RT Journal Article SR Electronic T1 A Two-Tiered In Vitro Approach to De-Risk Drug Candidates for Potential Bile Salt Export Pump Inhibition Liabilities in Drug Discovery JF Drug Metabolism and Disposition JO Drug Metab Dispos FD American Society for Pharmacology and Experimental Therapeutics SP 1147 OP 1160 DO 10.1124/dmd.120.000086 VO 48 IS 11 A1 Michael J. Hafey A1 Robert Houle A1 Keith Q. Tanis A1 Ian Knemeyer A1 Jackie Shang A1 Qing Chen A1 Andreas Baudy A1 James Monroe A1 Frank D. Sistare A1 Raymond Evers YR 2020 UL http://dmd.aspetjournals.org/content/48/11/1147.abstract AB Hepatocellular accumulation of bile salts by inhibition of bile salt export pump (BSEP/ABCB11) may result in cholestasis and is one proposed mechanism of drug-induced liver injury (DILI). To understand the relationship between BSEP inhibition and DILI, we evaluated 64 DILI-positive and 57 DILI-negative compounds in BSEP, multidrug resistance protein (MRP) 2, MRP3, and MRP4 vesicular inhibition assays. An empirical cutoff (5 μM) for BSEP inhibition was established based on a relationship between BSEP IC50 values and the calculated maximal unbound concentration at the inlet of the human liver (fu*Iin,max, assay specificity = 98%). Including inhibition of MRP2–4 did not increase DILI predictivity. To further understand the potential to inhibit bile salt transport, a selected subset of 30 compounds were tested for inhibition of taurocholate (TCA) transport in a long-term human hepatocyte micropatterned co-culture (MPCC) system. The resulting IC50 for TCA in vitro biliary clearance and biliary excretion index (BEI) in MPCCs were compared with the compound’s fu*Iin,max to assess potential risk for bile salt transport perturbation. The data show high specificity (89%). Nine out of 15 compounds showed an IC50 value in the BSEP vesicular assay of <5μM, but the BEI IC50 was more than 10-fold the fu*Iin,max, suggesting that inhibition of BSEP in vivo is unlikely. The data indicate that although BSEP inhibition measured in membrane vesicles correlates with DILI risk, that measurement of this assay activity is insufficient. A two-tiered strategy incorporating MPCCs is presented to reduce BSEP inhibition potential and improve DILI risk.SIGNIFICANCE STATEMENT This work describes a two-tiered in vitro approach to de-risk compounds for potential bile salt export pump inhibition liabilities in drug discovery utilizing membrane vesicles and a long-term human hepatocyte micropatterned co-culture system. Cutoffs to maximize specificity were established based on in vitro data from a set of 121 DILI-positive and –negative compounds and associated calculated maximal unbound concentration at the inlet of the human liver based on the highest clinical dose.