Mitochondria share extensive evolutionary conservation across nearly all living species. This homology allows robust insights to be gained into pathophysiologic mechanisms and therapeutic targets for the heterogeneous class of primary mitochondrial diseases (PMDs) through the study of diverse in vitro cellular and in vivo animal models. Dramatic advances in genetic technologies, ranging from RNA interference to achieve graded knock‐down of gene expression to CRISPR/Cas‐based gene editing that yields a stable gene knock‐out or targeted mutation knock‐in, have enabled the ready establishment of mitochondrial disease models for a plethora of individual nuclear gene disorders. These models are complemented and extended by the use of pharmacologic inhibitor‐based stressors to characterize variable degrees, onset, duration, and combinations of acute on chronic mitochondrial dysfunction in individual respiratory chain enzyme complexes or distinct biochemical pathways within mitochondria. Herein is described the rationale for, and progress made in, “therapeutic cross‐training,” a novel approach meant to improve the validity and rigor of experimental conclusions when testing therapies by studying treatment effects in multiple, evolutionarily‐distinct species, including Caenorhabditis elegans (invertebrate, worm), Danio rerio (vertebrate, zebrafish), Mus musculus (mammal, mouse), and/or human patient primary fibroblast cell line models of PMD. The goal of these preclinical studies is to identify lead therapies from candidate molecules or library screens that consistently demonstrate efficacy, with minimal toxicity, in specific subtypes of mitochondrial disease. Conservation of in vitro and in vivo therapeutic effects of lead molecules across species has proven extensive, where molar concentrations found to be toxic or efficacious in one species are often consistent with therapeutic effects at similar doses seen in other mitochondrial disease models. Phenotypic outcome studies in all models are prioritized at the level of survival and function, to reflect the ultimate goal of developing highly potent therapies for human mitochondrial disease. Lead compounds that demonstrate significant benefit on gross phenotypes may be further scrutinized in these same models to decipher their cellular targets, mechanism(s), and detailed biochemical effects. High‐throughput, automated technologic advances will be discussed that enable efficient, parallel screening in a diverse array of mitochondrial disease disorders and overarching subclasses of compounds, concentrations, libraries, and combinations. Overall, this therapeutic cross‐training approach has proven valuable to identify compounds with optimal potency and safety profiles among major biochemical subtypes or specific genetic etiologies of mitochondrial disease. This approach further supports rational prioritization of lead compounds, target concentrations, and specific disease phenotypes, outcomes, and subgroups to optimally inform the design of clinical trials that test their efficacy in human mitochondrial disease subjects.

中文翻译：

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追求精准线粒体医学：利用临床前细胞和动物模型优化线粒体疾病治疗发现

线粒体在几乎所有生物物种中都有广泛的进化保护。通过对多种体外细胞和体内动物模型的研究，这种同源性使我们能够深入了解异质类原发性线粒体疾病 (PMD) 的病理生理机制和治疗靶点。遗传技术的巨大进步，从 RNA 干扰实现基因表达的分级敲低，到基于 CRISPR/Cas 的基因编辑产生稳定的基因敲除或靶向突变敲入，使线粒体疾病的建立成为可能多种个体核基因疾病的模型。这些模型通过使用基于药物抑制剂的压力源来补充和扩展，以表征不同程度、发作、持续时间、以及线粒体内单个呼吸链酶复合物或不同生化途径中急性对慢性线粒体功能障碍的组合。本文描述了“治疗性交叉训练”的基本原理和取得的进展，这是一种新方法，旨在通过研究多种进化上不同的物种的治疗效果来提高实验结论的有效性和严谨性，包括Caenorhabditis elegans（无脊椎动物、蠕虫）、斑马鱼（脊椎动物、斑马鱼）、Mus musculus（哺乳动物、小鼠）和/或人类患者的 PMD 原代成纤维细胞系模型。这些临床前研究的目标是从候选分子或文库筛选中确定先导疗法，这些疗法在线粒体疾病的特定亚型中始终表现出功效，且毒性最小。已证明在物种间保留先导分子的体外和体内治疗效果是广泛的，其中发现在一个物种中有毒或有效的摩尔浓度通常与在其他线粒体疾病模型中看到的类似剂量的治疗效果一致。所有模型中的表型结果研究都优先考虑生存和功能水平，以反映为人类线粒体疾病开发高效疗法的最终目标。可以在这些相同的模型中进一步检查对总体表型有显着益处的先导化合物，以破译其细胞靶点、机制和详细的生化效应。将讨论高通量、自动化的技术进步，这些进步能够在各种线粒体疾病和化合物、浓度、文库和组合的总体亚类中进行有效、平行的筛选。总体而言，这种治疗性交叉训练方法已被证明对于在主要生化亚型或线粒体疾病的特定遗传病因中鉴定具有最佳效力和安全性的化合物是有价值的。这种方法进一步支持对先导化合物、目标浓度和特定疾病表型、结果、