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: 10251895
• 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 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10251895
• 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.

Viviana Gradinaru，加州理工学院 随着CRISPR/CAS9等技术的出现，两者的基因组工程 基础研究和治疗应用已成为现实。一个出色的挑战 是均值安全有效地将大基因组传递到所需细胞的均值 跨度。我们已经开发了一个基于体内CRE的选择平台（创建） 鉴定有效转导遗传定义的腺相关病毒（AAVS） 人群。我们使用创建来选择在转导大脑的病毒 血管内递送，发现一个载体，该载体非特异性地将大多数细胞跨越 成人大脑。由于血脑屏障的限制性是主要的 障碍治疗中枢神经系统疾病我们的发现有可能实现 通过CRISPR-CAS或RNA干扰对基因编辑/替换的令人兴奋的进步 恢复患病的CNS电路，如果需要的效率和特异性水平是 为患病靶标而设计。 我们计划通过创建基于病毒的解决方案来实现此类努力 大脑在从胚胎到成人的血脑屏障上的大型货物传递： 1。生成用于细胞类型和区域特异性基因递送的AAV 屏障，通过成年啮齿动物的血液无创的神经变性 申请。 2。生成能够用简单的 全身注射对神经发育研究和治疗的怀孕大坝。 3。将AAV的包装能力提高约2倍以实现大型 基因治疗和研究的基因组。 4。通过使用全身性启用非侵入电路特定的深脑调制 向量和遗传编码的活性调节剂（例如，通过化学遗传学或其他 现在开发）。 长期我们在实验室和合作者中计划，以贡献我们的 神经技术（包括病毒载体，组织清除和光遗传学 控制和成像）朝着阐明不良适应性神经回路有助于 神经退行性和神经发育中的脑病理学。