Resveratrol and its sulfated conjugates are Substrates of Organic Anion Transporting Polypeptides (OATPs): impact on resveratrol disposition

Resveratrol (3,4',5-trihydroxy-trans-stilbene) is a naturally occurring polyphenolic compound found in grapes, wine and medicinal plants with a variety of biological and pharmacological activities. These effects are observed despite its extremely low bioavailability and rapid clearance from the circulation due to extensive phase II metabolism. Whether resveratrol and its metabolites can accumulate to bioactive levels in organs and tissues by active transport mechanisms is not known yet. As OATPs mediate the uptake of endogenous compounds and clinically important drugs in various tissues, thereby affecting drug disposition, we investigated the transport of resveratrol and its three major sulfate conjugates in OATP1B1, OATP1B3 and OATP2B1-expressing Chinese Hamster Ovary (CHO) cells. Cells were cultured in 12 well plates and transport assays were performed using trans-resveratrol, trans-resveratrol-3-O-sulfate, trans-resveratrol-4-O-sulfate, and trans-resveratrol-disulfate at increasing concentrations. Samples were then analyzed by HPLC-MS/MS. For inhibition studies, cells were incubated in presence and absence of the OATP substrates bromosulphthalein and rifampicin. Results revealed 4- to 6-fold higher uptake rates for resveratrol in OATP1B1, OATP1B3 and OATP12B1- transfected CHO cells within 1 min, compared to wild type cells. Net OATP-mediated accumulation rates followed Michaelis-Menten kinetics with Km and Vmax values of 79.77 µM and 1149 pmol/mg protein/min for OATP1B1, 42.94 µM and 653.5 pmol/mg protein/min for OATP1B3, and 168.2 µM and 1216 pmol/mg protein/min for OATP2B1, respectively. Resveratrol-3-O-sulfate was only transported by OATP1B3 (Km: 277.0 µM; Vmax: of 186.8 pmol/mg protein/min), while resveratrol-disulfate was equally transported by OATP1B1 Km: 44.23 µM; Vmax: 100.7 pmol/mg protein/min) and OATP1B3 (Km: 41.4 µM; Vmax: 126.8 pmol/mg protein/min). Resveratrol-4-O-sulfate had a low affinity to all three OATPs as uptake was below detection limit in the tested cell lines. Furthermore, uptake of resveratrol, resveratrol-3-O-sulfate and resveratrol-disulfate was efficiently inhibited by bromsulfphthalein and rifampicin with IC50 values below 1 µM. In conclusion, our results elucidate, for the first time, the relevance of OATP1B1 and OATP1B3 as transporters for resveratrol and its sulfates