Data Sheet 1_Peptide-mimetics derived from leucyl-tRNA synthetase are potential agents for the therapy of mt-tRNA related diseases.pdf

IntroductionMitochondrial diseases caused by point mutations in mitochondrial tRNA (mt-tRNA) genes, including MELAS and MERRF syndromes, represent a significant unmet clinical need, due to the lack of effective treatments. We previously identified peptide molecules derived from human leucyl-tRNA synthetase, whose features make them attractive leads for the development of therapeutic agents against mt-tRNA point mutations-related diseases. Indeed, we demonstrated that, upon exogenous administration, these peptides penetrate human cell and mitochondrial membranes; stabilize mitochondrial tRNA structures; and rescue severe mitochondrial defects in cells bearing the point mutations m.3243A>G and m.8344A>G, responsible for MELAS and MERRF syndromes, respectively.
ResultsTo progress towards therapeutic applications, in this work we designed three peptide-mimetic derivatives (PMTs). These are composed entirely of D-amino acids and potentially endowed with enhanced stability in human plasma and resistance to enzymatic degradation. We show that, like the parent peptide, the PMTs have mitochondrial localization and improve cell viability and oxygen consumption in human cybrid cell lines bearing the aforementioned point mutations. Additionally, as anticipated, the PMTs had significantly higher plasma stability than the parent peptide. The most promising PMT was radiolabelled with Cu-64 and used in in vivo biodistribution and tolerability studies. Importantly, i. v. administered PMT reached all body districts, including heart, muscle and even brain, thus revealing an intrinsic ability to cross the blood-brain barrier. Finally, PMT was safe in adult wild-type mice at dosages up to 10 mg/kg.
DiscussionThese findings represent a significant step towards the implementation of therapeutic strategies for mttRNA-related mitochondrial diseases.

## 引言
由线粒体转运RNA（mitochondrial tRNA, mt-tRNA）基因点突变引发的线粒体疾病，涵盖MELAS综合征与MERRF综合征，因缺乏有效治疗手段，存在显著的未被满足的临床需求。我们此前已鉴定出源自人类亮氨酰-tRNA合成酶的肽类分子，其特性使其成为针对线粒体转运RNA点突变相关疾病的治疗药物开发的极具吸引力的先导化合物。事实上，我们已证实，经外源性给药后，此类肽类可穿透人类细胞膜与线粒体膜；稳定线粒体转运RNA结构；并挽救携带分别引发MELAS与MERRF综合征的m.3243A>G、m.8344A>G点突变的细胞中的严重线粒体功能缺陷。

## 结果
为推进至治疗应用阶段，本研究设计了三种肽模拟衍生物（peptide-mimetic derivatives, PMTs）。此类衍生物完全由D-氨基酸构成，理论上具备更高的人血浆稳定性与抗酶降解能力。我们的实验结果显示，与母本肽一致，PMTs可定位于线粒体，并可改善携带上述点突变的人类胞质杂合细胞系的细胞存活率与耗氧量。此外，如预期所示，PMTs的血浆稳定性显著高于母本肽。我们选取最具开发前景的PMT，采用铜-64（Cu-64）进行放射性标记，并开展了体内生物分布与耐受性研究。重要的是，经静脉给药的PMT可抵达全身各组织部位，包括心脏、肌肉甚至脑部，这表明其具备跨越血脑屏障的内在能力。最终，在剂量高达10 mg/kg时，PMT对成年野生型小鼠无明显毒性。

## 讨论
上述发现为实施针对线粒体转运RNA相关线粒体疾病的治疗策略迈出了重要一步。