Brain-Wide Genome Editing Enabled by Intravenously Administered Non-Viral Nanovectors As a Potential Therapy for Alzheimer’s Disease

静脉注射非病毒纳米载体实现全脑基因组编辑作为阿尔茨海默病的潜在疗法

• 批准号: 10630541
• 负责人: SHAOQIN GONG
• 金额: $ 186.03万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-05-15 至 2026-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10630541
• 关键词: Affect; Alzheimer' s Disease; Alzheimer' s disease therapy; Amyloid beta-Protein Precursor; Biodistribution; Biological Assay; Biological Products; Blood - brain barrier anatomy; Brain; Brain region; Bypass; CRISPR/Cas technology; Chemistry; Clustered Regularly Interspaced Short Palindromic Repeats; Collaborations; DNA; Data; Disease Progression; Elderly; Encapsulated; Engineering; Gene Targeting; Genes; Genetic; Genetic Engineering; Genome; Glucose; Glutathione; Guide RNA; Hippocampus; Human; Intravenous; Knock-in; Ligands; Messenger RNA; Monitor; Mus; Neurodegenerative Disorders; Neurons; Non-Viral Vector; Nucleic Acids; Organ; Particle Size; Pathologic; Peptides; Pharmacologic Substance; Population; Preclinical Testing; Production; Property; Proteins; Resolution; Ribonucleoproteins; Safety; Silicon Dioxide; Stimulus; Surface; Synapsins; System; Techniques; Therapeutic; Therapeutic Studies; Treatment Efficacy; United States National Institutes of Health; Viral Vector; Wild Type Mouse; amyloid imaging; behavior change; behavior test; biomaterial compatibility; blood-brain barrier crossing; brain cell; cell type; delivery vehicle; dosage; familial Alzheimer disease; gene therapy; genome editing; imaging platform; immunogenicity; in vivo; innovation; intravenous administration; mouse model; nanocapsule; nanovector; novel; plasmid DNA; preclinical evaluation; prevent; programs; promoter; rabies virus glycoprotein G; screening; systemic toxicity; therapeutically effective; therapy outcome; tomography; two-photon

Project Summary Currently, there is no effective way to slow down the progress of Alzheimer’s disease (AD) or prevent it. CRISPR genome editing is a revolutionary and versatile genetic engineering technique, making it possible to treat the root causes of genetic neurodegenerative diseases (NDDs) such as AD. However, the promise of brain gene therapy relies on the efficient delivery of biologics to the brain, which is extremely challenging due to the blood- brain barrier (BBB). To date, in vivo brain gene therapy has mostly been achieved using viral vectors that require laborious customization and have troublesome safety profiles. Non-viral vectors are largely administered via intracranial administration, which is invasive and can only enable gene therapy in a small and localized brain region. Similar to other NDDs, AD affects multiple brain regions. Thus, there is an urgent need to develop efficient non-viral delivery vehicles capable of bypassing the BBB for safe and efficient brain-wide gene therapy. The objectives of this project are (1) to engineer glutathione (GSH)-responsive silica nanocapsules (SNCs) capable of bypassing the BBB and delivering CRISPR genome editors to the whole brain systemically, and (2) to evaluate the therapeutic efficacy and biosafety of brain-wide genome editing enabled by the optimized SNC for the treatment of AD using a novel amyloid precursor protein (APP) knock-in AD mouse model and a unique gene target for APP modulation. The unique SNC possess a long list of desirable properties including versatile payload types, versatile surface chemistry for ligand conjugation, high payload loading content and efficiency, small particle sizes, excellent in vivo stability, good biocompatibility, and scalable production. Our preliminary data has shown that intravenously administered SNCs can efficiently deliver mRNA, DNA, Cas9 mRNA/sgRNA, and Cas9/gRNA ribonucleoprotein (RNP) to the whole brain of healthy mice with intact BBB. We aim to further optimize the amounts of the dual brain-targeting ligands (i.e., glucose and rabies virus glycoprotein (RVG) peptide) and dosages of the SNCs for enhanced brain-wide systemic delivery of two types of CRISPR genome editors (i.e., (1) Cas9 mRNA/sgRNA, and (2) plasmid DNA with a neuron-specific human synapsin 1 (SYN1) promoter and expressing both Cas9 and sgRNA). We will further determine their therapeutic efficacy and biosafety in treating AD using a novel APP knock-in AD mouse model while employing a unique gene target for APP modulation. The gene editing efficiency and biosafety profiles of the SNC, and the pathological and behavior changes of the AD mice will be monitored by various techniques including a novel serial two-photon tomography whole-brain amyloid imaging platform. With promising initial studies, the best- performing SNC will be submitted to the Preclinical Testing Core of the NIA-sponsored STOP-AD program for comprehensive preclinical evaluation. This project will pave the road for a new, safe, non-invasive and effective therapeutic approach for familial AD. Given the modularity and versatility of the SNCs, and ease of targeting different genes by the CRISPR system, we anticipate that our SNCs will be applicable for a wide range of NDDs.

