Activation of Soluble Adenylyl Cyclase as a Novel Therapeutic Strategy for the Treatment of Age-Related Neurodegenerative Disorders

项目摘要/摘要 全世界65岁以上的人口正在继续增长，预计还会翻一番 在接下来的30年里。由于这种增加，与年龄相关的神经退行性疾病的相关增加 (NDS)，如阿尔茨海默氏症(AD)和帕金森氏病，已经被观察到。尽管进行了数十年的研究， 目前还没有FDA批准的可以阻止或逆转疾病进展的疗法。积累 错误折叠的蛋白质聚集体是年龄相关性NDS的共同特征，被认为与 在疾病的病理生理学方面。蛋白质聚集体可以通过自噬从细胞中清除， 生物降解的主要途径之一。蛋白质聚集体在最后一步中被降解 自噬，它们被运送到称为溶酶体的细胞器中。溶酶体维持酸性 PH在4-5之间，以保持酸性水解酶的最佳环境。在老化的细胞中和在细胞模型中 AD，溶酶体变得不那么酸性，它们的水解酶变得不那么活跃，结果是， 通过自噬(即自噬通量)和未消化物质的积累而降解。在模型中 在AD病理中，通过添加外源性cAMP，溶酶体pH重新酸化，逆转了这一表型。在……里面 与这些发现一致，我们已经证明cAMP产生于胞浆中的腺酰环化酶同功体，已知 作为可溶性腺酰环化酶(SAC)，促进溶酶体酸化。此外，不表达SAC的细胞 显示自噬通量降低。SAC的这一生理作用以及其他生理作用已通过使用 为研究SAC生物学而开发的各种抑制剂和各种遗传工具。然而，“工具箱”也就是 目前用于研究SAC的药物缺乏一个关键成分：SAC的药理激活剂。识别SAC的步骤 激活剂，我们进行了400,000种不同化学成分的高通量筛选。从这个图书馆 我们发现了13个可能的SAC激活剂。在这项建议中，我描述了体外和基于细胞的分析， 我将用来确认、表征并进一步发展这13个化合物，使其成为“第一个”小分子。 对SAC有选择性的激活剂。使用新发现的SAC激活剂，我将测试刺激 SAC可以增强溶酶体的酸化，刺激自噬，从而减少溶酶体的蓄积 蛋白质聚集体。如果成功，这些研究将证实小分子SAC激活剂是一种潜在的新型药物 治疗神经退行性疾病的治疗策略。

Project Summary/Abstract The worldwide population of individuals over the age of 65 is continuing to grow and is expected to double over the next 30 years. Due to this increase, an associated rise in age-related neurodegenerative diseases (NDs), such as Alzheimer’s (AD) and Parkinson’s disease, has been observed. Despite decades of research, there are currently no FDA-approved therapies that can stop or reverse disease progression. The accumulation of misfolded protein aggregates is a common feature of age-related NDs and is thought to be heavily involved in the pathophysiology of the diseases. Proteins aggregates can be cleared from the cell through autophagy, one of the major biological degradation pathways. Protein aggregates are degraded in the final step of autophagy, where they are delivered to cellular organelles known as lysosomes. Lysosomes maintain an acidic pH between 4-5 to maintain an optimal environment for acid hydrolases. In aging cells and in a cellular model of AD, lysosomes become less acidic, their hydrolases become less active and, as a result, there is a decrease in degradation through autophagy (i.e., autophagic flux) and an accumulation of undigested materials. In models of AD pathology, re-acidification of lysosomal pH, via addition of exogenous cAMP, reversed this phenotype. In line with these findings, we have shown that cAMP generated from a cytosolic adenylyl cyclase isoform, known as Soluble Adenylyl Cyclase (sAC), promotes lysosomal acidification. In addition, cells that do not express sAC show decreased autophagic flux. This, and other, physiological roles of sAC have been determined via the use of inhibitors and various genetic tools that were developed to study sAC biology. However, the “toolbox” that is currently used to study sAC is lacking a key component: a pharmacological activator of sAC. To identify a sAC activator, we conducted a high-throughput screen of 400,000 chemically diverse compounds. From this library we discovered 13 presumptive sAC activators. In this proposal, I describe the in vitro and cell-based assays that I will use to confirm, characterize, and further develop these 13 compounds as “first-of-their kind” small molecule activators selective for sAC. Using the newly discovered sAC activators, I will test the hypothesis that stimulating sAC can enhance lysosomal acidification and stimulate autophagy, and as a result, decrease accumulation of protein aggregates. If successful, these studies will validate small molecule sAC activators as a potential novel therapeutic strategy to treat neurodegenerative disorders.