Bosentan alters endo- and exogenous bile salt disposition in sandwich-cultured human hepatocytes

Bosentan, a well-known cholestatic agent, was not identified as cholestatic at concentrations up to 200 µM based on the drug-induced cholestasis (DIC) index value, determined in a sandwich-cultured human hepatocyte (SCHH)-based DIC assay. To obtain further quantitative insights into the effects of bosentan on cellular bile salt handling by human hepatocytes, the present study determined the effect of 2.5-25 µM bosentan on endogenous bile salt levels and on the disposition of 10 µM chenodeoxycholic acid (CDCA) added to the medium in SCHH. Bosentan reduced intracellular as well as extracellular concentrations of both endogenous glycochenodeoxycholic acid (GCDCA) and glycocholic acid in a concentration-dependent manner. When exposed to 10 µM CDCA, bosentan caused a shift from canalicular efflux to sinusoidal efflux of GCDCA. CDCA levels were not affected. Our mechanistic model confirmed the inhibitory effect of bosentan on canalicular GCDCA clearance. Moreover, our results in SCHH also indicated reduced GCDCA formation. We confirmed the direct inhibitory effect of bosentan on CDCA conjugation with glycine in liver S9 fraction. Significance Statement Bosentan was evaluated at therapeutically relevant concentrations (2.5-25 µM) in sandwich-cultured human hepatocytes. It altered bile salt disposition and inhibited canalicular secretion of glycochenodeoxycholate (GCDCA). Within 24 h, bosentan caused a shift from canalicular to sinusoidal efflux of GCDCA. Our results also indicated reduced GCDCA formation. We confirmed a direct effect of bosentan on chenodeoxycholic acid conjugation with glycine in liver S9 fraction.