Screening of OATP1B1/3 and OCT1 inhibitors in cryopreserved hepatocytes in suspension

Drug–drug interactions involving hepatic drug transporters may have clinical consequences and jeopardize development of promising drug candidates. Organic anion transporting polypeptides (OATP/Oatp) and the organic cation transporters (OCT/Oct) are among the most important transporters involved in xenobiotic uptake in the liver. In the present study, 179 molecules have been tested as inhibitors of the uptake of estradiol-17βD-glucuronide (E217βG), substrate of OATP1B1/3 (rOatp), or 1-methyl-4-phenylpyridinium (MPP+), substrate of OCT1 (rOct1), into suspended cryopreserved hepatocytes from humans and rats. Uptake was assessed in 96-well plates by measuring intracellular accumulation of radioactive substrate in hepatocytes in presence or absence of inhibitor.In rat hepatocytes 140 compounds were identified as inhibitors (inhibition at 20 μM ≥ 30%) of E217βG uptake and 77 compounds inhibitors of MPP+ uptake. The most potent inhibitors of rOatp and rOct1 were dantrolene sodium (Ki = 2 ± 9 μM) and bepridil (Ki = 14 ± 2 μM), respectively. In human hepatocytes, the most potent inhibitors of E217βG and MPP+ uptake were capsazepine (Ki = 14 ± 5 μM) and cyproheptadine (Ki = 19 ± 3 μM), respectively.Structure–activity relationship (SAR) analysis of all tested compounds suggested that lipophilicity, polarity, pKa and the number of hydrogen bond donors and acceptors play a role in their interaction with the transporters investigated.The method used here is a simple tool to screen large number of compounds as inhibitors of the uptake of substrates into suspended hepatocytes.