erythromycin could inhibit the metabolism of bromocriptine. Pooled human liver microsomes (BD Biosciences, San Jose, Calif) containing 6200 pmol/mg · min testosterone 6␤-hydroxylation activity were used. The assay was conducted at 0.8-mg/mL protein with or without an NADPH-regenerating system (BD Biosciences) for 10 minutes at 37°C in triplicate. The products were analyzed by HPLC. 3 Even at a high concentration of erythromycin (100 mol/L), the inhibition of bromocriptine (0.5 mol/L) metabolism was only 8%.Organic anion transporting polypeptide C (OATP-C) is a hepatic uptake transporter with a wide range of substrates and inhibitors. 4 We used the OATP-C-overexpressed HEK-293 cells (HEK-OATP-C) 5 to investigate whether bromocriptine is an inhibitor of OATP-C and found that both bromocriptine (50% inhibitory concentration, 0.12 mol/L) and erythromycin (50% inhibitory concentration, 9.20 mol/L) significantly inhibited the OATP-C-mediated uptake of [ 3 H]estradiol-17␤-D-glucuronide (E 2 17␤G). Caffeine, which showed no interaction with bromocriptine, 1 did not affect the uptake of E 2 17␤G. In HEK-OATP-C cells erythromycin significantly inhibited the uptake of bromocriptine by OATP-C. At 0.5 and 1 mol/L of bromocriptine, the values of inhibition of its OATP-C uptake by 100 mol/L of erythromycin were 65% and 58%, respectively. A higher magnitude of inhibition was anticipated in the therapeutic range of bromocriptine (low nano-molar ranges). At the same bromocriptine concentration (0.5 mol/L), the magnitude of erythromycin inhibition on OATP-C is much higher than that for CYP3A4. Rather than inhibiting bromocriptine metabolism, erythromycin is more likely to have inhibited the hepatic uptake of bromocriptine. Nelson et al 1 suggested that erythromycin decreases first-pass metabolism and increases the systemic bioavailability of bromocriptine. Our data support their findings and suggest a mechanism.In conclusion, the results suggested that bromocriptine is a substrate of OATP-C. In the pharmacotherapeutics of Parkinsonism, the bromocriptine dose may need to be adjusted when patients are coadministered other OATP-C inhibitors or substrates. Furthermore, erythromycin may inhibit OATP-C uptake and CYP3A4 metabolism when it is coadministered with other drugs. When dr...

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