Bioengineering of highly effective AAV vectors for noninvasive gene delivery to the nervous system

高效 AAV 载体的生物工程，用于将基因非侵入性传递至神经系统

• 批准号: 10597682
• 负责人: SHUXIN LI
• 金额: $ 39.63万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-01 至 2027-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10597682
• 关键词: Adult; Age; Alzheimer' s Disease; Astrocytes; Axon; Biomedical Engineering; Blood - brain barrier anatomy; Capsid; Capsid Proteins; Cd68; Cells; Central Nervous System Diseases; Chimeric Proteins; Chondroitin Sulfate Proteoglycan; Cicatrix; Clinical; Effectiveness; Engineering; Evaluation; Failure; Gene Delivery; Gene Targeting; Genes; Glial Fibrillary Acidic Protein; Growth; Growth Inhibitors; Injections; Intravenous; Invaded; Locomotor Recovery; Mammals; Mediating; MicroRNAs; Microglia; Molecular Target; Motor; Mouse Strains; Mus; Mutagenesis; Natural regeneration; Nervous System; Nervous System Trauma; Neurons; Neurosciences; Neurosciences Research; Oligodendroglia; PTEN gene; Pathway interactions; Penetration; Peptides; Pericytes; Population; Probability; Protein Tyrosine Phosphatase; Recovery; Recovery of Function; Research Personnel; Rodent; SOX11 gene; Signal Transduction; Spinal cord injury; Synapsins; Technology; Transduction Gene; Transgenic Mice; Translating; Tropism; Up-Regulation; Viral Vector; Virus; Wild Type Mouse; adeno-associated viral vector; age related; axon growth; axon regeneration; blood-brain barrier crossing; c-myc Genes; cellular transduction; central nervous system injury; design; effective therapy; improved; in vivo; inhibitor; intravenous injection; nervous system disorder; neural; neuronal growth; novel; promoter; receptor; receptor function; regenerative; regenerative approach; regenerative therapy; repaired; technology validation; tool; transduction efficiency; transgene expression; vector

Abstract: This project is designed to identify a noninvasive and highly effective gene delivery strategy to target specific neural cells in the CNS and to validate this technology by delivering regenerative molecules to mammals with CNS injury. We will determine whether systemic delivery of our newly engineered AAV9 vectors can transduce most target CNS cells and whether noninvasive delivery of the genes that target neuronal intrinsic and extrinsic factors can promote robust axon regeneration and functional recovery in rodents with spinal cord injury (SCI). A major challenge in neuroscience research is to deliver target genes to specific types of neural cells widely distributed in CNS. Engineered AAV9 vectors usually show limited efficacy after intravenous (IV) injection by transducing cells only in some CNS regions. We thus created new AAV9 vectors that include multiple features of engineered AAV9 capsids, aiming to develop highly efficient BBB-crossing AAV9 vectors (HEBC-AAV9) that can transduce most target CNS cells after IV injection. In Aim 1, we will study efficiency of our novel HEBC-AAV9-GFP vectors for selectively transducing each type of neural cells (neurons, astrocytes, oligodendrocytes, and microglia) in several strains of adult mice. To solve a crucial issue in neuroscience research with this technology, in Aim 2 we will develop a regenerative therapy for SCI by systemic delivery of genes to target neuronal let-7 miRNA. After SCI, severed axons fail to regenerate partly because of reduced intrinsic growth capacity of mature neurons. Many genes are known to control the growth ability of mature neurons, but none have been translated to clinical use. The best targets are probably those with potential to impact multiple genes. Among them, let-7 is important for regulating age-dependent decline in axon regeneration in worms. In Aim 2, we propose to use unique HEBC-AAV9-synapsin vectors to target neurons selectively for inducing expression of let-7 inhibitor, lin28, and lin41, aiming to promote robust regeneration of multiple axon tracts by enhancing growth capacity of mature neurons in SCI rodents. Chondroitin sulfate proteoglycans (CSPGs) generated by glial scars strongly suppress axon extension and are major extrinsic molecular targets for treating CNS injury. Our lab designed small peptides to block functions of CSPG receptor LAR, PTPσ, and PTPδ by targeting their critical activity domains and demonstrated their high efficiency for promoting axon regrowth. In Aim 3, we will induce astrocytic expression of secreted 3 peptides for each of LAR, PTPσ, and PTPδ with HEBC- AAV9-GFAP vectors, aiming to promote robust axon regeneration after SCI by targeting extrinsic CSPGs alone or combined with intrinsic let-7 signals. Our new viral vectors should provide a powerful tool for gene delivery in CNS and for developing effective regenerative therapies for SCI and other neurological disorders.

抽象的： 该项目旨在确定一种无创和高效的基因输送策略来靶向 中枢神经系统中的特定神经细胞，并通过将再生分子传递到 中枢神经系统损伤的哺乳动物。我们将确定我们新设计的AAV9的系统交付是否 向量可以翻译大多数目标中枢神经系统细胞，以及靶向基因的无创递送 神经元内和外在因素可以促进稳健的轴突再生和功能恢复 在脊髓损伤（SCI）的啮齿动物中。神经科学研究的主要挑战是实现目标 特定类型的神经细胞的基因广泛分布在中枢神经系统中。通常设计的AAV9向量 仅在某些中枢神经系统区域中通过转导细胞来显示静脉内（IV）注射后的有限效率。 因此，我们创建了新的AAV9向量，其中包含工程AAV9 CAPSIDS的多个功能，瞄准 开发高效的BBB跨AAV9矢量（HEBC-AAV9），可以转化大多数目标 注射后CNS细胞。在AIM 1中，我们将研究新型HEBC-AAV9-GFP载体的效率 用于选择性转导每种类型的神经元（神经元，星形胶质细胞，少突胶质细胞和 小鼠中的小胶质细胞）。解决神经科学研究的关键问题 技术，在AIM 2中，我们将通过全身传递基因来开发SCI的再生疗法 靶神经元let-7 miRNA。 SCI之后，严重的轴突无法部分再生，部分原因是 成熟神经元的内在生长能力。已知许多基因可以控制 成熟的神经元，但没有人转化为临床用途。最好的目标可能是 可能会影响多个基因。其中，Let-7对于调节年龄依赖性很重要 蠕虫中轴突再生的下降。在AIM 2中，我们建议使用独特的HEBC-AAV9-Synapsin 靶向神经元的向量选择性地诱导Let-7抑制剂，Lin28和Lin41的表达 通过增强成熟神经元的生长能力来促进多个轴突的稳健再生 在Sci啮齿动物中。神经胶质疤痕产生的硫酸软骨蛋白蛋白聚糖（CSPG）强烈抑制 轴突延伸是治疗中枢神经系统损伤的主要外部分子靶标。我们的实验室设计 小肽通过靶向临界 活动域并证明了它们在促进轴突后悔的高效率。在AIM 3中，我们将 用HEBC-诱导每种LAR，PTPσ和PTPδ的分泌3肽的星形细胞表达 AAV9-GFAP矢量，旨在通过靶向外部的SCI后促进稳健的轴突再生 单独使用CSPG或与固有的Let-7信号结合使用。我们的新病毒载体应提供强大的 中枢神经系统中基因递送的工具，并开发SCI和其他有效的再生疗法 神经系统疾病。