gms | German Medical Science

MATE1 (multidrug and toxin extrusion protein 1) and MATE2K, encoded respectively by the SCL47A1 and SCL47A2 genes, are transporters in mammals mediating the efflux of organic cations and zwitterions. They are located preferentially on the apical membrane of epithelia of the renal proximal tubules. MATE1 is also expressed on the apical membrane of hepatocytes [1]. Under physiological conditions, the high proton concentration in the tubular lumen and bile canaliculi drives the MATE antiporters.

Many drugs have been reported as potential substrates of MATE1 and MATE2K, suggesting that their functions may determine the pharmacokinetics and relate to drug-drug interactions. The objective of this study was to assess the function of these human transporters in a uniform system.

We transfected HEK293 cells respectively with MATE1 or MATE2K via a suitable vector for single transfection, pEBTetD [2]，which provides a stable episomal expression of the transporters; the expression can be regulated by doxycycline. The switched-off cells were used as control. To quantify the uptake, instead of the efflux in vivo, the transfected cells were incubated at 37°C for 1 minute with 1 µmol/L of the following potential substrates : MPP⁺, guanidine, estrone-3-sulfate, metformin, captopril, cephalexin, and quinine. The amount of accumulated compound in the cells was subsequently measured with LC-MS/MS using Atlantis® HILIC sillica 5µm, 3,0 x 50mm column and Atlantis®HILIC sillica 5µm, 3,9 x 20mm guard column. The experiments were repeated on MATE2K-transfected cells with 10 µmol/L MPP⁺, metformin, cephalexin and guanidine. The compound concentrations inside the cells were measured by LC-MS/MS with ZIC-HILIC® 5µm, 100mm x 2,1mm column. Only a clear difference of concentration between switched on and off cells indicates an uptake of the substance through the transporters and was precisely quantified. According to it and the average protein content of the cells in each Petri dish, the rate of uptake was calculated, to indicate the transport efficiency.

The measurements showed, MPP⁺, metformin and cephalexin were taken up by the MATE1- transfected cells at a mean rate of 422 ± 15, 34 ± 5 and 2,1 ± 0,2 pmol/mg protein/minute. At 1 µM, MPP⁺ was also transported by MATE2K, but metformin was not. MATE2K transported MPP⁺ at a rate of 216 ± 28 and metformin at 129 ± 6 pmol/mg protein/minute in the cells when increasing the concentration in incubation to 10 µmol/L. High levels of estrone-3-sulfate, quinine and guanidine were detected in the lysates of both switched-on and -off cells without significant differences. Captopril could not be measured inside the cells.

In conclusion, MPP⁺ and metformin are substrates of both transporters. Cephalexin is a poor substrate of MATE1, but not transported by MATE2K. Estrone-3-sulfate, quinine, captopril and guanidine are no substrates.