GENETICALLY ENGINEERED T CELLS FOR DIPG

用于 DIPG 的基因工程 T 细胞

• 批准号: 10672286
• 负责人: Giedre Krenciute
• 金额: $ 50.36万
• 依托单位: ST. JUDE CHILDREN'S RESEARCH HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-01 至 2027-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10672286
• 关键词: Ablation; Address; Adoptive Transfer; Adult; Affect; Antigens; Antitumor Response; Brain Neoplasms; CAR T cell therapy; CD276 gene; CD28 gene; CRISPR screen; Cancer Etiology; Cell Communication; Cell Therapy; Cell physiology; Cell surface; Cells; Cellular biology; Child; Childhood Brain Neoplasm; Clinical; Clinical Research; Data; Development; Diffuse intrinsic pontine glioma; Disease model; Endoplasmic Reticulum; Engineering; Evaluation; Flow Cytometry; Frequencies; GRP78 gene; Gene Deletion; Genes; Genetic Engineering; Goals; Golgi Apparatus; Human; Immune; Immune response; Immunocompetent; Immunologic Deficiency Syndromes; Immunooncology; Immunotherapy; In Vitro; Intervention; Intrinsic factor; Knock-out; Knowledge; Learning; Literature; Macrophage; Mediating; Methods; Modeling; Molecular; Molecular Target; Mouse Strains; Mus; Myeloid Cells; Outcome; Pathway interactions; Patients; Phase; Phenotype; Population; Proteins; Publishing; Role; Safety; Signal Pathway; Solid Neoplasm; Subgroup; Surface; T-Cell Activation; T-Cell Receptor; T-Lymphocyte; TNFRSF5 gene; Testing; Translations; Tumor Antigens; Xenograft Model; antigen-specific T cells; cancer cell; childhood cancer mortality; chimeric antigen receptor; chimeric antigen receptor T cells; clinically relevant; cytokine; design; diffuse midline glioma; effective therapy; effector T cell; efficacy evaluation; endoplasmic reticulum stress; engineered T cells; eosinophil; exhaustion; experience; genome wide screen; genome-wide; glioma cell line; glucose-regulated proteins; human disease; immune cell infiltrate; improved; improved outcome; in vitro activity; in vivo; innovative technologies; monocyte; mouse model; multidisciplinary; neoplastic cell; novel; novel therapeutics; pediatric patients; pre-clinical; prototype; response; safety testing; tool; tumor; tumor heterogeneity; tumor-immune system interactions

PROJECT ABSTRACT Genetically engineered T cells for DIPG The overall objective of this study is to engineer an effective and safe chimeric antigen receptor (CAR) T-cell therapy for diffuse intrinsic pontine glioma (DIPG), a subgroup of diffuse midline gliomas for which currently there is no cure. 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 adult patients demonstrated the safety of CAR T-cell therapy for brain tumors, yet found only limited benefits. This lack of efficacy is most likely multifactorial and includes T-cell exhaustion, the immunosuppressive tumor microenvironment, and the paucity of brain tumor-specific antigens. Thus, we propose to develop CAR T cells targeting glucose-regulated protein 78 (GRP78), a novel DIPG-specific antigen. To further improve the effector function of GRP78-CAR T cells, we also propose to target a novel negative T-cell regulator called RASA2. We hypothesize that CAR T cells targeting GRP78 on the surface of tumor cells and lacking RASA2 can be developed as a safe, effective treatment for DIPGs and that evaluation of this intervention in DIPG mouse models that closely mimic human disease will identify T-cell extrinsic negative regulators of CAR T-cell function. Here we propose 3 interrelated Specific Aims to test this hypothesis. The rationale of each Aim is outlined below. In Aim 1, we will first target GRP78 and optimize the CAR design. To that end, we will generate CARs containing different co-stimulatory domains (41BB.ζ, MyD88.ζ, or MyD88.CD40.ζ) and compare their activity in vitro. Then we will evaluate the efficacy and safety of the active CAR(s) in vivo by using immunodeficient and immunocompetent mouse models of DIPG. A genome-wide screen in primary T cells has identified key regulators of T-cell activation after T-cell receptor stimulation. One of the identified genes, RASA2, has not been studied in T cells. In Aim 2, we will elucidate the mechanism by which the deletion of RASA2 enhances the effector function of CAR T cells and then delete the gene from GRP78-CAR T cells to enhance their function. Finally, in Aim 3, we will characterize the components of the DIPG microenvironment and examine the interactions between endogenous DIPG immune cells and CAR T–cell treatment. State-of-the-art methods will be used in all 3 Aims to study not only the function and in vivo fate of CAR T cells but also their antitumor activity and how they interact with DIPG-infiltrating immune cells (i.e., macrophages, eosinophils, monocytes). Our preliminary studies indicate that prototype GRP78-CAR T cells readily recognize and kill DIPG cells in vitro and have antitumor activity against solid tumors in vivo, highlighting that the developed models are well suited for the proposed Aims. To support the rapid translation of findings from this project to the clinical setting, we will use multiple DIPG mouse models that closely recapitulate human disease. Upon completion of this study, we will have defined the optimal GRP78-CAR design that safely eliminates DIPG tumors and persists long term in the DIPG immune microenvironment.

