Hepatic Transporter Alterations by Nuclear Receptor Agonist T0901317 in Sandwich-cultured Human Hepatocytes: Proteomic Analysis and PBPK Modeling to Evaluate Drug-Drug Interaction Risk

In vitro approaches for predicting drug-drug interactions (DDIs) caused by alterations in transporter protein regulation are not well established. However, reports of transporter regulation via nuclear receptor (NR) modulation by drugs are increasing. This study examined alterations in transporter protein levels in sandwich-cultured human hepatocytes (SCHH; n = 3 donors) measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomic analysis after treatment with T0901317, the first described synthetic liver X receptor (LXR) agonist. T0901317 treatment (10 μM, 48 h) decreased the levels of organic cation transporter (OCT) 1 (0.22-, 0.43- and 0.71-fold of control) and organic anion transporter (OAT) 2 (0.38-, 0.38- and 0.53-fold of control), and increased multidrug resistance protein (MDR) 1 (1.37-, 1.48 and 1.59-fold of control). The induction of NR downstream gene expression support the hypothesis that T0901317 off-target effects on farnesoid X receptor (FXR) and pregnane X receptor (PXR) activation are responsible for the unexpected changes in OCT1, OAT2 and MDR1. Uptake of the OCT1 substrate metformin in SCHH was decreased by T0901317 treatment. Effects of decreased OCT1 levels on metformin were simulated using a physiologically-based pharmacokinetic (PBPK) model. Simulations showed a clear decrease in metformin hepatic exposure resulting in a decreased pharmacodynamic effect. This DDI would not be predicted by the modest changes in simulated metformin plasma concentrations. Altogether, the current study demonstrated that an approach combining SCHH, proteomic analysis, and PBPK modeling is useful for revealing tissue concentration-based DDIs due to unexpected regulation of hepatic transporters by NR modulators. SIGNIFICANCE STATEMENT This study utilized an approach combining sandwich-cultured human hepatocytes, proteomic analysis, and physiologically-based pharmacokinetic modeling to evaluate alterations in pharmacokinetics (PK) and pharmacodynamics (PD) due to transporter regulation by nuclear receptor modulators. The importance of this approach from a mechanistic and clinically-relevant perspective is that it can reveal drug-drug interactions (DDIs) due to unexpected regulation of hepatic transporters, and enable prediction of altered PK and PD changes, especially for tissue concentration-based DDIs.