Transcriptomic Analysis in Liver Spheroids Identifies a Dog-Specific Mechanism of Hepatotoxicity for Amcenestrant

Therapeutic drugs can sometimes cause adverse effects in a nonclinical species that do not translate to other species, including human. Species-specific (rat, dog, and human) in vitro liver spheroids were employed to understand the human relevance of cholestatic liver injury observed with a selective estrogen receptor degrader (amcenestrant) in dog, but not in rat, during preclinical development. Amcenestrant showed comparable cytotoxicity in liver spheroids from all three species; however, its M5 metabolite (RA15400562) showed dog preferential cytotoxicity after seven days of treatment. Whole genome transcript profiles generated from liver spheroids revealed downregulation of genes related to bile acid synthesis and transport indicative of strong farnesoid X receptor (FXR) antagonism following treatment with both amcenestrant and its M5 metabolite in the dog but not in rat or human. In human spheroids, upregulation of genes for detoxification enzymes indicative of pregnane X receptor (PXR) agonism was observed following amcenestrant treatment, whereas in the dog these genes were downregulated. The M5 metabolite showed gene dysregulation indicating PXR agonism in both rat and human, and antagonism in dog. Analysis of liver samples from a three-month dog toxicity study conducted with amcenestrant showed downregulation of several genes associated with PXR and FXR, corroborating the in vitro results. These results support the hypothesis that dogs are uniquely susceptible to cholestatic hepatotoxicity following administration of amcenestrant due to species-specific antagonism of FXR and highlight the value of in vitro liver spheroids to investigating mechanisms of toxicity and possible species differences.