MITOLOGICS SAS, Hôpital Robert Debré, F-75019 Paris, France.
Toxicol Sci. 2012 Oct;129(2):332-45. doi: 10.1093/toxsci/kfs197.
Drug-induced liver injury (DILI) in humans is difficult to predict using classical in vitro cytotoxicity screening and regulatory animal studies. This explains why numerous compounds are stopped during clinical trials or withdrawn from the market due to hepatotoxicity. Thus, it is important to improve early prediction of DILI in human. In this study, we hypothesized that this goal could be achieved by investigating drug-induced mitochondrial dysfunction as this toxic effect is a major mechanism of DILI. To this end, we developed a high-throughput screening platform using isolated mouse liver mitochondria. Our broad spectrum multiparametric assay was designed to detect the global mitochondrial membrane permeabilization (swelling), inner membrane permeabilization (transmembrane potential), outer membrane permeabilization (cytochrome c release), and alteration of mitochondrial respiration driven by succinate or malate/glutamate. A pool of 124 chemicals (mainly drugs) was selected, including 87 with documented DILI and 37 without reported clinical hepatotoxicity. Our screening assay revealed an excellent sensitivity for clinical outcome of DILI (94 or 92% depending on cutoff) and a high positive predictive value (89 or 82%). A highly significant relationship between drug-induced mitochondrial toxicity and DILI occurrence in patients was calculated (p < 0.001). Moreover, this multiparametric assay allowed identifying several compounds for which mitochondrial toxicity had never been described before and even helped to clarify mechanisms with some drugs already known to be mitochondriotoxic. Investigation of drug-induced loss of mitochondrial integrity and function with this multiparametric assay should be considered for integration into basic screening processes at early stage to select drug candidates with lower risk of DILI in human. This assay is also a valuable tool for assessing the mitochondrial toxicity profile and investigating the mechanism of action of new compounds and marketed compounds.
药物性肝损伤（DILI）在人类中很难通过经典的体外细胞毒性筛选和监管动物研究来预测。这解释了为什么在临床试验期间或由于肝毒性而有许多化合物被停止或从市场上撤出。因此，提高对人类 DILI 的早期预测是很重要的。在这项研究中，我们假设通过研究药物诱导的线粒体功能障碍可以实现这一目标，因为这种毒性作用是 DILI 的主要机制之一。为此，我们开发了一种使用分离的小鼠肝线粒体的高通量筛选平台。我们的广谱多参数测定法旨在检测全局线粒体膜通透性（肿胀）、内膜通透性（跨膜电位）、外膜通透性（细胞色素 c 释放）以及琥珀酸或苹果酸/谷氨酸驱动的线粒体呼吸的改变。选择了 124 种化学物质（主要是药物）的混合物，包括 87 种有记录的 DILI 和 37 种没有报道的临床肝毒性。我们的筛选测定法对 DILI 的临床结果具有出色的敏感性（取决于截止值，分别为 94%或 92%）和高阳性预测值（89%或 82%）。计算出药物诱导的线粒体毒性与患者 DILI 发生之间存在高度显著的关系（p < 0.001）。此外，该多参数测定法还可鉴定出几种以前从未描述过线粒体毒性的化合物，甚至有助于澄清某些已被认为是线粒体毒性的药物的作用机制。用这种多参数测定法研究药物诱导的线粒体完整性和功能丧失，应考虑纳入早期基本筛选过程，以选择在人类中 DILI 风险较低的候选药物。该测定法也是评估线粒体毒性特征和研究新化合物和市售化合物作用机制的有价值的工具。
