Usefulness of <i>in vitro</i> combination assays of mitochondrial dysfunction and apoptosis for the estimation of potential risk of idiosyncratic drug induced liver injury

Abstract

-Drug-induced liver injury (DILI) is one of the serious and frequent drug-related adverse events. This adverse event is a main reason for regulatory action pertaining to drugs, including restrictions in clinical indications and withdrawal from clinical trials or the marketplace. Idiosyncratic DILI especially has become a major clinical concern because of its unpredictable nature, frequent hospitalization, need for liver transplantation and high mortality. The estimation of the potential for compounds to induce idiosyncratic DILI is very difficult in non-clinical studies because the precise mechanism of idiosyncratic DILI is still unknown. Recently, many in vitro assays which indicate a possibility of the prediction of the idiosyncratic DILI have been reported. Among these, some in vitro assays focus on the effects of compounds on mitochondrial function and the apoptotic effects of compounds on human hepatocytes. In this study, we measured oxygen consumption rate (OCR) and caspase-3/7 activity as an endpoint of mitochondrial dysfunction and apoptosis, respectively, with human hepatocytes after treatment with compounds causing idiosyncratic DILI (troglitazone, leflunomide, ranitidine and diclofenac). Troglitazone and leflunomide decreased the OCR but did not affect caspase-3/7 activity. Ranitidine increased caspase-3/7 activity but did not affect the OCR. Diclofenac decreased the OCR and increased caspase-3/7 activity. Acetaminophen and ethanol, which are also hepatotoxicants but do not induce idiosyncratic DILI, did not affect the OCR or caspase-3/7 activity. These results indicate that a combination assay of mitochondrial dysfunction and apoptosis is useful for the estimation of potential risk of compounds to induce idiosyncratic DILI.

Evaluation of the Potential Risk of Drugs to Induce Hepatotoxicity in Human—Relationships between Hepatic Steatosis Observed in Non-Clinical Toxicity Study and Hepatotoxicity in Humans-

“…Finally, we estimated the potential risk of compound X to induce DILI in humans using in vitro combination assays of mitochondrial dysfunction and apoptosis as endpoints, which we have previously reported as a useful tool for estimation of risk of DILI [30]. Compound X induced both mitochondrial dysfunction and apoptosis in human primary hepatocytes in this assay (Figure 6 and Figure 7).…”

“…To sort these lipid ions, |w| > 0.05 and |p (corr)| > 0.8 in the loading s-plot of the OPLS-DA score, which represent the magnitude of contribution (weight) and reliability (correlation), respectively, were selected as cut-off values. Subsequently, the lipid ions identified as discriminant factors were subjected to identification of lipid molecules as described previously [30,31]. …”

Ether-phosphatidylcholine characterized by consolidated plasma and liver lipidomics is a predictive biomarker for valproic acid-induced hepatic steatosis

“…In the development of drugs, we sometimes encounter fatty change in the hepatocytes, which is not accompanied by any degenerative changes in the liver, in nonclinical toxicity studies, especially in rodents. There are many drugs that cause hepatic fatty change (steatosis) in rodents and some of them have the potential to induce DILI in humans through mitochondrial toxicity (McCarthy et al, 2004;Gudbrandsen et al, 2006;Ulrich et al, 2001;Silva et al, 2008;Le Dinh et al, 1988;Luiken et al, 2009;Choi et al, 2015;Sahini et al, 2014;Goda et al, 2016;Sahi et al, 2010). This is reasonable because mitochondria take part in the metabolism of lipids, including the oxidation of fatty acids, and fatty acids or other endogenous metabolites related to lipid metabolism will accumulate in the hepatocytes when the oxidation of fatty acids is inhibited by compounds as a consequence of their mitochondrial toxicity.…”

Mitochondrial dysfunction as a mechanism of drug-induced hepatotoxicity: current understanding and future perspectives

“…In addition, some studies suggest that the uncoupling effect of diclofenac stimulates mitochondrial respiration and enhances superoxide generation, with the increased ROS oxidizing membrane thiols, which play important roles in opening of the MPT [94]. Diclofenac was also shown to decrease the oxygen consumption rate in human hepatocytes [102] as well as in isolated rat liver mitochondria, accompanied by increased mitochondrial swelling [103]. In addition, diclofenac caused a decrease in ATP levels in human hepatocytes [104].…”