A Novel Vector Strategy to Feasibilize Cellular Therapy for Heart Disease

一种使心脏病细胞治疗变得可行的新型载体策略

• 批准号: 10746971
• 负责人: Zhi Hong Lu
• 金额: $ 19.44万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-01 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10746971
• 关键词: Address; Adenoviruses; Adverse effects; Biomedical Engineering; CAR T cell therapy; Capsid; Cell Therapy; Chemistry; Clinical; Clustered Regularly Interspaced Short Palindromic Repeats; Coupling; DNA; Dependovirus; Development; Disease; Engineering; Fiber; Gene Expression; Generations; Genes; Genetic; Genetic Transcription; Genome; Guide RNA; Heart; Heart Diseases; Human; Imagination; In Situ; In Vitro; Intervention; Knock-in; Link; Mediating; Methods; Mus; National Heart, Lung, and Blood Institute; Organ; Pan Genus; Patients; Production; Proteins; Reporting; Running; Safety; Serotyping; Simian Adenoviruses; Specificity; Sum; Surface; System; T-Lymphocyte; Techniques; Technology; Time; Tissues; Transgenes; Viral; Virus; base editing; cell type; chimeric antigen receptor T cells; clinical practice; coronary fibrosis; cost; delivery vehicle; design; gene delivery system; improved; in vivo; mouse model; nanovector; neoplastic; novel; practical application; pragmatic implementation; receptor; repaired; technology platform; technology validation; therapeutic genome editing; transduction efficiency; transgene expression; vector

ABSTRACT Recently, the potential of cellular therapy based upon genetic reprogramming of T cells via CAR-T technology has been explored for diseases of the heart, and the promise embodied in this strategy has captured popular and scientific imagination. However, in current clinical CAR-T cell therapy practice, extracorporeal methods must be applied, which represent a major time and cost barrier restricting wider implementation of this technology. Thus, robust methods to generate CAR-T in situ, within the patients, would greatly facilitate the practical application of this promising approach. For this, we propose to develop a robust gene delivery system consisting of a conjoined “SAd.AAV” nano-vector with multiple T cell-tropic adeno-associated viruses (AAVs) conjugated to a clinically approved, T-cell tropic simian (chimpanzee) adenovirus 36 (SAd) on its capsid's surface. Of note, both capsid-engineered SAd and AAVs are able to selectively target various tissues including T cells with high transduction efficiencies, and we hypothesized that SAd.AAV may therefore provide superior targeting through the combined effects of both engineered viral capsids targeting different receptors on the same tissue or cell type. Importantly, AAVs carrying single-stranded knock-in donor DNA templates have been widely shown to support high editing efficiency of homology directed repair (HDR). Furthermore, the robust but transient expression of gene editor by SAd also provides highly desirable “hit-and-run” gene editing reducing potential adverse effects associated with prolonged editor expression after the on-target editing is achieved. Therefore, by virtue of the advantages of each virus in gene editing, SAd.AAV potentially increases the tissue targeting specificity and efficiency and improves safety of existing CRISPR-Cas gene editing therapies. As a proof of concept, after constructing and characterizing the first T cell-targeted SAd.AAV, we plan to achieve efficient production of the heart disease-specific CAR-T cells in vivo with an SAd.AAV gene editing platform to knock-in a switchable CARi transgene in the T cells in vivo. A proof-of-principle demonstration of in vivo production of gene-edited switchable CARi-T cells will establish a key platform for follow-on studies of CARi-T interventions in murine models of heart diseases. We furthermore anticipate that the SAd.AAV can be rapidly redesigned to target a wide variety of clinically important organs relevant to the NHLBI for gene editing-based therapies.

抽象的 最近，基于T细胞的基因重编程的细胞疗法的潜力 已经探索了心脏疾病的探索，这种策略所体现的诺言捕获了流行 和科学的想象力。但是，在当前的临床CAR-T细胞治疗实践中，体外方法必须 应用，这代表了限制该技术更广泛实施的主要时间和成本障碍。 这是在患者内部生成car-t原位的强大方法，将极大地促进实际 这种有前途的方法的应用。为此，我们建议开发一个稳健的基因输送系统 与多个T细胞胶片相关病毒（AAVS）相连的“ Sad.aav”纳米矢量 经临床批准的T细胞热带Simian（Chimpanzee）腺病毒36（SAD）在其Capsid表面上。值得注意的是， 衣壳设计的SAD和AAV都能够选择性地靶向各种组织，包括高的T细胞 转导效率，我们假设Sad.aav因此可以通过 两种工程病毒式衣壳的综合作用，靶向不同的组织或细胞上不同受体 类型。重要的是，携带单链敲门供体DNA模板的AAV已被广泛显示为 支持同源性维修（HDR）的高编辑效率。此外，强大但短暂的 SAD的基因编辑器表达还提供了高度理想的“击球和跑”基因编辑，从而降低了潜力 实现靶向编辑后，与延长的编辑器表达相关的不良影响。所以， 由于每个病毒在基因编辑中的优势，sad.aav可能会增加组织靶向 特异性和效率，并提高现有CRISPR-CAS基因编辑疗法的安全性。作为证明 概念，在构建和表征第一个T细胞靶向的Sad.aav之后，我们计划实现高效 使用SAD.AAV基因编辑平台在体内生产心脏病特异性的CAR-T细胞 在体内T细胞中的可切换CARI转换。体内生产的原则证明 基因编辑的可切换CARI-T细胞将建立一个关键平台，用于跟随CARI-T干预措施 心脏病的鼠模型。我们此外，预计可悲的aav可以迅速重新设计为 针对与NHLBI相关的各种临床重要器官，用于基于基因编辑的疗法。