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T细胞 本研究的总体目标是设计一种有效和安全的嵌合抗原受体(CAR)T细胞 弥漫性固有桥脑胶质瘤(DIPG)的治疗，这是弥漫性中线胶质瘤的一个亚群，目前在 是无法治愈的。CAR T细胞疗法是一项基于过继转移抗原特异性T细胞的创新技术 经改造后可在临床上引发针对肿瘤细胞的有效和特异的免疫反应。早期临床研究 在成人患者中证明了CAR T细胞治疗脑瘤的安全性，但发现益处有限。 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，以及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(GRP78)，一种新的DIPG特异性抗原。进一步改进效应器 为了进一步研究GRP78-CAR T细胞的功能，我们还提出了一种新的负性T细胞调节因子RASA2。我们 假设靶向肿瘤细胞表面GRP78且缺乏RASA2的CAR T细胞 作为一种安全、有效治疗DIPG的方法及其在DIPG小鼠中的干预评价 接近模拟人类疾病的模型将识别T细胞对CAR T细胞的外在负调控 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说。每个目标的基本原理是 概述如下。在目标1中，我们将首先瞄准GRP78，并优化汽车设计。为此，我们将产生 包含不同共刺激结构域(41BB.ζ、MyD88.ζ或MyD88.CD40.ζ)的CARS，并比较它们的 体外活性。然后，我们将评估活体CAR(S)的有效性和安全性 免疫缺陷和免疫活性的DIPG小鼠模型。在初级T细胞中的全基因组筛查 确定了T细胞受体刺激后T细胞激活的关键调节因子。其中一个被确认的基因RASA2， 还没有在T细胞中进行过研究。在目标2中，我们将阐明RASA2缺失的机制 增强CAR T细胞的效应功能，然后从GRP78-CAR T细胞中删除该基因以增强 它们的功能。最后，在目标3中，我们将描述DIPG微环境的组成并检查 内源性DIPG免疫细胞与CAR T细胞治疗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，而且研究它们的抗肿瘤作用 以及它们如何与渗透DIPG的免疫细胞(即巨噬细胞、嗜酸性粒细胞、单核细胞)相互作用。 我们的初步研究表明，原型GRP78-CAR T细胞在体外很容易识别和杀伤DIPG细胞 并在体内对实体瘤具有抗肿瘤活性，突出表明所开发的模型非常适合 对于提议的目标。为了支持从这个项目到临床环境的研究成果的快速转化，我们将 使用与人类疾病相似的多种DIPG小鼠模型。在完成这项研究后，我们 将定义最优的GRP78-CAR设计，安全地消除DIPG肿瘤并长期存在 DIPG免疫微环境。