Epigenetic modulation of cellular aging to improve CAR-T therapy for solid tumors

表观遗传调节细胞衰老以改善实体瘤的 CAR-T 疗法

• 批准号: 10326136
• 负责人: Benedetta Nicolis di Robilant
• 金额: $ 33.56万
• 依托单位: DORIAN THERAPEUTICS, INC.
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-23 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10326136
• 关键词: 3-Dimensional; Affect; American; Animal Model; Antigens; CAR T cell therapy; CD19 gene; Cell Aging; Cell Line; Cell physiology; Cells; Chromatin; Clinical; Coculture Techniques; DNA cassette; Deubiquitinating Enzyme; Doxorubicin; Engineering; Epigenetic Process; Excision; Frequencies; Genetic; Glioblastoma; Goals; Grant; Hematologic Neoplasms; Histones; Immune; Immunosuppression; In Vitro; Injections; Lead; Luciferases; Malignant Neoplasms; Mediating; Memory; Modeling; Monitor; Myeloid-derived suppressor cells; Neuroblastoma; Outcome; Patients; Phenotype; Resistance; Small Business Innovation Research Grant; Solid Neoplasm; Stimulus; System; T-Lymphocyte; Testing; Therapeutic; Tissues; Toxicology; Transforming Growth Factor beta; Tumor-Derived; Tumorigenicity; Ubiquitin; Universities; Xenograft procedure; base; cancer cell; cell age; cell stroma; chimeric antigen receptor T cells; cytokine release syndrome; early phase clinical trial; effector T cell; engineered T cells; exhaustion; fitness; improved; in vitro Model; in vivo; innovation; leukemia; mouse model; neoplastic cell; novel; novel strategies; osteosarcoma; performance tests; pre-clinical; programs; public health relevance; response; scaffold; screening; self-renewal; senescence; small hairpin RNA; stem cell self renewal; stem cells; success; tool; transcriptome sequencing; tumor; tumor microenvironment

Abstract CAR-T cell therapy has had incredible clinical success in the treatment of hematological malignancies. However, very limited activity against solid tumors has been achieved so far, despite targeting a variety of antigens and tumor types. Importantly, solid tumors account for more than 90% of all cancers, affecting more than 16 million Americans in 2020. The goal of Dorian Therapeutic’s SBIR proposal is to investigate the impact of reducing cellular senescence in CAR-T cells to improve solid tumor targeting. We have proven that T cells age the equivalent of 30 years during CAR-T manufacturing, which is detrimental to their ability to expand and kill cancer cells. Moreover, solid tumors are characterized by a strong immunosuppressive and pro-senescent microenvironment, strongly challenging T cells effector functions. Old T cells are particularly prone to be suppressed and die due to the hostile microenvironment. Rejuvenating T cells by reducing cellular aging at the epigenetic level is a novel strategy to improve T cell fitness and clinical outcomes. The goal of this project is to identify a shRNA-based genetic cassette targeting the epigenetic regulator USP16 to increase CAR-T cell fitness for the treatment of solid tumors. Dorian’s founders identified USP16 at Stanford University as an epigenetic regulator able to reduce senescence by enhancing stem cell self-renewal in multiple tissues. USP16 is a deubiquitinating enzyme responsible for the removal of ubiquitin moieties from histone H2AK119, increasing chromatin accessibility to pro-senescent programs. The company has developed in vitro and in vivo proof-of-concept that targeting USP16 by means of a shRNA co-expressed within the CD19.CAR construct reduces T cell aging and increases stem cell memory (Tscm) frequency upon manufacturing, without affecting proliferation. Most importantly, the engineered CAR-T cells resulted in improved tumor killing in a preclinical mouse model of leukemia. The Aims of this proposal are to (1) Create an in vitro model to study the effect of tumor microenvironment on T cells aging and functions, (2) Identify genetic cassettes targeting USP16 and increasing T cell fitness in vitro, and (3) Demonstrate that engineered CAR-T cells expressing a shRNA for USP16 have a better anti-tumor activity in a tumor model of osteosarcoma and in a 3D system of patient-derived tumor cells. At the successful completion of this SBIR project we will have identified the best genetic cassettes targeting USP16 and improving T cell fitness ready for toxicology studies (tumorigenicity and cytokine release syndrome (CRS) studies).

摘要 CAR-T细胞疗法在治疗血液恶性肿瘤方面取得了令人难以置信的临床成功。然而，在这方面， 尽管靶向多种抗原， 肿瘤类型重要的是，实体瘤占所有癌症的90%以上，影响超过1600万人 2020年的美国人Dorian Therapeutic的SBIR提案的目标是调查减少 CAR-T细胞中的细胞衰老以改善实体瘤靶向。 我们已经证明，在CAR-T制造过程中，T细胞老化相当于30年，这是有害的。 与它们扩张和杀死癌细胞的能力有关。此外，实体瘤的特征在于强的 免疫抑制和促衰老微环境，强烈挑战T细胞效应子功能。旧T 由于不利的微环境，细胞特别容易受到抑制并死亡。再生T细胞 通过在表观遗传水平上减少细胞衰老是一种改善T细胞适应性和临床 结果。该项目的目标是确定一个基于shRNA的基因盒，靶向表观遗传学， 调节子USP 16以增加CAR-T细胞适应性用于治疗实体瘤。 Dorian的创始人在斯坦福大学发现USP 16是一种能够减少衰老的表观遗传调节因子 通过增强多个组织中的干细胞自我更新。USP 16是一种去泛素化酶，负责 从组蛋白H2 AK 119中去除泛素部分，增加染色质对促衰老细胞的可及性， 程序.该公司已经开发了体外和体内概念验证，通过 在CD19.CAR构建体内共表达的shRNA减少T细胞老化并增加干细胞记忆 （Tscm）频率，而不影响增殖。最重要的是， 细胞在白血病的临床前小鼠模型中导致改善的肿瘤杀伤。 本研究的目的是：（1）建立一种体外模型，研究肿瘤微环境对T细胞增殖的影响， 细胞老化和功能，（2）鉴定靶向USP 16并在体外增加T细胞适应性的基因盒， 和（3）证明表达USP 16的shRNA的工程化CAR-T细胞具有更好的抗肿瘤活性， 在骨肉瘤的肿瘤模型和患者来源的肿瘤细胞的3D系统中的活性。 在成功完成SBIR项目后，我们将确定最佳的基因盒靶向 USP 16和改善T细胞适应性，可用于毒理学研究（致瘤性和细胞因子释放综合征 (CRS)研究）。