Circuit-Specific Delivery of Large Cargo Across the Nervous Systems of Adult Mammals and Embryos via Novel Engineered Systemic Vectors

通过新型工程系统载体在成年哺乳动物和胚胎的神经系统中进行大型货物的特定电路递送

• 批准号: 9789711
• 负责人: Viviana Gradinaru
• 金额: $ 117.25万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-30 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9789711
• 关键词: Adult; Anatomy; Basic Science; Blood; Blood - brain barrier anatomy; Blood Circulation; Brain; Brain Pathology; CRISPR/Cas technology; Cells; Central Nervous System Diseases; Clustered Regularly Interspaced Short Palindromic Repeats; Dependovirus; Development; Disease; Embryo; Engineering; Gene Delivery; Genes; Genetic; Genome; Genome engineering; Image; Injections; Laboratories; Life; Longevity; Mammals; Modality; Nature; Nerve Degeneration; Nervous system structure; Placenta; Population; RNA Interference; Research; Rodent; Specificity; Technology; Therapeutic; Therapeutic Studies; Tissues; Viral; Viral Vector; Virus; aging brain; base; cell type; gene therapy; genome editing; in utero; in vivo; neural circuit; neurodevelopment; neurotechnology; novel; optogenetics; pregnant; repaired; tool; vector

Viviana Gradinaru, Caltech With the advent of technologies such as CRISPR/Cas9, genome engineering for both basic research and therapeutic applications is becoming reality. An outstanding challenge is the mean to safely and efficiently transfer large genomes to desired cells across life span. We have developed an in vivo Cre-based selection platform (CREATE) for identifying adeno-associated viruses (AAVs) that efficiently transduce genetically defined populations. We used CREATE to select for viruses that transduce the brain after intravascular delivery and found a vector that nonspecifically transduces most cells across the adult brain. Since the restrictive nature of the blood brain barrier presents a major impediment toward treating CNS disorders our discovery has the potential to enable exciting advances in gene editing/replacement via CRISPR-Cas or RNA interference to restore diseased CNS circuits if the needed level of efficiency and specificity can be engineered for diseased targets. We plan to enable such efforts by creating viral-based solutions to non-invasive whole- brain large cargo delivery across the blood-brain barrier from embryo to adult by: 1. Generating AAVs for cell-type and region specific gene delivery across the blood-brain- barrier, noninvasively via the bloodstream in the adult rodent for neurodegeneration applications. 2. Generate AAVs capable of transducing the developing brain in utero with a simple systemic injection to the pregnant dam for neurodevelopment research and therapy. 3. Increase the packaging capability of AAVs by about 2-fold to enable delivery of large genomes for gene therapy and research. 4. Enable non-invasive circuit specific deep brain modulation by the use of systemic vectors and genetically encoded activity modulators (e.g. by chemogenetics or others in development now). Longer term we plan, in our laboratory and also with collaborators, to contribute our neurotechnologies (including, in addition to viral vectors, tissue clearing and optogenetic control and imaging) towards elucidating maladaptive neural circuits that contribute to brain pathology in neurodegeneration and neurodevelopment.

维维安娜·格拉迪纳鲁，加州理工学院 随着 CRISPR/Cas9 等技术的出现，基因组工程 基础研究和治疗应用正在成为现实。一项杰出的挑战 是将大型基因组安全有效地转移到生命周期中所需细胞的方法 跨度。我们开发了一个基于 Cre 的体内选择平台 (CREATE) 鉴定可有效转导基因定义的腺相关病毒 (AAV) 人口。我们使用 CREATE 来选择转导大脑的病毒 血管内递送并发现了一种非特异性转导大多数细胞的载体 成人的大脑。由于血脑屏障的限制性是一个主要的限制因素 治疗中枢神经系统疾病的障碍我们的发现有可能使 通过 CRISPR-Cas 或 RNA 干扰进行基因编辑/替换方面取得了令人兴奋的进展 如果可以达到所需的效率和特异性水平，则可以恢复患病的中枢神经系统回路 专为患病目标而设计。 我们计划通过创建基于病毒的非侵入性整体解决方案来实现这些努力 大脑大货物通过血脑屏障从胚胎到成人的运输通过： 1. 生成 AAV，用于跨血脑的细胞类型和区域特异性基因传递 屏障，通过成年啮齿动物的血流无创治疗神经退行性变 应用程序。 2. 生成能够通过简单的方法在子宫内转导发育中的大脑的 AAV 对怀孕母鼠进行全身注射，用于神经发育研究和治疗。 3. 将AAV的包装能力提高约2倍，以实现大批量交付 用于基因治疗和研究的基因组。 4. 通过使用系统性的方法实现非侵入性电路特定的深部脑调制 载体和基因编码的活性调节剂（例如通过化学遗传学或其他方法） 现在正在开发）。 从长远来看，我们计划在我们的实验室以及合作者的帮助下，贡献我们的力量 神经技术（除了病毒载体之外，还包括组织清除和光遗传学） 控制和成像）以阐明导致适应不良的神经回路 神经变性和神经发育中的脑病理学。