Biology subjects

Fontana, R. J.

Publications and source records attributed to Fontana, R. J..

3 recordsLinked to original sources

bioRxiv2026

High-Throughput Morphological Profiling of Patient-Derived Human Liver Organoids for Hepatotoxicity Hazard Identification and Compound Prioritization

Drug-induced liver injury (DILI) remains a major challenge in drug development because existing in vitro models poorly predict clinical hepatotoxicity. We coupled patient-derived human liver organoids (HLOs) with high-content imaging and automated machine learning (ML) to detect hepatotoxicity-associated phenotypes by morphological profiling. HLOs from six DILI patients and one healthy donor (n = 7) retained donor-specific baseline phenotypes in hepatocyte maturity, metabolism, and morphology. ML models trained per line on MAD-robustized, well-level-aggregated median features achieved compound-level AUROCs of 0.88 (95% CI 0.86-0.90) to 0.91 (95% CI 0.89-0.93) and balanced accuracies of 0.81-0.84 in distinguishing 54 hepatotoxicants from 20 non-hepatotoxic controls. Averaging predictions across all seven lines at an optimized threshold gave 94.4% sensitivity (95% CI 84.6-98.8%) and 85.0% specificity (95% CI 62.1-96.8%), substantially outperforming HLO cytotoxicity assays and cancer cell line profiling. Lines were moderately-to-highly concordant (Pearson r = 0.66-0.85), and benchmark concentration modeling revealed reproducible inter-line differences in phenotypic sensitivity (median 4.0-fold, several compounds exceeding 20-fold). Exposure anchoring of benchmark concentrations to human-equivalent plasma concentrations, using in vitro mass-balance dosimetry and curated plasma free fractions, placed organoid points of departure within 100-fold of clinical Cmax for 60 of 66 compounds (23-fold residual offset, 95% CI 12-45), and placed two of the three misclassified negative controls an order of magnitude above clinical Cmax. Sulindac injury signatures were significantly enhanced by TNF co-treatment, demonstrating capacity to detect pro-inflammatory-mediated DILI. This platform preserves inter-line variability while enabling reproducible, scalable multi-line consensus hazard classification; because compounds were tested over a common nominal concentration range, the readout defines hepatocellular hazard, which must be combined with exposure as an independent input to estimate clinical DILI risk.

pharmacology and toxicology↗

bioRxiv2023

A High-Throughput Microphysiological Liver Chip System to Model Direct and Idiosyncratic Drug-Induced Liver Injury Using Human Liver Organoids

1.ObjectiveDrug-induced liver injury (DILI) is a major failure mode in pharmaceutical development. This study aims to address the limitations of existing preclinical models by introducing a high-throughput, microfluidic liver-on-a-chip system, termed as "Curio Barrier Liver Chips," seeded with human liver organoids to enable metabolic and phenotypic morphologic characterization. MethodsCurio Barrier liver chips, fabricated in an 8x2 well configuration, were utilized to establish 3D liver organoid cultures. Human-induced pluripotent stem cells (iPSCs) were differentiated into hepatic-like organoids (HLOs), and their viability, liver-specific functions, and pharmacological responses were assessed over 28 days. ResultsThe Curiochips successfully maintained liver physiology and function, showing strong albumin secretion and cytochrome P450 (CYP) activities through 28 days. Unlike traditional models requiring millimolar drug concentrations to detect hepatotoxicity, this platform showed increased sensitivity for APAP and FIAU at micromolar concentrations. In situ differentiation of foregut spheroids to liver organoids was also achieved, further simplifying the establishment of liver chips. Furthermore, the chips demonstrated viability, function and DILI responsiveness for 28 days making this an improved model for studying idiosyncratic DILI with prolonged drug exposure and high-throughput capabilities compared to other available systems or primary human hepatocytes. ConclusionsThe Curiochips offer an advanced, miniaturized in vitro model for early-stage drug development and a sensitive, responsive and cost-effective means to detect direct hepatotoxicity. The high-throughput capability, coupled with robust functionality and pharmacological responses make it a promising tool for improving the prediction and understanding of DILI mechanisms in general and those that required prolonged drug exposure. The model also opens new avenues for research in other chronic liver diseases.

pharmacology and toxicology↗

bioRxiv2021

A Multi-Omics Human Liver Organoid Screening Platform for DILI Risk Prediction

Background and AimsDrug-induced liver injury (DILI), both intrinsic and idiosyncratic, causes frequent morbidity, mortality, clinical trial failures and post-approval withdrawal. This suggests an unmet need for improved in vitro models for DILI risk prediction that can account for diverse host genetics and other clinical factors. In this study, we evaluated the utility of human liver organoids (HLOs) for high-throughput DILI risk prediction and in an organ-on-chip system. MethodsHLOs were derived from 3 separate iPSC lines and benchmarked on two platforms for their ability to model in vitro liver function and identify hepatotoxic compounds using biochemical assays for albumin, ALT, and AST, microscopy-based morphological profiling, and single-cell transcriptomics: 1) HLOs dispersed in 384-well formatted plates and exposed to a library of compounds. 2) HLOs adapted to a liver-on-chip system. ResultsO_LIDispersed HLOs derived from the 3 iPSC lines had similar DILI predictive capacity to intact HLOs in a high-throughput screening format allowing for measurable IC50 values of compound cytotoxicity. Distinct morphological differences were observed in cells treated with drugs exerting differing mechanisms of toxicity. C_LIO_LIOn-chip HLOs significantly increased albumin production, CYP450 expression, and ALT/AST release when treated with known DILI drugs compared to dispersed HLOs and primary human hepatocytes. On-chip HLOs were able to predict the synergistic hepatotoxicity of tenofovir-inarigivir and showed steatosis and mitochondrial perturbation via phenotypic and transcriptomic analysis with exposure to FIAU and acetaminophen, respectively. C_LI ConclusionsThe high throughput and liver-on-chip system exhibit enhanced in vivo-like function and demonstrate the potential utility of these platforms for hepatotoxicity risk assessment. Tenofovir-inarigivr associated hepatotoxicity was observed and correlates with the clinical manifestation of DILI observed in patients. LAY SUMMARYIdiosyncratic (spontaneous, patient-specific) drug-induced liver injury (DILI) is difficult to study due to the lack of liver models that function as human liver tissue and are adaptable for large-scale drug screening. Human liver organoids grown from patient stem cells respond to known DILI-causing drugs in both a high-throughput and on a physiological "chip" culture system. These platforms show promise in their use as predictive model for novel drugs before entering clinical trials.

pharmacology and toxicology↗