Long-term and mechanistic evaluation of drug-induced liver injury in Upcyte human hepatocytes

Abstract

Drug-induced liver injury (DILI) constitutes one of the most frequent reasons of restricted-use warnings as well as withdrawals of drugs in postmarketing and poses an important concern for the pharmaceutical industry. The current hepatic in vivo and in vitro models for DILI detection have shown clear limitations, mainly for studies of long-term hepatotoxicity. For this reason, we here evaluated the potential of using Upcytes human hepatocytes (UHH) for repeated-dose long-term exposure to drugs. The UHH were incubated with 15 toxic and non-toxic compounds for up to 21 days using a repeated-dose approach, and, in addition to conventional examination of effects on viability, the mechanisms implicated in cell toxicity were also assessed by means of high-content screening. The UHH maintained the expression and activity levels of drug-metabolizing enzymes for up to 21 days of culture and became more sensitive to the toxic compounds after extended exposures, showing inter-donor differences which would reflect variability among the population. The assay also allowed to detect the main mechanisms implicated in the toxicity of each drug as well as identifying special susceptibilities depending on the donor. UHH can be used for a long-term repeated detection of DILI at clinically relevant concentrations and also offers key mechanistic features of drug-induced hepatotoxicity. This system is therefore a promising tool in preclinical testing of human relevance that could help to reduce and/or replace animal testing for drug adverse effects.

Prediction of human drug-induced liver injury (DILI) in relation to oral doses and blood concentrations

“…In vitro methods with primary human hepatocytes (PHH) represent a well-established tool to identify concentrations of test compounds that induce toxicity or that cause gene expression alterations (Gebhardt et al 2003;Vinken and Hengstler 2018;Braeuning et al 2018). In addition, human hepatocytes engineered to allow in vitro expansion and cell lines are frequently used in this context (Tolosa et al 2019;Wink et al 2018;O'Brien et al 2006). However, the predictive performance of four so far published in vitro studies with PHH in 2D and 3D spheroid culture is limited, resulting in sensitivities of 51, 66, 59 and 69% and accuracies of 71, 71, 67 and 82%, respectively (Xu et al 2008;Khetani et al 2013;Proctor et al 2017;Vorrink et al 2018).…”

Identification of significant protein markers by mass spectrometry after co‐treatment of cells with different drugs: An in vitro survey platform

“…In this study, the protein basigin was identified, which is associated with liver injury. A group has previously reported that ketotifen has hepatotoxicity . After ethosuximide and metformin co‐treatment, 8 candidate protein markers were identified (Table ).…”

“…A group has previously reported that ketotifen has hepatotoxicity. 34 After ethosuximide and metformin co-treatment, 8 candidate protein markers were identified (Table 4). Tropomodulin-2 is associated with actin cytoskeleton, 35 and modification of the actin cytoskeletal organization by metformin has been reported.…”

The application of omics-based human liver platforms for investigating the mechanism of drug-induced hepatotoxicity in vitro

“…Thus, these prototypic short-term in vitro studies (24-48 h) predominantly reflect acute toxicity. The development of DILI, however, may take several weeks or months (Tolosa et al 2019). This delay may be related to drug metabolism, adaptation to drug intake (development of drug tolerance and physical dependence) or chronic liver injury caused by deposition of bilirubin (drug-induced cholestasis) or lipids (steatosis) (Schuemie et al 2016).…”