Focused Ultrasound-mediated Delivery of Gene-editing Elements to the Brain for Neurodegenerative Disorders

聚焦超声介导的基因编辑元件递送至大脑以治疗神经退行性疾病

• 批准号: 9810901
• 负责人: KAM W LEONG
• 金额: $ 79.85万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-01 至 2022-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9810901
• 关键词: Acoustics; Adverse effects; Alzheimer' s Disease; Alzheimer' s disease model; Amyloid beta-Protein; Amyloid beta-Protein Precursor; Astrocytes; Brain; Brain region; CRISPR/Cas technology; Cells; Clustered Regularly Interspaced Short Palindromic Repeats; Dependovirus; Development; Disease; Disease model; Elements; Expression Profiling; Focused Ultrasound; Gases; Gene Delivery; Generations; Genes; In Situ; In Vitro; Injections; Knock-out; Length; Mediating; Methodology; Microbubbles; Minor; Modeling; Mus; Neurodegenerative Disorders; Neurons; Parkinson Disease; Pathway interactions; Phase; Phenotype; Physiologic pulse; Plasmids; Play; Pluronics; Proteins; Proteolysis; Reporting; Rodent; Safety; Senile Plaques; Specificity; Structure; System; Techniques; Technology; Testing; Therapeutic; Tissues; Transcription Coactivator; Transcription Repressor/Corepressor; Treatment Efficacy; Viral Genome; Viral Packaging; Virion; adeno-associated viral vector; aged; base; beta-site APP cleaving enzyme 1; cell type; design; dopaminergic neuron; dosage; gene delivery system; immunogenicity; improved; improved outcome; in vivo; intravenous injection; mutant; non-viral gene delivery; nonhuman primate; novel therapeutic intervention; pressure; prevent; small molecule; success; therapeutic gene; transcription factor; vector

Focused Ultrasound-mediated Delivery of Gene-editing Elements to the Brain for Neurodegenerative Disorders Abstract CRISPR technology as a transformative toolkit for precise gene editing may tackle many intractable neurodegenerative diseases. However, delivery of CRISPR-based gene editing elements to the brain is highly inefficient. We previously pioneered the use of focused ultrasound (FUS) technology to achieve noninvasive, brain-specific delivery. We demonstrated that FUS in conjunction with monodispersed, gas-filled microbubbles could deliver therapeutic payloads, including small molecules, proteins and adeno-associated virus (AAV), to a specific region of the brain through intravenous injection in both rodent and non-human primate (NHP) models. Furthermore, we have also redesigned polyplex systems to improve the nonviral CRISPR-mediated gene editing efficiency and specificity both in vitro and in vivo. With those preliminary studies, we propose to integrate the merits of tissue-specific (FUS) and cell-specific (AAV and non-viral polyplex vectors) delivery systems to enable CRISPR delivery to the brain and its disease-related cell types. The overall objective of this project is to develop a non-invasive, FUS-mediated technology for delivering AAV vectors and non-viral polyplexes carrying CRISPR elements to the brain and to evaluate the efficacy on two major neurodegenerative disorders, Alzheimer’s and Parkinson’s diseases. We propose to pursue three specific aims in the UG3 developmental phase and a fourth aim in the UH3 demonstration phase: (1) Optimize the FUS system with defined microbubble composition, acoustic parameters and optimize the AAV vector carrying the CRISPR knockout, suppression as well as activation elements to achieve high gene editing efficacy in mouse brain; (2) Develop an efficient CRISPR delivery non-viral polyplex system with a transient expression profile for delivery into the brain via FUS technology; (3) Evaluate the therapeutic efficacy and the safety on both Alzheimer’s and Parkinson’s disease models with the optimized AAV and non-viral CRISPR delivery systems; and (4) Evaluate the delivery efficiency and safety of the optimized delivery system in the NHP model. As current CRISPR delivery to the brain relies only on intracranial injection, the proposed project will be the first study developing a noninvasive, efficient approach to achieve gene editing in the brain. Success of this project will stimulate new strategic approaches of tacking neurodegenerative disorders that remain challenging or even untreatable.

聚焦超声介导的基因编辑元件递送至大脑以治疗神经退行性疾病 抽象的 CRISPR 技术作为精确基因编辑的变革工具包可能会解决许多棘手的神经退行性疾病 疾病。然而，将基于 CRISPR 的基因编辑元件传递到大脑的效率非常低。我们之前 率先使用聚焦超声 (FUS) 技术来实现无创、针对大脑的递送。我们 证明 FUS 与单分散、充气微泡结合可以提供治疗有效负载， 包括小分子、蛋白质和腺相关病毒（AAV），通过静脉注射到达大脑的特定区域 在啮齿动物和非人类灵长类动物（NHP）模型中进行注射。此外，我们还重新设计了 Polyplex 系统 提高非病毒 CRISPR 介导的基因编辑在体外和体内的效率和特异性。与那些 初步研究中，我们建议整合组织特异性（FUS）和细胞特异性（AAV 和非病毒）的优点 Polyplex 载体）递送系统，使 CRISPR 能够递送至大脑及其疾病相关细胞类型。整体 该项目的目标是开发一种非侵入性、FUS 介导的技术，用于传递 AAV 载体和非病毒 将 CRISPR 元件携带到大脑的多聚体，并评估对两种主要神经退行性疾病的功效， 阿尔茨海默病和帕金森病。我们建议在 UG3 发展阶段追求三个具体目标 UH3 演示阶段的第四个目标：(1) 通过确定的微泡成分、声学特性来优化 FUS 系统 参数并优化携带 CRISPR 敲除、抑制和激活元件的 AAV 载体 在小鼠大脑中实现高基因编辑效率； (2) 开发高效的 CRISPR 递送非病毒复合物系统 通过 FUS 技术传递至大脑的瞬时表达谱； (3) 疗效评价及疗效评价 通过优化的 AAV 和非病毒 CRISPR 传递系统，确保阿尔茨海默病和帕金森病模型的安全性； (4)评估NHP模型中优化后的输送系统的输送效率和安全性。作为目前的 CRISPR 输送到大脑仅依赖于颅内注射，拟议的项目将是第一个开发 非侵入性、有效的方法来实现大脑中的基因编辑。该项目的成功将激发新的战略 治疗仍然具有挑战性甚至无法治愈的神经退行性疾病的方法。