Reversal of Tau Pathology with MSUT2 siRNA Conjugates

项目摘要 “纯”tau蛋白病的遗传和病理学证据，如进行性核上性麻痹（PSP）， 皮质基底动脉变性（CBD）和额颞叶痴呆（FTD）的一些病例直接涉及tau蛋白 导致神经元细胞死亡，而在阿尔茨海默病（AD）中，tau蛋白的积累与发育相关 以及认知障碍的进展。由于（迄今为止）针对淀粉样蛋白β的药物失败，tau蛋白 现在成为寻找疾病修饰药物的“下一个最佳”治疗靶点。目前有 正在进行的2项在AD和PSP中使用抗tau抗体的2期临床研究， 正在开发的靶向制剂。在这里，我们提出了一种新的替代策略来改善神经元tau蛋白， 通过靶向MSUT 2基因表达的毒性。支持这种方法的知识体系包括 当在表达病理性tau蛋白的转基因动物中敲除MSUT 2时， tau种类和体外，其中敲低MSUT 2的siRNA是细胞保护性的。有效的体内递送 降低MSUT 2水平的siRNA缀合物将提供干预tau蛋白病的独特手段， 其优点是具有用于开发人类治疗剂可行途径。我们假设 siRNA介导的MSUT 2水平降低将在体内保护并可能逆转 由tau积累驱动的神经变性。DTx Pharma创造了一种新颖的专有脂肪酸基序 其在体内将siRNA递送到包括CNS神经元在内的多种细胞/组织中， 击倒。在这里，我们概述了与Brian Kraemer合作的药物发现计划的初始步骤， 使用DTx基序将MSUT 2特异性siRNA递送到体内神经元，以测试MSUT 2的敲低是否被 在充分表征的tau介导的变性的小鼠模型中具有神经保护作用。如果成功，这种方法 将被推进以产生铅，脂肪酸修饰的，MSUT 2 siRNA，用于潜在的治疗应用， tau蛋白病我们提出了3个具体目标：SA 1，MSUT 2 siRNA的产生和筛选，用于基因敲除 MSUT 2表达细胞中的活性。最有活性的siRNA与DTx脂肪酸基序的缀合， 在细胞系和原代神经元中的敲低效率的评价。SA 2，体内实验优化 给药，评价活性持续时间并评估DTx偶联的MSUT 2 siRNA在 野生型小鼠SA 3是在SA 2中鉴定的有效、安全和有效的siRNA，将用于治疗突变的tau蛋白。 在治疗试验中表达小鼠（PS19）。分析将包括记忆功能的测量， 磷酸化tau的积累、聚集/寡聚tau的沉积、神经炎症，以及 神经退行性变我们将比较siRNA介导的MSUT 2敲低对tau病理学的影响， 与先前使用抗tau抗体（C2 N/Abbvie）和tau靶向反义寡核苷酸的相同小鼠模型中的结果相同， 寡核苷酸（Ionis/Biogen），支持目前处于II期的新型治疗方法的进展 临床试验如果我们有类似或更好的活性，我们将寻求推进领先的MSUT 2靶向siRNA。

Project Summary Genetic and pathologic evidence in 'pure' tauopathies such as Progressive Supranuclear Palsy (PSP), Corticobasilar Degeneration (CBD) and some cases of Fronto-temporal Dementia (FTD) directly implicate tau as causing neuronal cell death, while in Alzheimer’s Disease (AD) tau accumulation correlates with development and progression of cognitive impairment. Because of failure (to date) of drugs targeting amyloid beta, tau has now emerged as 'the next best' therapeutic target in the search for disease modifying drugs. There are currently 2 active phase 2 clinical studies using anti-tau antibodies in both AD and PSP underway, with additional tau targeting agents under development. Here, we propose a novel, alternative strategy to ameliorate neuronal tau toxicity by targeting MSUT2 gene expression. The body of knowledge supporting this approach includes amelioration of tau toxicity in vivo when MSUT2 is knocked out in transgenic animals expressing a pathologic tau species and in vitro where siRNA's that knock down MSUT2 are cytoprotective. Effective in vivo delivery of siRNA conjugates that lower MSUT2 levels will provide a distinct means of intervening against tauopathy with the advantage of having a viable pathway forward for development of a human therapeutic. We hypothesize that siRNA-mediated reduction of MSUT2 levels will protect against and potentially reverse in vivo neurodegeneration driven by tau accumulation. DTx Pharma has created a novel, proprietary fatty acid motif that delivers siRNA in vivo into multiple cells/tissues, including CNS neurons, allowing efficient target gene knockdown. Here we outline the initial steps of a collaborative drug discovery program with Brian Kraemer that uses the DTx motif to deliver MSUT2 specific siRNA to neurons in vivo to test whether knockdown of MSUT2 is neuro-protective in a well-characterized mouse model of tau-mediated degeneration. If successful, this approach would be advanced to generate lead, fatty acid-modified, MSUT2 siRNAs for potential therapeutic application in tauopathies. We propose 3 specific aims: SA1, generation and screening of MSUT2 siRNAs for gene knockdown activity in MSUT2 expressing cells. Conjugation of the most active siRNAs to the DTx fatty acid motif for evaluation of knockdown efficiency in cell lines and primary neurons. SA2, in vivo experiments to optimize dosing, evaluate duration of activity and assess for potential toxicities of DTx-conjugated MSUT2 siRNAs in wildtype mice. SA3, a potent, safe and efficacious siRNA identified in SA2 will be used to treat mutant tau expressing mice (PS19) in a therapeutic trial. Analyses will include the measurement of memory function, accumulation of phosphorylated tau, deposition of aggregated/oligomeric tau, neuroinflammation, and neurofibrillary degeneration. We will compare the effect of siRNA mediated MSUT2 knockdown on tau pathology with prior results in the same mouse model using anti-tau antibodies (C2N/Abbvie) and tau-targeted antisense oligonucleotides (Ionis/Biogen) that supported advancement of novel therapeutics that are currently in phase 2 clinical trials. If we have similar or better activity—we will seek to advance a lead MSUT2 targeted siRNA.