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

基本信息

批准号：
10322192
负责人：
Dalia Haydar
金额：
$ 12.5万
依托单位：
ST. JUDE CHILDREN'S RESEARCH HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-01-01 至 2022-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10322192
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 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 Mediating Mediator of activation protein 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 Radiation Receptor Signaling Research Research Personnel Resistance Role Safety Saint Jude Children&apos s Research Hospital Shapes Subgroup T-Cell Activation T-Cell Receptor T-Lymphocyte Techniques Testing Training Transmembrane Domain Tumor Antigens Tumor-associated macrophages Tumor-infiltrating immune cells Xenograft Model antigen-specific T cells antitumor effect base career chemotherapy chimeric antigen receptor chimeric antigen receptor T cells cytokine cytotoxicity design engineered T cells exhaustion extracellular genome wide screen human disease improved improved outcome in vitro activity in vivo innovative technologies interest macrophage 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

项目摘要

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.

项目概要/摘要这项研究的总体目标是设计一种有效的 CAR T 细胞疗法来治疗高级别胶质瘤 (HGG)， CAR T 细胞疗法是一种治疗效果不佳的脑肿瘤亚组，是一项创新技术。基于经过工程设计的抗原特异性 T 细胞的过继转移，以引发临床上有效且特异性的 HGG 患者的免疫反应证明了 CAR T 细胞的安全性。脑肿瘤治疗；然而，仅观察到有限的疗效，这很可能是多因素造成的。包括异源抗原表达、肿瘤的有限归巢和渗透、T 细胞耗竭和有限的持久性，以及免疫抑制肿瘤微环境（TME）。该提案的中心假设是 CAR 的优化设计和额外的基因修饰 T 细胞将提高其抗肿瘤效果，并且这些干预措施将在免疫中进行评估提出了三个相互关联的研究目标，即密切模仿人类疾病的小鼠模型。为了检验这个假设，每个假设的基本原理如下：首先，必须优化 CAR 设计。在免疫活性神经胶质瘤模型中进行评估在目标 1 中，我将生成含有不同辅酶的 CAR T 细胞。刺激结构域（CD28. z、41BB. z、CD28.mut z 和 41BB.mut z），并比较它们的体外和体内活性其次，对原代 T 细胞进行全基因组筛选，确定了 T 细胞激活后的关键调节因子。细胞受体 (TCR) 刺激基因属于调节细胞周期、增殖和细胞增殖的分子。下游 TCR 信号传导，包括 SOCS1、RASA2 或 CBLB，因此，在目标 2 中，我将探索 CRISPR 是否可以。 Cas9 介导的 Socs1、Rasa2 和/或 Cblb 沉默可增强 CAR T 细胞的效应功能。独立的Aim 3随后将探索一种双CAR靶向方法，其中我不仅会靶向神经胶质瘤细胞这些研究还集中在肿瘤微环境中的免疫抑制细胞。相关巨噬细胞（TAM），因为它们大量存在于神经胶质瘤中，并且在塑造为了支持该项目的可行性，我采用了成熟的免疫能力 GL261。胶质瘤模型来研究针对相关肿瘤抗原 B7-H3 的 CAR T 细胞疗法，这不仅是 GL261 细胞表达，而且也在广泛的儿童和成人脑肿瘤中表达。初步研究表明“原型”B7-H3 CAR T 细胞在体外很容易识别并杀死 GL261 细胞并具有体内抗肿瘤活性，强调所开发的模型非常适合所提出的目标该项目将在所有三个目标中使用最先进的技术，不仅研究功能和体内命运。 CAR T 细胞的作用，以及它们的抗肿瘤活性，以及 CAR T 细胞如何与胶质瘤浸润免疫相互作用这项研究的完成将确定最优化的 CAR 设计，以最好地控制 HGG 肿瘤和此外，结果将说明是否针对 TAM。将克服 TME 对 B7-H3 CAR T 细胞的抑制作用。