Translational Toxicology: Adapting Liver Microphysiological System for Cross-Species Evaluation of Hepatotoxicity Risk

Drug-induced liver injury (DILI) remains a leading cause of drug candidate attrition during development, underscoring the need for improved translational models that can predict human-specific toxicities. Traditional animal models, while informative for systemic toxicity, often fall short in forecasting human liver responses due to inherent interspecies differences. To address this translational gap, we leveraged the PhysioMimix® Organ-on-Chip (OOC) Core System to develop a robust DILI assay using primary hepatocytes from human, rat, and dog sources.
Methods
Using the PhysioMimix Core System, human, rat or dog hepatocytes were cultured as 3D liver microtissues under dynamic perfusion in the Liver-12 plate for up to 14 days, maintaining stable species-specific functionality (CYP activity, albumin, urea). A panel of six reference compounds (Troglitazone, Pioglitazone, Nefazodone, Buspirone, Tolcapone and Entacapone) recommended by the IQ MPS Consortium, including high-risk DILI agents and structural analogues with low human liability, was applied across a 7-point dosing regimen over four days. Functional in vitro and clinical biomarkers (albumin, urea, LDH, ALT) were monitored to assess hepatocellular health.
Results
The assay successfully captured known species-specific toxicities. For instance, nefazodone and troglitazone (DILI Rank 8) induced hepatotoxicity in human and rat models, while dog hepatocytes showed reduced sensitivity. Conversely, buspirone and entacapone exhibited minimal toxicity in animal models, though the human MPS detected subtle hepatotoxic signals at higher entacapone concentrations, highlighting its sensitivity to rare DILI events. Albumin consistently emerged as a sensitive cross-species marker, with dose-dependent decreases aligning with known toxic profiles.
Conclusions
These findings demonstrate the value of integrating human and animal liver MPS models to enhance cross-species interpretation and improve in vitro–in vivo extrapolation (IVIVE). The PhysioMimix system can be simply adapted to test multiple species in parallel to determine species-specific toxicity. As regulatory interest in MPS technologies grows, this approach offers a promising path to refine liver safety assessments, reduce reliance on and numbers of animal models used, and better predict human hepatotoxicity risks earlier in the drug development pipeline.

Presenter: Dr Anthony Berger

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Recapitulating Immune-Driven Hepatotoxicity Using a Liver Microphysiological Platform

With the April 2025 U.S. Food and Drug Administration (FDA) roadmap promoting New Approach Methodologies (NAMs) for regulatory testing of monoclonal antibodies (mAbs) and other drugs, there is a need for models with greater complexity and human relevance. For biologics such as mAbs, immune-mediated drug-induced liver injury (iDILI) remains a significant concern in drug development. Human-specific immune system interactions with new modalities, such as therapeutic antibodies, can lead to unpredictable hepatotoxicity, which often fail to be captured in traditional animal models and high-throughput cell line models, leading to poor translational outcomes and high attrition rates. The FDA-recognised PhysioMimix® DILI assay was modified to incorporate peripheral blood mononuclear cells (PBMCs) to address this with an immune-competent, human liver relevant MPS model. The model aims to enhance translational relevance and improve safety profiling in drug development pipelines.
Methods
Primary human hepatocytes were cultured under dynamic perfusion in Liver-12 plates by PhysioMimix Core to form 3D liver microtissues. The liver microtissues were analysed for functionality (CYP activity, albumin, urea) and clinically relevant injury markers (ALT, AST). Following four days of hepatocyte culture, HLA-matched PBMCs were added to the circulating media with the liver microtissues for a further four days and functionality assessed (cytokine profiling). Ipilimumab (anti-CTLA-4) or infliximab (anti-TNF-alpha) was dosed into the media of hepatocyte only and co-cultures for 48 hours and samples were taken at 1-, 4-, 8-, 24- and 48-hours post-dosing. The liver microtissues were evaluated for hepatotoxic responses while immune activation and inflammation was assessed by cytokine release.
Results
Immune cell interaction and recovery from the Liver-12 was first completed to determine the ability of the PBMC to transverse the microfluidic channels and scaffolds unhindered (60-70% recovery). Following initiation of co-culture, maintenance of liver microtissue functionality and stable immune phenotypes was demonstrated over four days of co-culture. Upon dosing with the mAbs in the co-culture model, immune-mediated hepatotoxicity was detected with significant elevation in LDH and ALT/AST, and reduction in the functional biomarkers, albumin and urea. Interestingly, significant cytokine increases were only seen in ipilimumab-treated conditions, aligning with clinical data which demonstrated a pro-inflammatory phenotype via activation of T-cells.
Conclusion
This proof-of-concept study using two clinically relevant monoclonal antibodies demonstrates the ability of the PhysioMimix DILI assay to predict immune-mediated hepatotoxicity. Future studies will look to further profile immune alterations and interactions with the liver microtissues. Together, this assay provides a human-specific solution to the assessment of hepatotoxicity of large molecules. The FDA’s endorsement of NAMs for investigational new drug (IND) applications marks a paradigm shift toward human-relevant, non-animal testing strategies, positioning the PhysioMimix® Core System as a pivotal tool for the future of immunotoxicity assessment and safer drug development.

Presenter: Dr. Justina Then

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