Myosin Activation as a Novel Therapeutic Strategy for Polycystic Kidney Disease

摘要 常染色体显性多囊肾病（ADPKD）影响全世界每400-1000人中就有1人， 导致肾脏、肝脏和其他器官中逐渐产生充满液体的囊肿，导致器官衰竭。 PKD是由多囊蛋白多囊蛋白-1（PC 1）或多囊蛋白-2（PC 2）突变引起的。一个明确 对于多囊蛋白功能障碍如何在机制上导致囊肿的理解仍然是难以捉摸的，部分原因是， 缺乏精确表型复制疾病的包囊产生模型系统。干细胞衍生的人肾 类器官以特定的方式模拟PKD囊肿发生，并有可能加速药物发现， PKD的发展。 使用这些PKD类器官，进行药物筛选，将肌球蛋白抑制剂blebbistatin鉴定为药物。 强效包囊激动剂随后，我发现EMD，一种肌球蛋白激活剂，显示出临床前的希望， 在疾病的早期和晚期治疗PKD类器官中的PKD囊肿形成。根据初步 根据文献中的数据和线索，我假设PC 1和PC 2正调控非肌肉肌球蛋白 小管上皮内的收缩性，以防止囊肿形成。 利用独特的肾类器官体外系统，我建议分析肌球蛋白调节剂化合物， 类器官中的非肌肉肌球蛋白动力学，以确定控制囊肿扩张的肌球蛋白结构域， 告知临床前药物开发。第二，我的目标是优化体内递送和治疗策略， PKD小鼠模型中的EMD。最后，我将利用PC 1在结构上类似于非典型的 粘附G-蛋白偶联受体，以分析生物化学激活的PC 1是否可以刺激类器官 收缩以将多囊蛋白功能与肌球蛋白控制联系起来。总之，这些实验将建立肌球蛋白 激活作为治疗PKD的治疗策略，并确定PC 1作为肌球蛋白调节剂的机制 收缩性。这项工作的目标是通过利用最好的模型系统来加速PKD的药物发现， 代表疾病，以及告知多囊蛋白在囊形成中的功能的机制研究。

ABSTRACT Autosomal dominant polycystic kidney disease (ADPKD) affects 1 in every 400-1000 people worldwide, causing progressive fluid-filled cyst production in the kidneys, liver, and other organs, resulting in organ failure. PKD is caused by mutations in the polycystin proteins polycystin-1 (PC1) or polycystin-2 (PC2). A clear understanding of how polycystin dysfunction leads to cysts mechanistically is still elusive, due, in part, to the lack of cyst producing model systems that accurately phenocopy the disease. Stem cell derived human kidney organoids model PKD cystogenesis in a specific way and have potential to accelerate drug discovery and development for PKD. Using these PKD organoids, a drug screen was performed identifying blebbistatin, a myosin inhibitor, as a potent cyst agonist. Subsequently, I discovered that EMD, a myosin activator, showed pre-clinical promise in treating PKD cystogenesis in PKD organoids at both early and late stages of disease. Based upon preliminary data and clues from the literature, I hypothesize that PC1 and PC2 positively regulate non-muscle myosin contractility within the tubular epithelium to prevent cystogenesis. Utilizing the unique kidney organoid in vitro system, I propose to analyze myosin modulator compounds and non-muscle myosin dynamics in organoids to pinpoint the myosin domain that governs cyst expansion to inform pre-clinical drug development. Second, I aim to optimize in vivo delivery and treatment strategies of EMD in a mouse model of PKD. Finally, I will utilize the insight that PC1 structurally resembles an atypical adhesion G-protein coupled receptor to analyze whether biochemically activating PC1 can stimulate organoid contraction to link polycystin function to myosin control. Together, these experiments will establish myosin activation as a therapeutic strategy to treat PKD and determine mechanisms of PC1 as a regulator of myosin contractility. The goal of this work is to expedite drug discovery for PKD by utilizing model systems that best represent the disease, and mechanistic studies that inform polycystin function in cystogenesis.