BACKGROUND: During clinical trials bosentan, the first orally active endothelin receptor antagonist, caused asymptomatic transaminase elevations in some patients. In this study we investigated whether inhibition of the hepatocanalicular bile salt export pump (rodents, Bsep; humans, BSEP ABCB11) could account for bosentan-induced liver injury. METHODS: We reanalyzed the safety database of the bosentan trials for cholestatic liver injury, determined the cholestatic potency of bosentan in the rat, and studied the effects of bosentan and its metabolites on Bsep-mediated taurocholate transport in vitro. RESULTS: Bosentan caused dose-dependent and reversible liver injury in 2% to 18% of patients and caused a significant increase of serum bile salt levels (P <.01). Concomitant administration of glyburide (INN, glibenclamide) enhanced the cholestatic potency of bosentan. Similar effects were seen in rats, in which serum bile salt levels were increased by glyburide less than by bosentan, which increased the levels less than a combination of bosentan and glyburide. In vitro, Bsep-mediated taurocholate transport was inhibited by bosentan (inhibition constant, approximately 12 micromol/L) and metabolites (inhibition constant, approximately 8.5 micromol/L for metabolite Ro 47-8634). CONCLUSIONS: These results indicate that bosentan-induced liver injury is mediated, at least in part, by inhibition of Bsep/BSEP-causing intracellular accumulation of cytotoxic bile salts and bile salt induced liver cell damage. The data further emphasize the pathophysiologic importance of drug-Bsep interactions in acquired forms of cholestatic liver injury.