Liver safety evaluation of endothelin receptor antagonists using HepatoPac® : A single model impact assessment on hepatocellular health, function and bile acid disposition.

Marketed (bosentan, ambrisentan) and discontinued (sitaxsentan, CI-1034) endothelin receptor antagonists were examined in the human micropatterned hepatocyte co-culture (MPCC) model HepatoPac® . Differences across hepatocellular health (cellular adenosine triphosphate/glutathione content), function (urea production/albumin secretion) and taurocholic acid transport (biliary clearance/excretion index) were compared using amiodarone and ciclosporin A as positive controls. Ambrisentan had the weakest potency in all six endpoints, while sitaxsentan, bosentan and CI-1034 had more potent effects on hepatobiliary transport than health/function endpoints. Normalization to clinical Cmax gave the following relative rank order of safety based on margins for each endpoint: ambrisentan ≥ CI-1034 ~ bosentan > sitaxsentan. These data suggested impaired hepatobiliary disposition might contribute to a more prominent role in liver injury associated within sensitive human populations exposed to these compounds than direct hepatocellular toxicity. Rat, dog and monkey MPCCs also showed greater sensitivity potential to disrupted hepatobiliary disposition compared with hepatocellular health/functional endpoints. Drug metabolism competency was exhibited across all species. In vivo, rats and dogs appear more resistant to transaminase elevations and/or histological evidence of liver injury caused by these mechanisms even at exceedingly high systemic exposures relative to sensitive humans. Rats and dogs are resistant to hepatobiliary toxicants due to physiological differences in bile composition/handling. Although traditional animal testing provides adequate safety coverage for advancement of novel pharmaceuticals into clinical trials, supplemental assays employing human MPCCs may strengthen weight-of-evidence predictions for sensitive human populations. Proving the predictive value of this single impact assessment model in advance of clinical trial information for human liver injury risk is needed across more pharmaceuticals.

Key Challenges and Opportunities Associated with the Use of In Vitro Models to Detect Human DILI: Integrated Risk Assessment and Mitigation Plans.

Drug-induced liver injury (DILI) is a major cause of late-stage clinical drug attrition, market withdrawal, black-box warnings, and acute liver failure. Consequently, it has been an area of focus for toxicologists and clinicians for several decades. In spite of considerable efforts, limited improvements in DILI prediction have been made and efforts to improve existing preclinical models or develop new test systems remain a high priority. While prediction of intrinsic DILI has improved, identifying compounds with a risk for idiosyncratic DILI (iDILI) remains extremely challenging because of the lack of a clear mechanistic understanding and the multifactorial pathogenesis of idiosyncratic drug reactions. Well-defined clinical diagnostic criteria and risk factors are also missing. This paper summarizes key data interpretation challenges, practical considerations, model limitations, and the need for an integrated risk assessment. As demonstrated through selected initiatives to address other types of toxicities, opportunities exist however for improvement, especially through better concerted efforts at harmonization of current, emerging and novel in vitro systems or through the establishment of strategies for implementation of preclinical DILI models across the pharmaceutical industry. Perspectives on the incorporation of newer technologies and the value of precompetitive consortia to identify useful practices are also discussed.

Establishment of a hepatocyte-kupffer cell coculture model for assessment of proinflammatory cytokine effects on metabolizing enzymes and drug transporters.

Elevated levels of proinflammatory cytokines associated with infection and inflammation can modulate cytochrome P450 enzymes, leading to potential disease-drug interactions and altered small-molecule drug disposition. We established a human-derived hepatocyte-Kupffer cell (Hep:KC) coculture model to assess the indirect cytokine impact on hepatocytes through stimulation of KC-mediated cytokine release and compared this model with hepatocytes alone. Characterization of Hep:KC cocultures showed an inflammation response after treatment with lipopolysaccharide and interleukin (IL)-6 (indicated by secretion of various cytokines). Additionally, IL-6 exposure upregulated acute-phase proteins (C-reactive protein, alpha-1-acid glycoprotein, and serum amyloid A2) and downregulated CYP3A4. Compared with hepatocytes alone, Hep:KC cocultures showed enhanced IL-1ß-mediated effects but less impact from both IL-2 and IL-23. Hep:KC cocultures treated with IL-1ß exhibited a higher release of proinflammatory cytokines, an increased upregulation of acute-phase proteins, and a larger extent of metabolic enzyme and transporter suppression. IC50 values for IL-1ß-mediated CYP3A4 suppression were lower in Hep:KC cocultures (98.0-144 pg/ml) compared with hepatocytes alone (IC50 > 5000 pg/ml). Cytochrome suppression was preventable by blocking IL-1ß interaction with IL-1R1 using an antagonist cytokine or an anti-IL-1ß antibody. Unlike IL-1ß, IL-6-mediated effects were comparable between hepatocyte monocultures and Hep:KC cocultures. IL-2 and IL-23 caused a negligible inflammation response and a minimal inhibition of CYP3A4. In both hepatocyte monocultures and Hep:KC cocultures, IL-2RB and IL-23R were undetectable, whereas IL-6R and IL-1R1 levels were higher in Hep:KC cocultures. In summary, compared with hepatocyte monocultures, the Hep:KC coculture system is a more robust in vitro model for studying the impact of proinflammatory cytokines on metabolic enzymes.