Evaluating the utility of hepatic in vitro models for the assessment of the multi-mechanistic toxicity of drug-induced liver injury

药品不良反应(ADR)是国家卫生服务(NHS)的主要负担，2004年占住院人数的6.5%。据估计，由于ADR是第七大死因，NHS每年要花费4.66亿英镑。然而，ADR不仅仅是医院的问题；它们代表着一个主要的制药问题。2015年，据估计，将一种药物从替补推向市场的成本为26亿美元。1975-1999年间，在获得批准的548种新药中，10.2%获得了黑盒警告或因ADR而退出市场。出于这个原因，有一个巨大的制药问题，因为数百万英镑和时间可能被浪费。虽然ADR会影响身体的不同器官，但肝脏是报道最多的两个病例之一。药物性肝损伤(DILI)表现为多种病理状态，20-40%的报告病例会导致胆汁淤积。药物诱导的线粒体毒性(DIMT)已被报道为DILI的决定因素，有50%的药物黑盒警告DILI也有线粒体负担。DILI与多种毒性机制有关。因此，不能采用‘一刀切’的方法，这增加了临床前检测DILI的难度。然而，线粒体毒性和胆汁转运蛋白的影响被认为是肝脏毒性的主要机制。线粒体具有丰富的结构和功能特征，可被化合物靶向并导致毒性。这些可能包括电子传递链抑制，氧化磷酸化解偶联，线粒体通透性转换(MPT)孔的开放，线粒体动力学的改变和线粒体基因组的枯竭。由于线粒体有自己的基因组，线粒体DNA(MtDNA)变异对DILI易感性的影响是另一个需要研究的潜在因素。该项目的目的是开发和验证用于评估帝力线粒体毒性和转运体功能障碍的筛查模型。HepaRG细胞是来源于人肝祖细胞系的终末分化的肝细胞，它保留了原代人肝细胞(PHH)的许多特征，包括异种代谢酶、药物转运体和功能性胆管结构。可以操纵HepaRG细胞来绕过克拉布特里效应，从而可以研究线粒体毒性。因此，将评估HepaRG细胞连接线粒体和转运蛋白功能障碍之间的相互作用的能力。此外，首个针对线粒体遗传变异的个人化、肝脏特异性体外模型的开发，将使人们能够对个体对DILI的易感性的影响进行新的研究。对于DILI及其与线粒体毒性和转运体改变的联系，更具预测性和敏感性的模型将在预防晚期药物磨损和开发更安全的药物方面证明是非常宝贵的。

Adverse drug reactions (ADRs) are a major burden to the National Health Services (NHS), accounting for 6.5 % of hospital admissions in 2004. It is estimated that ADRs cost the NHS £466 million annually due to them being the 7th leading cause of death. However, ADRs are not just a hospital problem; they represent a major pharmaceutical concern. In 2015, it was estimated that the cost of getting a drug from bench to market cost $2.6 billion. During 1975 - 1999, of the 548 new drugs that were approved, 10.2 % acquired black box warnings or were withdrawn from the market due to ADRs. For this reason, there is a great pharmaceutical concern as millions of pounds and time could be wasted. Whilst ADRs can affect different organs in the body, the liver is one of the two most reported cases. Drug-induced liver injury (DILI) presents itself as a variety of pathological conditions, with 20 - 40 % of reported cases causing cholestasis. Drug-induced mitochondrial toxicity (DIMT) has been reported as a determinant of DILI with 50 % of drugs with black box warnings for DILI also having mitochondrial burdens. DILI is associated with multiple mechanisms of toxicity. For this reason, a 'one test fits all' approach cannot be used, which has contributed to the difficulty in detecting DILI preclinically. Nevertheless, mitochondrial toxicity and biliary transporter implications are recognised as major mechanisms of hepatotoxicity. The mitochondria have a wealth of structural and functional features which can be targeted by a compound and lead to toxicity. These can include electron transport chain inhibition, oxidative phosphorylation uncoupling, opening of the mitochondrial permeability transition (MPT) pore, alterations in mitochondrial dynamics and the depletion of the mitochondrial genome. Due to the mitochondria having their own genome, the effects of mitochondrial DNA (mtDNA) variation upon susceptibility to DILI is another potential factor requiring investigation. The aim of the project is to develop and validate screening models for the assessment of mitochondrial toxicity and transporter dysfunction in DILI. HepaRG cells are terminally differentiated hepatic cells derived from a human hepatic progenitor cell line that retains many characteristics of primary human hepatocytes (PHH), including xenobiotic metabolism enzymes, drug transporters and functional biliary structures. HepaRG cells can be manipulated to circumvent the Crabtree effect, allowing investigations of mitochondrial toxicity. Therefore, the ability of HepaRG cells to connect the interplay between mitochondria and transporter dysfunction will be evaluated. Additionally, the development of the first personalised, liver-specific in vitro model for mitochondrial genetic variation, coined transmitochondrial HepG2 cybrids will allow novel investigations of the effects of individual susceptibility to DILI. More predictive and sensitive models for DILI and their links with mitochondrial toxicity and transporter alterations would prove invaluable in the prevention of late-stage drug attrition and the development of safer drugs.