Translating Pathomechanisms into Treatment for Spinal Muscular Atrophies

项目总结 脊髓性肌萎缩症(SMA)是导致衰弱的单基因运动神经元(MN)疾病 肌肉无力，常常过早死亡。我的研究计划专注于推动两人疗法的发展 SMA的形式：由存活运动神经元的隐性、功能丧失突变引起的近端SMA 1 瞬时受体电位显性突变引起的SMN1基因和远端SMA基因 香草素4基因(TRPV4)。我们最重要的方法是将人类患者的发现与 在动物和IPSC衍生模型上的实验，以阐明与 人类疾病，并确定有希望的治疗机会。在这里，我们将利用独特的资源和 最先进的技术来定义限制当前SMA疗法疗效的因素，表征细胞 以及驱动SMA病理的分子机制，并识别和验证新的治疗策略。 近端SMA处于快速发展的基因靶向治疗的前沿，最近批准了两种 其中一种是SMN诱导治疗，第三种正在FDA审查中。虽然取得了变革性的成功，但 这些治疗方法差异很大，从正常达到早期运动里程碑到不治疗。 运动功能的改善。在过去的5年里，我们的研究揭示了近端SMA的病理 从子宫开始，在目前患者开始治疗之前。在人类和小鼠中，SMA MNS 表现出妊娠期成熟受损和新生儿急剧变性，同时伴有明显的 SMN表达下降。在这里，我们将在这些观察的基础上1)剖析具体的机制 在发育和治疗过程中调节SMN的表达，2)确定导致 SMA MN的成熟和退化受损，以及3)利用这些见解开发新的和在子宫内的 SMA治疗策略。在对dsma的平行研究中，我们最近证明了神经致病 细胞表面阳离子通道TRPV4的突变扰乱了调节蛋白质-蛋白质的相互作用，并导致 通道函数的增益。现有的TRPV4拮抗剂在人体内具有良好的耐受性，使该通道成为 前景看好的治疗靶点。引人注目的是，突变的TRPV4基因敲入小鼠模型会发展成严重的神经系统 由选择性遗传挽救的血-神经屏障(BNB)局灶性崩溃所致的表型 从内皮细胞(ECs)中删除TRPV4或用TRPV4拮抗剂治疗有症状的小鼠。 这些研究表明，TRPV4的激活可以通过以下方式以非细胞自主的方式驱动神经病理 监管BNBs。在这里，我们将1)描述调节TRPV4通道活性的蛋白质相互作用，2) 评估TRPV4在调节EC屏障功能中的作用，以及3)评估TRPV4拮抗剂作为 Dsma小鼠的治疗策略以及最终以BNB中断为特征的其他疾病。一起， 我们的研究将进一步加深我们对SMA病理机制的理解，勾画出新的治疗靶点 和策略，并推进对SMAS和相关神经肌肉疾病患者的护理。

PROJECT SUMMARY Spinal muscular atrophies (SMAs) are monogenetic motor neuron (MN) diseases that cause debilitating muscle weakness and often early mortality. My research program focuses on advancing therapeutics for two forms of SMA: proximal SMA caused by recessive, loss-of-function mutations of the survival motor neuron 1 gene (SMN1) and distal SMA (dSMA) caused by dominant mutations of the transient receptor potential vanilloid 4 gene (TRPV4). Our overarching approach is to integrate findings from human patients with experimentation in animal and iPSC-derived models to elucidate pathomechanistic pathways relevant to human disease and identify promising therapeutic opportunities. Here, we will leverage unique resources and state-of-the-art technologies to define factors limiting efficacy of current SMA therapeutics, characterize cellular and molecular mechanisms driving SMA pathology, and identify and validate novel therapeutic strategies. Proximal SMA is at the forefront of rapidly evolving gene-targeting therapeutics, with two recently approved SMN-inducing treatments and a third under FDA review. While a transformative success, the clinical efficacy of these treatments is highly variable, ranging from normal attainment of early motor milestones to no improvement in motor function. In the last 5 years, our studies have revealed that proximal SMA pathology begins in utero, before treatments are currently initiated in patients. In both humans and mice, SMA MNs exhibit impaired maturation during gestation and precipitous neonatal degeneration, paralleled by a marked decline in SMN expression. Here, we will build on these observations to 1) dissect the specific mechanisms regulating SMN expression during development and treatment, 2) identify the molecular mechanisms causing impaired maturation and degeneration of SMA MNs, and 3) use these insights to develop novel and in utero SMA therapeutic strategies. In parallel studies on dSMA, we have recently demonstrated that neuropathogenic mutations in TRPV4, a cell surface cation channel, disrupt regulatory protein-protein interactions and cause a gain of channel function. Existing TRPV4 antagonists have good tolerability in humans, making the channel a promising therapeutic target. Strikingly, mutant TRPV4 knock-in mouse models develop severe neurological phenotypes due to focal breakdown of blood-neural barriers (BNBs), which are rescued by selective genetic deletion of TRPV4 from endothelial cells (ECs) or treatment of symptomatic mice with TRPV4 antagonists. These studies suggest that TRPV4 activation can drive neuropathology in a non-cell autonomous manner by regulating BNBs. Here, we will 1) characterize protein interactions regulating TRPV4 channel activity, 2) evaluate the role of TRPV4 in modulating EC barrier function, and 3) assess TRPV4 antagonists as a therapeutic strategy in dSMA mice and ultimately other disorders characterized by BNB disruption. Together, our studies will further our mechanistic understanding of SMA pathology, delineate novel therapeutic targets and strategies, and advance care of patients with SMAs and related neuromuscular diseases.