Fang Minggang, Banday Shahid, Deibler Sara K, Simone Tessa M, Coleman Madison, O'Connor Emerald, Li Rui, Zhu Lihua Julie, Green Michael R
Departments of Molecular, Cell and Cancer Biology, University of Massachusetts Chan Medical School, Worcester, Massachusetts 01605
Departments of Molecular, Cell and Cancer Biology, University of Massachusetts Chan Medical School, Worcester, Massachusetts 01605.
J Neurosci. 2025 Jun 25;45(26):e2307242025. doi: 10.1523/JNEUROSCI.2307-24.2025.
Friedreich ataxia (FA) is an autosomal recessive disease characterized by progressive damage to the nervous system and severe cardiac abnormalities. The disease is caused by a GAA•TTC triplet repeat expansion in the first intron of the gene, resulting in epigenetic repression of transcription and reduction in FXN (frataxin) protein which results in mitochondrial dysfunction. Factors and pathways that promote repression represent potential therapeutic targets whose inhibition would restore transcription and frataxin protein levels. Here, we performed a candidate-based RNAi screen to identify kinases, a highly druggable class of proteins, that when knocked down upregulate expression. Using this approach, we identified Rho kinase ROCK1 as a critical factor required for repression. ShRNA-mediated knockdown of ROCK1, or the related kinase ROCK2, increases mRNA and frataxin protein levels in FA patient-derived induced pluripotent stem cells (iPSCs) and differentiated neurons and cardiomyocytes to levels observed in normal cells. We demonstrate that small molecule ROCK inhibitors, including the FDA-approved drug belumosudil and fasudil, reactivate expression in cultured FA iPSCs, neurons, cardiomyocytes, and FA patient primary fibroblasts and ameliorate the characteristic mitochondrial defects in these cell types. Remarkably, treatment of transgenic FA mice of both sexes with belumosudil or fasudil upregulates expression, ameliorates the mitochondrial defects in the brain and heart tissues, and improves motor coordination and muscle strength. Collectively, our study identifies ROCK kinases as critical repressors of expression and provides preclinical evidence that FDA-approved ROCK inhibitors may be repurposed for treatment of FA.
弗里德赖希共济失调（FA）是一种常染色体隐性疾病，其特征为神经系统进行性损伤和严重心脏异常。该疾病由基因第一内含子中的GAA•TTC三联体重复扩增引起，导致转录的表观遗传抑制以及FXN（共济蛋白）蛋白减少，进而造成线粒体功能障碍。促进这种抑制的因子和信号通路代表了潜在的治疗靶点，抑制这些靶点可恢复转录和共济蛋白水平。在此，我们进行了一项基于候选基因的RNA干扰筛选，以鉴定激酶（一类具有高度可成药潜力的蛋白质），这些激酶在敲低后可上调基因表达。通过这种方法，我们确定Rho激酶ROCK1是基因抑制所需的关键因子。短发夹RNA介导的ROCK1或相关激酶ROCK2的敲低，可使FA患者来源的诱导多能干细胞（iPSC）以及分化的神经元和心肌细胞中的基因mRNA和共济蛋白水平增加至正常细胞中的水平。我们证明，包括美国食品药品监督管理局（FDA）批准的药物贝林妥欧单抗和法舒地尔在内的小分子ROCK抑制剂，可在培养的FA iPSC、神经元、心肌细胞以及FA患者原代成纤维细胞中重新激活基因表达，并改善这些细胞类型中的特征性线粒体缺陷。值得注意的是，用贝林妥欧单抗或法舒地尔治疗两性转基因FA小鼠，可上调基因表达，改善脑和心脏组织中的线粒体缺陷，并改善运动协调性和肌肉力量。总的来说，我们的研究确定ROCK激酶是基因表达的关键抑制因子，并提供了临床前证据表明FDA批准的ROCK抑制剂可重新用于治疗FA。
