Testing Optimal Gene Editor for an Alzheimer's CRISPR therapeutic.

测试阿尔茨海默病 CRISPR 疗法的最佳基因编辑器。

• 批准号: 10746716
• 负责人: Subhojit Roy
• 金额: $ 211.28万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2026-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10746716
• 关键词: Advisory Committees; Alzheimer' s Disease; Alzheimer' s disease therapeutic; Alzheimer' s disease therapy; Amyloid; Amyloid beta-Protein; Amyloid beta-Protein Precursor; Attenuated; Biological Assay; Brain; CRISPR therapeutics; CRISPR/Cas technology; Cell surface; Cells; Clinic; Clinical; Clinical Paths; Clinical Trials; Clustered Regularly Interspaced Short Palindromic Repeats; Consensus; Consultations; Deposition; Disease; Dose; Down Syndrome; Early Onset Alzheimer Disease; Endosomes; Equilibrium; Etiology; Event; Exons; Eye diseases; FDA approved; Failure; Future; Gene Proteins; Genes; Genetic; Genome; Goals; Guide RNA; Hand; Human; Human Amyloid Precursor Protein; Human Genetics; Injections; Kinetics; Knock-in; Knock-in Mouse; Lead; Letters; Measures; Mediating; Mission; Modality; Mouse Protein; Mus; Nerve Regeneration; Neurofibrillary Tangles; Neurons; Organoids; Outcome; Pathologic; Pathway interactions; Persons; Pharmaceutical Preparations; Phase I/II Trial; Physiological; Physiology; Positioning Attribute; Protein C; Protein Fragment; Protein Truncation; Proteins; Proteolysis; Public Health; Quantitative Reverse Transcriptase PCR; RNA; Recommendation; Research; Research Personnel; Retina; Role; Safety; Surface; Testing; Therapeutic; Time; Transcript; Transmembrane Domain; United States National Institutes of Health; Variant; adeno-associated viral vector; base editing; beta-site APP cleaving enzyme 1; cost; efficacy testing; experimental study; extracellular; genome-wide; in vivo; induced pluripotent stem cell; meetings; mouse model; neuroprotection; nonhuman primate; novel; novel therapeutics; prevent; response; secretase; tau Proteins; tau aggregation; therapeutic genome editing; translation to humans

Summary/Abstract: The overall goal of this U01 proposal is to advance a unique CRISPR-based therapeutic for Alzheimer’s disease (AD) towards the pre-IND stage, by determining the best genome-editor/gRNA combination that can be ultimately used in clinical trials. Though there is an enormous unmet need for developing therapeutics for AD, we do not have drugs that can unequivocally slow down the relentless course of AD. We have developed a gene-editing based strategy targeting APP – a gene with a central and indisputable role in AD; strongly supported by human genetics. We do not eliminate the APP gene, but edit out a pentapeptide YENPTY endocytic domain at the C- terminus of APP, which blocks the entire pathologic APP β-cleavage pathway (including β-amyloid). The transmembrane domain and the N-terminus remains intact, and the ∆C APP is retained on the cell surface, leading to an increase in APP α-cleavage, which in turn upregulates neuroprotective and neuroregenerative APP fragments. Thus, our strategy shifts the balance of APP cleavage from pathologic to physiologic, without eliminating the gene. Since the targeted events are upstream, our strategy should be applicable to forms of AD – sporadic and familial – and also to early-onset AD in Down syndrome, where APP triplication on Ch.21 invariably causes AD. So far, we have used the classical SaCas9 to demonstrate safety and efficacy of our approach in vivo, but SaCas9 is too big to fit into a single AAV, which will be necessary for ultimate delivery in humans. Moreover, since there is no FDA-approved gene-editing clinical trial in the brain yet, and the best genome-editor is unknown. Here, we propose to first test two promising small genome-editors (and corresponding gRNAs) that can fit into a single AAV – SaCas9 and NmCas9 – using genome-scale on- and off- target assays (Aim 1). The best Lead-Editor/gRNA combinations emerging from these experiments will be tested for efficacy, safety, predictability, and durability in human brain organoids and a novel APP mouse model where the entire mouse APP was replaced with human APP (Aim 2). Collectively, these studies will not only clarify the best genome-editor/RNA combination for editing the APP gene, but also help future trials in the brain gene- editing space by determining the best editor for brain-relevant applications. Towards the end of our projects, we propose to hold an INTERACT meeting with the FDA – in consultation with our Clinical/Translational Advisory Team – to get regulatory input and consensus on pre-IND NHP studies. Upon completion, our studies will not only offer a novel therapeutic in an arena of enormous unmet need and litany of failures, but also provide a clinical path for future gene-editing efforts in this therapeutic space.

总结/摘要： U 01提案的总体目标是推进一种独特的基于CRISPR的阿尔茨海默病治疗方法 (AD)通过确定最终可以实现的最佳基因组编辑器/gRNA组合， 用于临床试验。尽管开发AD治疗方法的需求巨大，但我们没有 有药物可以明确地减缓AD的无情进程。我们开发了一种基因编辑技术 基于靶向APP的策略-一种在AD中具有核心和无可争议的作用的基因;得到人类的强烈支持 遗传学我们没有消除APP基因，但编辑了一个五肽YENPTY内吞结构域在C- APP末端，阻断整个病理性APP β-裂解途径（包括β-淀粉样蛋白）。的 跨膜结构域和N-末端保持完整，并且CD 4C APP保留在细胞表面上， 导致APP α裂解增加，进而上调神经保护和神经再生APP 片段因此，我们的策略将APP切割的平衡从病理性转移到生理性， 消除基因。由于目标事件是上游事件，我们的策略应适用于各种形式的广告 - 散发性和家族性-以及唐氏综合征中的早发性AD，其中第21章APP三倍 总是导致AD。到目前为止，我们已经使用经典的SaCas 9来证明我们的药物的安全性和有效性。 虽然SaCas 9在体内的方法是可行的，但SaCas 9太大而不能装入单个AAV，这对于最终递送到体内是必要的。 人类此外，由于目前还没有FDA批准的大脑基因编辑临床试验， 基因编辑器是未知的。在这里，我们建议首先测试两个有前途的小基因组编辑器（和 相应的gRNA），可以使用基因组规模的开启和关闭来适应单个腺相关病毒（SaCas 9和NmCas 9） 靶向测定（Aim 1）。将测试从这些实验中出现的最佳Lead-Editor/gRNA组合 在人脑类器官中的有效性、安全性、可预测性和耐久性，以及一种新型APP小鼠模型， 用人APP替换整个小鼠APP（Aim 2）。总的来说，这些研究不仅将澄清 最好的基因组编辑器/RNA组合编辑APP基因，但也有助于未来的大脑基因试验- 通过确定大脑相关应用程序的最佳编辑器来编辑空间。在项目结束时，我们 建议与FDA举行互动会议-与我们的临床/翻译顾问协商 团队-就IND前NHP研究获得监管意见和共识。完成后，我们的研究将不会 不仅在巨大的未满足需求和一连串失败的竞技场中提供了一种新的治疗方法，而且还提供了一种 为未来在这一治疗领域的基因编辑工作提供临床路径。