Improving CAR T cell therapy for brain tumors using immune competent models

使用免疫活性模型改善脑肿瘤的 CAR T 细胞疗法

• 批准号: 10732795
• 负责人: Dalia Haydar
• 金额: $ 24.9万
• 依托单位: CHILDREN'S RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-01-01 至 2025-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10732795
• 关键词: Adoptive Transfer; Adult; Adverse effects; American; Antigen Targeting; Antigens; Biological; Brain Neoplasms; CAR T cell therapy; CBLB gene; CD276 gene; CD28 gene; CRISPR screen; CRISPR/Cas technology; Cell Cycle; Cell surface; Cells; Cellular immunotherapy; Chemotherapy and/or radiation; Childhood; Childhood Brain Neoplasm; Clinical; Clinical Research; Core Facility; Disease; Down-Regulation; Engineering; Generations; Genes; Genetic; Glioma; Goals; Hematologic Neoplasms; Heterogeneity; Homing; IL3RA gene; Image; Immune; Immune response; Immunocompetent; Immunology; Immunotherapy; Impaired cognition; Impairment; In Vitro; Inflammatory; Intervention; Life; Macrophage; Mediating; Mediator; Memory; Modeling; Modification; Molecular; Mus; Mutate; Operative Surgical Procedures; Outcome; Patients; Penetration; Performance; Persons; Phase; Phenotype; Play; Positioning Attribute; Pre-Clinical Model; Primary Brain Neoplasms; Proliferating; Receptor Signaling; Research; Research Personnel; Resistance; Role; Safety; Saint Jude Children' s Research Hospital; Shapes; Subgroup; T-Cell Activation; T-Cell Homing Receptors; T-Cell Receptor; T-Lymphocyte; TCR Activation; Techniques; Testing; Training; Transmembrane Domain; Tumor Antigens; Tumor-associated macrophages; Xenograft Model; antigen-specific T cells; antitumor effect; career; chimeric antigen receptor; chimeric antigen receptor T cells; cytokine; cytotoxicity; design; engineered T cells; exhaustion; extracellular; genome wide screen; human disease; immune cell infiltrate; improved; improved outcome; in vitro activity; in vivo; innovative technologies; interest; motor impairment; mouse model; neoplastic cell; novel; prototype; receptor; research clinical testing; success; targeted treatment; tool; trafficking; transcriptomics; tumor; tumor microenvironment; tumor xenograft; tumor-immune system interactions

PROJECT SUMMARY/ABSTRACT The overall objective of this study is to engineer a potent CAR T cell therapy for high grade glioma (HGG), a subgroup of brain tumors for which outcomes remain poor. CAR T cell therapy is an innovative technology based on adoptive transfer of antigen-specific T cells engineered to elicit a clinically effective and specific immune response against tumor cells. Early clinical studies in HGG patients demonstrated safety of CAR T cell therapy for brain tumors; yet, only limited benefits were observed. Lack of efficacy is most likely multifactorial and include heterogenous antigen expression, limited homing to and penetration of tumors, T cell exhaustion and limited persistence, as well as the immunosuppressive tumor microenvironment (TME). Therefore, the central hypothesis of this proposal is that optimized design and additional genetic modifications of CAR T cells will improve their antitumor effects, and that these interventions will be evaluated in immune competent mouse models that closely mimic human disease. Three interrelated research aims are proposed to test this hypothesis and the rationale of each is outlined below. First, CAR design has to be optimized and evaluated in immune competent glioma models. In Aim 1, I will generate CAR T cells containing different co- stimulatory domains (CD28.ζ, 41BB.ζ, CD28.mutζ, and 41BB.mutζ), and compare their activity in vitro and in vivo. Second, a genome wide screen in primary T cells has identified key regulators of T cell activation post T- cell receptor (TCR) stimulation. Identified genes belong to molecules that regulate cell cycle, proliferation, and downstream TCR signaling including SOCS1, RASA2, or CBLB. In Aim 2, I will therefore explore if CRISPR- Cas9 mediated silencing of Socs1, Rasa2, and/or Cblb enhances the effector functions of CAR T cells. The independent Aim 3 will then explore a dual CAR targeting approach in which I will not only target glioma cells but also immunosuppressive cells within the glioma microenvironment. These studies are focused on tumor associated macrophages (TAMs), since they are abundantly present in gliomas and play a critical role in shaping the TME. To support the feasibility of this project, I have adapted the well-established immune competent GL261 glioma model to study CAR T cell therapies targeting the relevant tumor antigen B7-H3, which is not only expressed by GL261 cells but also in a broad range of pediatric and adult brain tumors. In addition, my preliminary studies indicate that `prototype' B7- H3 CAR T-cells readily recognize and kill GL261 cells in vitro and have antitumor activity in vivo, highlighting that the developed model is well suited for the proposed aims of this project. State of the art technique will be used in all three Aims to not only study the function and in vivo fate of CAR T-cells, but also their antitumor activity, and how CAR T-cells interact with glioma- infiltrating immune cells. Completion of this study will define the most optimal CAR design that best controls HGG tumors and persists longer in the context of inflammatory brain tumors. Additionally, results will illustrate if targeting TAMs will overcome the suppressive effects of TME on B7-H3 CAR T cells.