项目概要 目前，尚无有效的方法可以减缓或预防阿尔茨海默病（AD）的进展。 基因组编辑是一种革命性的、多功能的基因工程技术，使治疗 AD 等遗传性神经退行性疾病 (NDD) 的根本原因是大脑基因的希望。 治疗依赖于将生物制剂有效地输送到大脑，由于血液 迄今为止，体内脑基因治疗主要是使用需要的病毒载体来实现的。 非病毒载体主要通过繁琐的定制和安全性问题进行管理。 颅内给药是侵入性的，只能在较小的局部大脑中进行基因治疗 与其他 NDD 类似，AD 影响多个大脑区域，因此迫切需要开发有效的方法。 能够绕过血脑屏障进行安全有效的全脑基因治疗的非病毒递送载体。 该项目的目标是 (1) 设计谷胱甘肽 (GSH) 响应型二氧化硅纳米胶囊 (SNC) 能够绕过 BBB 并将 CRISPR 基因组编辑系统系统地传递到整个大脑，以及 (2) 评估优化的 SNC 实现的全脑基因组编辑的治疗效果和生物安全性 使用新型淀粉样前体蛋白 (APP) 敲入 AD 小鼠模型和独特的 独特的 SNC 具有一系列理想的特性，包括多功能性。 有效负载类型、用于配体缀合的多功能表面化学、高有效负载负载含量和效率， 粒径小，体内稳定性优异，生物相容性好，可规模化生产。 数据显示，静脉注射 SNC 可以有效递送 mRNA、DNA、Cas9 mRNA/sgRNA、 Cas9/gRNA 核糖核蛋白 (RNP) 作用于具有完整 BBB 的健康小鼠的整个大脑。 我们的目标是进一步优化双脑靶向配体（即葡萄糖和狂犬病病毒）的数量 糖蛋白（RVG）肽）和 SNC 的剂量，用于增强两种类型的全脑系统递送 CRISPR 基因组编辑器（即 (1) Cas9 mRNA/sgRNA，和 (2) 具有神经元特异性人类的质粒 DNA 突触蛋白 1 (SYN1) 启动子并表达 Cas9 和 sgRNA）我们将进一步确定其治疗作用。 使用新型 APP 敲入 AD 小鼠模型治疗 AD 的功效和生物安全性，同时采用独特的 APP 调制的基因靶标。SNC 的基因编辑效率和生物安全性概况。 AD小鼠的病理和行为变化将通过各种技术进行监测，包括一种新颖的技术 串行双光子断层扫描全脑淀粉样蛋白成像平台具有前景良好的初步研究，最好的- 执行 SNC 将提交给 NIA 赞助的 STOP-AD 计划的临床前测试核心 该项目将为新的、安全的、非侵入性的和有效的方法铺平道路。 鉴于 SNC 的模块化和多功能性以及易于靶向性，家族性 AD 的治疗方法。 通过 CRISPR 系统识别不同的基因，我们预计我们的 SNC 将适用于广泛的 NDD。