This report describes a potent and selective inhibitor of multidrug and toxin extrusion (MATE) protein, pyrimethamine (PYR), and examines its effect on the urinary and biliary excretion of typical Mate1 substrates in mice. In vitro inhibition studies demonstrated that PYR is a potent inhibitor of mouse (m)Mate1 (K_i = 145 nM) among renal organic cation transporters mOctn1 and mOctn2 (K_i > 30 μM), mOct1 (K_i = 3.6 μM), and mOct2 (K_i = 6.0 μM). PYR inhibited the uptake of metformin by kidney brush-border membrane vesicles (BBMVs) (K_i = 41 nM) and canalicular membrane vesicles in the presence of outward gradient of H⁺. PYR treatment significantly increased the kidney-to-plasma ratio of tetraethylammonium, and both the liver- and kidney-to-plasma ratios of metformin in mice, whereas it did not affect their plasma concentrations and urinary excretion rates. Furthermore, the plasma lactate concentration, a biomarker for inhibition of gluconeogenesis by metformin, was significantly higher in the PYR-treated group than in the control group. These results not only suggest the importance of mMate1 in the efflux of organic cations into the urine and bile in mice but also the importance of canalicular efflux mediated by MATE proteins for the therapeutic efficacy of metformin. PYR is a potent inhibitor of human (h)MATE1 and hMATE2-K (K_i = 77 and 46 nM, respectively) and H⁺ and organic cation exchanger in human kidney BBMVs (K_i = 31 nM) in the presence of outward gradient of H⁺. Taken together, PYR can be used as a potent probe inhibitor of human MATE transporters.

The purine salvage pathway plays a major role in the nucleotide production, relying on the supply of nucleobases and nucleosides from extracellular sources. Although specific transporters have been suggested to be involved in facilitating their transport across the plasma membrane in mammals, those which are specifically responsible for utilization of extracellular nucleobases remain unknown. Here we present the molecular and functional characterization of SLC43A3, an orphan transporter belonging to an amino acid transporter family, as a purine-selective nucleobase transporter. SLC43A3 was highly expressed in the liver, where it was localized to the sinusoidal membrane of hepatocytes and the lung. In addition, SLC43A3 expressed in MDCKII cells mediated the uptake of purine nucleobases such as adenine, guanine and hypoxanthine without requiring typical driving ions such as Na⁺ and H⁺, but it did not mediate the uptake of nucleosides. When SLC43A3 was expressed in APRT/HPRT1-deficient A9 cells, adenine uptake was found to be low. However, it was markedly enhanced by the introduction of SLC43A3 with APRT. In HeLa cells, knock-down of SLC43A3 markedly decreased adenine uptake. These data suggest that SLC43A3 is a facilitative and purine-selective nucleobase transporter that mediates the cellular uptake of extracellular purine nucleobases in cooperation with salvage enzymes.

Monocarboxylate transporter 7 (MCT7) is an orphan transporter expressed in the liver, brain, and in several types of cancer cells. It has also been reported to be a survival factor in melanoma and breast cancers. However, this survival mechanism is not yet fully understood due to MCT7’s unidentified substrate(s). Therefore, here we sought to identify MCT7 substrate(s) and characterize the transport mechanisms by analyzing amino acid transport in HEK293T cells and polarized Caco-2 cells. Analysis of amino acids revealed significant rapid reduction in taurine from cells transfected with enhanced green fluorescent protein-tagged MCT7. We found that taurine uptake and efflux by MCT7 was pH-independent and that the uptake was not saturated in the presence of taurine excess of 200 mM. Furthermore, we found that monocarboxylates and acidic amino acids inhibited MCT7-mediated taurine uptake. These results imply that MCT7 may be a low-affinity facilitative taurine transporter. We also found that MCT7 was localized at the basolateral membrane in polarized Caco-2 cells and that the induction of MCT7 expression in polarized Caco-2 cells enhanced taurine permeation. Finally, we demonstrated that interactions of MCT7 with ancillary proteins basigin/CD147 and embigin/GP70 enhanced MCT7-mediated taurine transport. In summary, these findings reveal that taurine is a novel substrate of MCT7 and that MCT7-mediated taurine transport might contribute to the efflux of taurine from cells.

Antibody–drug conjugates (ADCs) represent a new class of cancer therapeutics that enable targeted delivery of cytotoxic drugs to cancer cells. Although clinical efficacy has been demonstrated for ADC therapies, resistance to these conjugates may occur. Recently, SLC46A3, a lysosomal membrane protein, was revealed to regulate the efficacy of trastuzumab emtansine (T-DM1), a noncleavable ADC that has been widely used for treating breast cancer. However, the role of SLC46A3 in mediating T-DM1 cytotoxicity remains unclear. In this study, we discovered the function of SLC46A3 as a novel proton-coupled steroid conjugate and bile acid transporter. SLC46A3 preferentially recognized lipophilic steroid conjugates and bile acids as endogenous substrates. In addition, we found that SLC46A3 directly transports Lys-SMCC-DM1, a major catabolite of T-DM1, and potent SLC46A3 inhibitors attenuate the cytotoxic effects of T-DM1, suggesting a role in the escape of Lys-SMCC-DM1 from the lysosome into the cytoplasm. Our findings reveal the molecular mechanism by which T-DM1 kills cancer cells and may contribute to the rational development of ADCs that target SLC46A3.

In a recent clinical study, it was reported that the ingestion of apple juice (AJ) markedly reduced the plasma concentration of atenolol. However, our in vitro study showed that atenolol may not be a substrate of OATP2B1, so this AJ-atenolol interaction cannot be explained by inhibition of OATP2B1. On the other hand, we more recently showed that the solution osmolality influences gastrointestinal (GI) water volume, and this may indirectly affect intestinal drug absorption. In this study, we examined whether the osmolality-dependence of water dynamics can account for AJ-atenolol interactions by evaluating the GI water volume and the atenolol aborption in the presence of AJ in rats. Water absorption was the highest in purified water, followed by saline and isosmotic mannitol solution, and the lowest in AJ, confirming that water absorption is indeed osmolality-dependent. Interestingly, AJ showed apparent water secretion into the intestinal lumen. The intestinal concentration of FD-4, a non-permeable compound, after administration in AJ was lower than the initial concentration, while that in purified water was greater than the initial concentration. Further, the fraction absorbed of atenolol in intestine was significantly lower in AJ or hyperosmotic mannitol solution (adjusted to the osmolality of AJ) than after administration in purified water. Comparable results were observed in in vivo pharmacokinetic study in rats. Our results indicate that orally administered AJ has a capacity to modulate luminal water volume depending on the osmolality, and this effect may result in significant AJ-atenolol interactions.