Abstract
Multidrug and toxin extrusion (MATE) transporters are expressed on the luminal membrane of renal proximal tubule cells and extrude their substrates into the luminal side of the tubules. Inhibition of MATE1 can reduce renal secretory clearance of its substrate drugs, and lead to drug-drug interactions (DDI). In order to address if IC50 values of MATE1 inhibitors with regard to their extracellular concentrations are affected by the direction of MATE1-mediated transport, we established an efflux assay of 1-methyl-4-phenylpyridinium (MPP+) and metformin using the HEK293 model transiently expressing human MATE1. The efflux rate was defined by reduction of the cellular amount of MPP+ and metformin for 0.25 min shortly after the removal of extracellular MPP+ and metformin. Inhibition potencies of 12 inhibitors towards MATE1-mediated transport were determined in both uptake and efflux assays. When MPP+ was used as a substrate, 8 out of 12 inhibitors showed comparable IC50 values between assays (<4-fold). IC50 values from the efflux assays were higher for cimetidine (9.9-fold), trimethoprim (10-fold), famotidine (6.4-fold), and cephalexin (>3.8-fold). When metformin was used as a substrate, IC50 values of the tested inhibitors when evaluated using uptake and efflux assays were within 4-fold of each other, with an exception of cephalexin (>4.7-fold). IC50 values obtained from the uptake assay using metformin showed smaller IC50 values than those from efflux assay. Therefore, the uptake assay is recommended to determine IC50 values for the DDI predictions.
Significance Statement In this study, a new method to evaluate IC50 values of extracellular added inhibitors utilizing an efflux assay was established. IC50 values were not largely different between uptake and efflux directions but smaller in uptake. This study supports the rationale for commonly accepted uptake assay with metformin as an in vitro probe substrate for MATE1-mediated DDI risk assessment in drug development.
- drug transport
- drug-drug interactions
- efflux transporters (P-gp, BCRP, MRP, MATE, BSEP, etc)
- membrane transport
- Organic cation transport
- renal transport
- transporters
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