Engineered AAV vectors for combinatorial treatment of rare genetic brain diseases

用于罕见遗传性脑部疾病组合治疗的工程 AAV 载体

• 批准号: 10414342
• 负责人: VIRGINIA Eunice KIMONIS
• 金额: $ 60万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10414342
• 关键词: Engineered; AAV; vectors; combinatorial; treatment

Project Summary AAV-based gene therapy requires the development of safe, efficient, and target-specific vectors. AAV-mediated gene therapy for peripheral tissues (blood and skeletal muscle) has made great strides, focused mainly on gene replacement for loss of function diseases. However, there has been little research on dominantly inherited CNS disorders that are caused by toxic gene products. We have assembled a world class, multi-disciplinary academic research team supported by our industrial partners to develop innovative AAV-based gene knockdown and replacement treatments for rare genetic diseases including Spinocerebellar Ataxia Type 7 (SCA7) and valosin- containing protein (VCP) multisystem proteinopathy. We respond to the RFA Project Objectives and propose three Specific Aims. In Aim 1, we will design and manufacture new AAV vectors with improved critical quality attributes (safety, efficacy, target specificity) for gene therapy. We propose innovative neuron specific gene delivery, temporal control of gene expression and reduced immune responses in the CNS. We have constructed AAV vectors that express EGFP and mRNA barcodes for improved screening to support our proposed AAV treatments. In Aim 2, we will develop advanced quantitative analytics by combining next-generation sequencing and bar-coded AAVs for efficient assessment of in vivo gene delivery in the mouse model. We will screen and compare AAV-mediated gene expression with different capsid variants using different promoters / enhancers. Single-cell RNAseq also will be used to assess immunological responses of selected AAV vectors by different administration routes. In Aim 3, to effectively treat the SCA7 and VCP diseases, we will develop novel AAV vectors that simultaneously knockdown toxic gene products while replacing normal gene products that are codon optimized to be unaffected by knockdown. This proposed combinatorial treatment will establish a proof-of- concept for many other dominant inherited diseases, where loss of normal allele expression due to non-specific silencing causes its own problems. We have well developed models of the SCA7 and VCP diseases for Aim 3 studies. Our team has great expertise in developing, manufacturing and applying AAV vectors in basic research and preclinical application. Our published work and preliminary data establish the feasibility and key methodologies for the proposed research. Critically, we have an established viral production facility and distribution platform operating out of our UCI Center for Neural Circuit Mapping that supports viral reagent design, validation, and manufacturing.

项目概要 基于 AAV 的基因治疗需要开发安全、高效且针对特定目标的载体。 AAV介导的 外周组织（血液和骨骼肌）的基因治疗取得了长足的进步，主要集中在基因治疗上 替代功能丧失疾病。然而，对于显性遗传性中枢神经系统的研究却很少。 由有毒基因产物引起的疾病。我们聚集了世界一流的多学科学术团队 研究团队在我们的工业合作伙伴的支持下开发基于 AAV 的创新基因敲除和 罕见遗传病的替代疗法，包括脊髓小脑性共济失调 7 型 (SCA7) 和 valosin- 含蛋白（VCP）多系统蛋白病。我们响应 RFA 项目目标并提出建议 三个具体目标。在目标 1 中，我们将设计和制造具有更高关键质量的新 AAV 载体 基因治疗的属性（安全性、有效性、靶标特异性）。我们提出创新的神经元特异性基因 传递、基因表达的时间控制以及中枢神经系统中免疫反应的减少。我们已经建造了 表达 EGFP 和 mRNA 条形码的 AAV 载体，用于改进筛选以支持我们提出的 AAV 治疗。在目标 2 中，我们将通过结合下一代测序来开发先进的定量分析 和条形码 AAV，用于有效评估小鼠模型中的体内基因传递。我们将筛选并 使用不同的启动子/增强子比较 AAV 介导的基因表达与不同衣壳变体。 单细胞 RNAseq 还将用于评估不同类型 AAV 载体的免疫反应 给药途径。在目标3中，为了有效治疗SCA7和VCP疾病，我们将开发新型AAV 载体可同时敲除有毒基因产物，同时替换作为密码子的正常基因产物 优化后不受击倒的影响。这项拟议的组合治疗将建立一个证明- 许多其他显性遗传疾病的概念，其中由于非特异性而导致正常等位基因表达的丧失 沉默会导致其自身的问题。我们为目标 3 开发了完​​善的 SCA7 和 VCP 疾病模型 研究。我们的团队在 AAV 载体的开发、制造和基础研究应用方面拥有丰富的专业知识 和临床前应用。我们发表的工作和初步数据确立了可行性和关键 拟议研究的方法。至关重要的是，我们拥有成熟的病毒生产设施，并且 在我们的 UCI 神经回路映射中心运行的分销平台，支持病毒试剂设计， 验证和制造。