项目摘要/摘要 这项研究的总体目标是设计一种有效的CAR T细胞治疗高级别胶质瘤(HGG)， 结果仍然很差的脑肿瘤的一个亚群。汽车T细胞疗法是一项创新技术 基于过继转移抗原特异性T细胞的基因工程诱导临床有效和特异 针对肿瘤细胞的免疫反应。HGG患者的早期临床研究证明CAR T细胞是安全的 脑瘤的治疗；然而，只观察到有限的好处。疗效的缺乏很可能是多因素的 包括异质性抗原表达，肿瘤的有限归巢和穿透，T细胞耗竭 和有限的持久性，以及免疫抑制肿瘤微环境(TME)。因此， 这一提议的中心假设是，汽车的优化设计和额外的遗传修改 T细胞将提高其抗肿瘤效果，这些干预措施将在免疫学中进行评估 近乎模拟人类疾病的合格小鼠模型。提出了三个相互关联的研究目标 下面概述了检验这一假说和每种假说的基本原理。首先，必须优化汽车设计，并 在免疫活性胶质瘤模型中进行评估。在目标1中，我将生成包含不同co-1的CAR T细胞 刺激结构域(CD28.ζ、41Bb.ζ、CD28.mutζ和41BB.mutζ)，并比较它们在体外和体内的活性 活着。其次，在初级T细胞中的全基因组筛查已经确定了T细胞在T细胞死亡后激活的关键调节因子。 细胞受体(TCR)刺激。已识别的基因属于调节细胞周期、增殖和 下行TCR信令包括SOCS1、RASA2或CBLB。因此，在目标2中，我将探讨CRISPR是否- Cas9介导的Socs1、Rasa2和/或Cblb沉默增强了CAR T细胞的效应功能。这个 然后，《独立目标3》将探索一种双重CAR靶向方法，在这种方法中，我不仅将靶向胶质瘤细胞 还包括胶质瘤微环境中的免疫抑制细胞。这些研究都集中在肿瘤上。 相关巨噬细胞(TAM)，因为它们在胶质瘤中大量存在，并在形成过程中发挥关键作用 TME。为了支持这个项目的可行性，我对成熟的免疫功能GL261进行了改造 胶质瘤模型研究针对相关肿瘤抗原B7-H3的CAR T细胞治疗，这不仅是 不仅在GL261细胞中表达，而且在儿童和成人脑肿瘤中也有表达。此外，我的 初步研究表明“原型”B7-H3CAR T细胞在体外很容易识别和杀伤GL261细胞 并在体内具有抗肿瘤活性，突出表明所开发的模型非常适合所提出的目标 这个项目。最先进的技术将用于所有这三个目标，不仅是为了研究功能和体内命运 CAR T细胞及其抗肿瘤活性，以及CAR T细胞如何与胶质瘤浸润性免疫相互作用 细胞。这项研究的完成将确定最优的汽车设计，最好地控制HGG肿瘤和 在炎症性脑瘤的情况下持续时间更长。此外，结果将说明是否以TAMs为目标 将克服TME对B7-H3CAR T细胞的抑制作用。