Assessing drug-induced liver injury liabilities in human and animal hepatocytes under static and perfused spheroid culture conditions

Abstract

Abstract Drug-induced liver injury (DILI) remains a major challenge in drug development, highlighting the need for reliable in vitro tools to assess hepatotoxicity and improve translation from animal to human studies. Models using hepatocytes from preclinical species are also needed to evaluate species-specific toxicity. In this study, we evaluated the basal function and DILI sensitivity of primary hepatocytes from human, monkey, rat, and dog cultured for up to 14 days in static monolayers, static spheroids, and spheroids cultured in a microfluidics-based microphysiological system (MPS). Hepatocyte function and injury responses were assessed using albumin, urea, and liver enzymes. Cells were exposed to species-specific DILI compounds chlorpromazine (CPZ), bosentan (BOS), and fialuridine (FIAU). Across platforms, human and monkey hepatocytes exhibited greater functional stability and sensitivity to DILI compounds than rat and dog hepatocytes. CPZ and BOS induced cytotoxicity primarily in human and monkey hepatocytes, while FIAU produced species-dependent effects consistent with known in vivo outcomes. The microfluidics-based MPS exhibited modestly improved hepatocyte spheroid function relative to static models, although limited MPS throughput constrained our ability for testing drugs beyond FIAU. Overall, these results demonstrate that integrating multi-species hepatocyte spheroids across static and microfluidic platforms enables comparative DILI assessment and supports improved preclinical-to-clinical translation.