PT  - JOURNAL ARTICLE
AU  - Yi-an Bi
AU  - Sumathy Mathialagan
AU  - Laurie Tylaska
AU  - Myra Fu
AU  - Julie Keefer
AU  - Anna Vildhede
AU  - Chester Costales
AU  - A. David Rodrigues
AU  - Manthena Varma
TI  - Organic Anion Transporter 2 mediates hepatic uptake of tolbutamide, a Cytochrome P450 2C9 probe drug
AID  - 10.1124/jpet.117.245951
DP  - 2018 Jan 01
TA  - Journal of Pharmacology and Experimental Therapeutics
PG  - jpet.117.245951
4099  - http://jpet.aspetjournals.org/content/early/2018/01/11/jpet.117.245951.short
4100  - http://jpet.aspetjournals.org/content/early/2018/01/11/jpet.117.245951.full
AB  - Tolbutamide is primarily metabolized by cytochrome P450 (CYP)2C9, and thus, frequently applied as a clinical probe substrate for CYP2C9 activity. However, there is a marked discrepancy in the in vitro-in vivo extrapolation of its metabolic clearance implying potential for additional clearance mechanisms. The goal of this study was to evaluate the role of hepatic uptake transport in the pharmacokinetics of tolbutamide and identify the molecular mechanism thereof. Transport studies using singly-transfected cells expressing six major hepatic uptake transporters showed that tolbutamide is a substrate to organic anion transporter (OAT)2 alone -- with transporter affinity (Km) of 19.5±4.3 μM. Additionally, OAT2-specific transport was inhibited by ketoprofen (OAT2 inhibitor) and 1mM rifamycin SV (pan inhibitor), but not by cyclosporine and rifampicin (OATPs/NTCP inhibitors). Uptake studies in primary human hepatocytes confirmed the predominant role of OAT2 in the active uptake with significant inhibition by rifamycin SV and ketoprofen, but not by the other inhibitors. Concentration-dependent uptake was noted in human hepatocytes with active transport characterized by Km and Vmax of 39.3±6.6 μM and 426±30 pmol/min/mg-protein, respectively. Bottom-up physiologically based pharmacokinetic modeling was employed to verify the proposed role of OAT2-mediated hepatic uptake. In contrast to the rapid-equilibrium (CYP2C9-only) model, permeability-limited (OAT2-CYP2C9 interplay) model better described the plasma concentration-time profiles of tolbutamide. Additionally, the latter well described tolbutamide pharmacokinetics in carriers of CYP2C9 genetic variants and quantitatively rationalized its known drug-drug interactions. Our results provide first-line evidence for the role of OAT2-mediated hepatic uptake in the pharmacokinetics of tolbutamide; and imply the need for additional clinical studies in this direction.