项目摘要 用于DIPG的基因工程化T细胞 本研究的总体目标是设计有效和安全的嵌合抗原受体（CAR）T细胞 弥漫性内在脑桥胶质瘤（DIPG）的治疗，弥漫性中线胶质瘤的一个亚组，目前有 无法治愈CAR T细胞疗法是一种基于抗原特异性T细胞过继转移的创新技术 工程化以引发针对肿瘤细胞的临床有效和特异性免疫应答。早期临床研究 在成人患者中的研究证明了CAR T细胞治疗脑肿瘤的安全性，但仅发现有限的益处。 这种疗效的缺乏很可能是多因素的，包括T细胞耗竭，免疫抑制肿瘤 微环境和脑肿瘤特异性抗原的缺乏。因此，我们建议开发CAR T细胞， 靶向葡萄糖调节蛋白78（GRP 78），一种新的DIPG特异性抗原。为了进一步改善效应器 为了研究GRP 78-CAR T细胞的功能，我们还提出靶向一种称为RASA 2的新型负性T细胞调节因子。我们 假设靶向肿瘤细胞表面上的GRP 78且缺乏RASA 2的CAR T细胞可以 作为DIPG的安全、有效治疗开发，并在DIPG小鼠中评价这种干预 密切模拟人类疾病的模型将识别CAR T细胞的T细胞外在负调节因子 功能在这里，我们提出了三个相互关联的具体目标来检验这一假设。每个目标的基本原理是 概述如下。在目标1中，我们将首先针对GRP 78并优化CAR设计。为此，我们将 CD40.1）的汽车，并比较它们的表达。 体外活性然后，我们将通过使用以下方法评估活性CAR在体内的功效和安全性： DIPG的免疫缺陷和免疫活性小鼠模型。在原代T细胞中进行全基因组筛选 确定了T细胞受体刺激后T细胞活化的关键调节因子。其中一个已鉴定的基因RASA 2， 尚未在T细胞中进行研究。在目的2中，我们将阐明RASA 2缺失的机制。 增强CAR T细胞的效应子功能，然后从GRP 78-CAR T细胞中删除该基因以增强CAR T细胞的效应子功能。 它们的功能。最后，在目标3中，我们将描述DIPG微环境的组成部分，并检查 内源性DIPG免疫细胞和CAR T细胞治疗之间的相互作用。最先进的方法 将用于所有3个目的，不仅研究CAR T细胞的功能和体内命运，还研究它们的抗肿瘤作用。 活性以及它们如何与DIPG浸润免疫细胞相互作用（即，巨噬细胞、嗜酸性粒细胞、单核细胞）。 我们的初步研究表明，原型GRP 78-CAR T细胞在体外容易识别和杀死DIPG细胞， 并具有体内抗实体瘤的抗肿瘤活性，突出表明所开发的模型非常适合 为了提出的目标。为了支持将本项目的发现快速转化为临床环境，我们将 使用多种密切概括人类疾病的DIPG小鼠模型。在完成这项研究后，我们 将定义最佳的GRP 78-CAR设计，安全地消除DIPG肿瘤，并在 DIPG免疫微环境。