OATP-mediated hepatic uptake of glucuronide metabolites of androgens.

We previously established that androgen glucuronides are primarily effluxed by MRP2 and MRP3 in liver and intestine. However, no data exist on the mechanism of hepatic uptake of these metabolites. To fill this knowledge gap, the first goal of this study was to explore the role of hepatic uptake transporters and characterize transport kinetics of glucuronides of testosterone (TG), dihydrotestosterone (DHTG), androsterone (AG), and etiocholanolone (EtioG) using cell-lines overexpressing organic anion transporting polypeptide (OATP1B1, OATP1B3, and OATP2B1). Using quantitative proteomics-guided approach, we then estimated the relative contribution (ft) of individual OATPs in hepatic uptake of the androgen glucuronides. The transport screening assays revealed that the glucuronides were primarily influxed by OATP1B1 and OATP1B3. The Km values for OATP1B1-mediated uptake were low for EtioG (6.2 µM), followed by AG, TG, and DHTG (46.2, 56.7, and 71.3 µM, respectively). Whereas, the Km value for OATP1B3-mediated uptake for EtioG, AG, DHTG, and TG were 19.8, 29.3, 69.6, and 110.4 µM, respectively. Both OATP1B1 and OATP1B3 exhibited highest transport rate toward AG as compared to other glucuronides. When adjusted for transporter abundance in human livers, EtioG and DHTG were transported equally by OATP1B1 and OATP1B3, whereas TG and AG were preferentially (>68%) transported by OATP1B3. Collectively, this is the first report that elucidate mechanisms of hepatic uptake of androgen glucuronides. Perturbation in these processes by genetic polymorphisms, disease conditions or drug-interactions can lead to changes in enterohepatic recycling of androgen. The apparent higher selectivity of OATP1B3 towards TG and AG can be leveraged for establishing these metabolites as clinical biomarkers of OATP1B3 activity. SIGNIFICANCE STATEMENT: This is the first study to elucidate the mechanism of hepatic uptake of androgen glucuronides and estimate the relative contribution (ft) of individual OATPs using quantitative proteomics. Our results show that both OATP1B1 and OATP1B3 are responsible for the hepatic uptake of major circulating testosterone glucuronides. The apparent higher selectivity of OATP1B3 towards testosterone glucuronide and androsterone glucuronide can be leveraged for establishing these metabolites as clinical biomarkers of OATP1B3 activity. The American Society for Pharmacology and Experimental Therapeutics.