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，基于通过 CAR-T 技术对 T 细胞进行基因重编程的细胞疗法的潜力 已针对心脏疾病进行了探索，该策略所体现的希望已受到广泛欢迎 和科学想象力。然而，在目前的临床CAR-T细胞治疗实践中，体外方法必须 的应用，这是限制该技术更广泛实施的主要时间和成本障碍。 因此，在患者体内原位生成 CAR-T 的稳健方法将极大地促进实际应用。 应用这种有前途的方法。为此，我们建议开发一个强大的基因传递系统，包括 联合“SAd.AAV”纳米载体与多个 T 细胞嗜性腺相关病毒 (AAV) 缀合 其衣壳表面有一种经临床批准的 T 细胞热带猿猴（黑猩猩）腺病毒 36 (SAd)。值得注意的是， 衣壳工程化的 SAd 和 AAV 都能够选择性地靶向多种组织，包括具有高活性的 T 细胞。 转导效率，因此我们假设 SAd.AAV 可能通过以下方式提供卓越的靶向性： 两种工程病毒衣壳针对同一组织或细胞上不同受体的综合效应 类型。重要的是，携带单链敲入供体 DNA 模板的 AAV 已被广泛证明 支持同源定向修复（HDR）的高编辑效率。此外，稳健但短暂的 SAd 表达基因编辑器还提供了非常理想的“打了就跑”的基因编辑，减少了潜在的可能性 实现目标编辑后，与延长编辑器表达相关的不利影响。所以， 凭借每种病毒在基因编辑方面的优势，SAd.AAV 潜在地增加了组织靶向性 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，在构建并表征第一个 T 细胞靶向 SAd.AAV 后，我们计划实现高效 利用 SAd.AAV 基因编辑平台进行体内敲入，生产心脏病特异性 CAR-T 细胞 体内T细胞中可切换的CARi转基因。体内生产的原理验证演示 基因编辑的可切换CARi-T细胞将为CARi-T干预措施的后续研究建立一个关键平台 心脏病的小鼠模型。我们还预计 SAd.AAV 可以快速重新设计以 针对与 NHLBI 相关的多种临床重要器官进行基于基因编辑的治疗。