Genetic Approaches to Optimize CAR T cells for Glioblastoma Therapy

胶质母细胞瘤(GBM)是最常见和侵袭性最强的原发脑肿瘤。 不治之症。因此，迫切需要开发新的治疗方法。转基因T细胞表达 嵌合抗原受体(Car)有可能作为一种独特的细胞毒工具来特异性靶向。 GBM。CAR T细胞疗法已经成功地治疗了血液系统的恶性肿瘤，但带来了多重挑战 脑肿瘤环境需要多方面的方法，才能使CAR T细胞成功地治疗GBM。学习 为此，我们开发了一种针对白介素13Rα2的单链可变区(ScFv)，白介素13R是一种与基底膜相关的肿瘤 并产生了IL13Rα2-CAR。IL13Rα2-CAR T细胞只能识别IL13Rα2阳性胶质瘤 细胞，并在临床前异种移植和免疫活性动物模型中具有抗胶质瘤活性。然而， 肿瘤最终复发，与人类的情况相似。治疗失败的主要原因包括：(一) CAR T细胞在免疫抑制的肿瘤环境中不能持续存在，(Ii)抗原丢失变体 当以单一抗原为靶点时，以及(Iii)CAR T细胞无法有效地运输到肿瘤部位 肿瘤产生的趋化因子与CAR T细胞表达的趋化因子受体不匹配。在……里面 机制研究，我们已经证明了有限的IL13Rα2-CAR T细胞的持久性和发展 抗原丢失变异体。此外，我们在异种移植模型中显示，IL15在CAR T中的转基因表达 细胞增强了它们的持久性和抗胶质瘤活性。然而，这些异种移植研究是有限的；目标是 R01的目的是在具有免疫能力的动物模型中进行机制研究，并评估遗传 增强IL13Rα-CAR 2-CAR T细胞抗胶质瘤活性的途径因此，我们现在假设 IL13Rα2-CAR T细胞可以通过进一步的基因工程来优化其抗基底膜活性 增强其持久性，靶向多种肿瘤抗原，改善其向肿瘤的转运 网站。目的1研究表达IL15的CAR T细胞能否抵抗免疫抑制肿瘤 同基因基底膜模型中的环境。Aim 2优化CAR T细胞以同时针对IL13Rα2和EphA2，2 胶质瘤相关抗原。目标3调查CAR T细胞向GBM的贩运是否可以通过 CCR2的转基因表达，CCR2是一种识别CCL2的趋化因子受体，CCL2是一种由GBMS产生的趋化因子。 在赠款结束时，我们将解决CAR T细胞治疗GBM的三个主要障碍。而当 我们将使用我们的数据来证明使用优化的IL13Rα2-CAR T细胞进行未来临床研究的合理性 对于患有GBMS的患者；我们改进的T细胞疗法应该适用于广泛的固体 肿瘤。

Glioblastoma (GBM), the most frequently occurring and aggressive primary brain tumor, remains virtually incurable. Thus, there is an urgent need to develop new therapies. Genetically modified T cells expressing chimeric antigen receptors (CARs) have the potential to serve as a unique cytotoxic tool to specifically target GBM. CAR T cell therapy has been successful for hematological malignancies, but multiple challenges posed by the brain tumor environment require a multifaceted approach for CAR T cells to succeed for GBM. To study this, we have developed a single-chain variable fragment (scFv) specific for IL13Rα2, a GBM-associated tumor antigen, and have generated an IL13Rα2-CAR. IL13Rα2-CAR T cells only recognize IL13Rα2-positive glioma cells and had anti-glioma activity in preclinical xenograft and immune-competent animal models. However, tumors eventually recurred, paralleling the situation in humans. Major causes of treatment failure include (i) the inability of CAR T cells to persist within an immunosuppressive tumor environment, (ii) antigen-loss variants when a single antigen is targeted, and (iii) the inability of CAR T cells to efficiently traffic to tumor sites due to a mismatch between chemokines produced by the tumor and chemokine receptors expressed by CAR T cells. In mechanistic studies, we have demonstrated limited IL13Rα2-CAR T cell persistence and the development of antigen-loss variants. In addition, we showed in xenograft models that transgenic expression of IL15 in CAR T cells enhances their persistence and anti-glioma activity. However, these xenograft studies are limited; the goal of this R01 is to perform mechanistic studies in immune-competent animal models and evaluate genetic approaches to enhance the anti-glioma activity of IL13Rα2-CAR T cells. Thus, we now hypothesize that IL13Rα2-CAR T cells can be further genetically engineered to optimize their anti-GBM activity by enhancing their persistence, targeting multiple tumor antigens, and improving their trafficking to tumor sites. Aim 1 investigates whether IL15-expressing CAR T cells can resist the immunosuppressive tumor environment in syngeneic GBM models. Aim 2 optimizes CAR T cells to target both IL13Rα2 and EphA2, two glioma-associated antigens. Aim 3 investigates if trafficking of CAR T cells to GBMs can be improved by the transgenic expression of CCR2, a chemokine receptor that recognizes CCL2, a chemokine produced by GBMs. At the conclusion of the grant, we will have addressed three major hurdles of CAR T cell therapy for GBM. While we will use our data to justify the development of a future clinical study utilizing optimized IL13Rα2-CAR T cells for patients with GBMs; our modified approach to T cell therapy should be applicable to a broad range of solid tumors.