IL-15 and -21 armored GPC3-specific CAR T cells for children with solid tumors

CAR T细胞可以在血液病患者中诱导显著的抗肿瘤反应，但一直以来 对实体瘤患者基本无效。几个障碍限制了实体恶性肿瘤的CAR T疗效 包括有限数量的安全实体肿瘤特异性抗原和免疫抑制肿瘤 缺乏支持刺激的微环境(TME)。此外，尚不清楚CAR T细胞是如何 影响人类TME免疫逃逸机制的进化。我们开发并测试了一组 嵌合抗原受体(CARS)：1)靶向GPC3的嵌合抗原受体，GPC3是一种选择性表达的肿瘤抗原 2)共表达白介素15和白介素21， 细胞因子对T细胞存活、扩增、持久和保留抗肿瘤效应的重要作用 功能性。我们发现T细胞共表达第二代GPC3-CAR和IL15(15.GPC3-CAR) 与GPC3-CAR T细胞相比，显著提高了扩增、持久性和抗肿瘤活性，并且 IL15和IL21(15.21.GPC3-CAR)共同表达进一步增强了临床前的这些特性 模特们。此外，我们的临床前研究表明，15.21GPC3-CAR T细胞独特地维持表达 T细胞因子-1，它可以保护CAR T细胞免于衰竭，并保持以下的增殖能力 GPC3介导的重复激活。由于这些细胞因子表达载体的安全性和有效性参数 人类的T细胞是未知的，我们建议评估15.GPC3-CAR和15.21 GPC3-CAR的T细胞 应用贝叶斯最佳间隔剂量递增治疗复发或难治性儿童的序贯1期研究 GPC3阳性实体瘤。我们的多学科团队已经产生了有希望的安全性和有效性 结果单独治疗T细胞表达GPC3-CAR的儿童。我们假设15.GPC3-Car 15.21 GPC3-CAR T细胞将是安全的，IL15和IL21的共表达将使GPC3-CAR T细胞能够 克服抑制的TME，产生强大的扩张性、持久性和持久的抗肿瘤反应。在……里面 目的1、评价15.GPC3-CAR和15.21-GPC3-CAR T细胞在GPC3阳性患儿中的安全性 实体瘤。在目标2中，我们将测定CAR T细胞的体内持久性和抗肿瘤活性。最后，在 目的3，通过对CAR T细胞产物、外周血样本和输注后肿瘤的评估 活检，我们将定义GPC3-CAR T细胞，癌细胞和非-CAR细胞的转录和表型特征 肿瘤间质细胞。我们将确定CAR T细胞的生存计划，并定义免疫逃逸 使用单细胞RNA测序、流式细胞术和CO指数法检测TME中的机制。 通过拟议的研究，我们将确定安全性、体内扩展和持久性以及抗肿瘤 我们的治疗细胞的活性。结果将确定CAR T细胞的存活计划和免疫逃逸 人类TME的机制为开发提高疗效的有效措施提供了蓝图 针对实体瘤患者的CAR和其他免疫疗法。

CAR T cells can induce remarkable antitumor responses in patients with hematologic cancers but have been largely ineffective in patients with solid tumors. Several barriers limit CAR T efficacy in solid malignancies including the limited number of safe solid tumor-specific antigens and an immunosuppressive tumor microenvironment (TME) that lacks supporting stimuli. Furthermore, it remains unknown how CAR T cells influence the evolution of immune escape mechanisms in the human TME. We developed and tested a set of chimeric antigen receptors (CARs) that 1) target glypican-3 (GPC3), a tumor antigen selectively expressed by several pediatric solid tumors but not non-malignant tissues, and 2) co-express interleukin-15 (IL15) and IL21, cytokines that are important for T cell survival, expansion, persistence, and retention of antitumor effector functionality. We found that T cells co-expressing a second generation GPC3-CAR and IL15 (15.GPC3-CAR) have significantly improved expansion, persistence, and antitumor activity compared to GPC3-CAR T cells, and that co-expression of both IL15 and IL21 (15.21.GPC3-CAR) further enhances these properties in preclinical models. Additionally, our preclinical studies showed that 15.21.GPC3-CAR T cells uniquely maintain expression of T cell factor-1, which protects CAR T cells from exhaustion and preserves proliferative capacity following repeated GPC3-mediated activation. Since the safety and efficacy parameters of these cytokine-expressing CAR T cells in humans are unknown, we propose to evaluate 15.GPC3-CAR and 15.21.GPC3-CAR T cells in sequential phase 1 studies using Bayesian Optimal Interval dose escalation in children with relapsed or refractory GPC3-positive solid tumors. Our multidisciplinary team has already generated promising safety and efficacy results treating children with T cells expressing the GPC3-CAR alone. We hypothesize that 15.GPC3-CAR and 15.21.GPC3-CAR T cells will be safe and that co-expression of IL15 with IL21 will enable GPC3-CAR T cells to overcome the inhibitory TME, resulting in robust expansion, persistence, and durable antitumor responses. In Aim 1, we will evaluate the safety of 15.GPC3-CAR and 15.21.GPC3-CAR T cells in children with GPC3-positive solid tumors. In Aim 2, we will determine the in vivo persistence and antitumor activity of CAR T cells. Finally, in Aim 3, through the evaluation of CAR T cell products, peripheral blood samples, and post-infusion tumor biopsies, we will define the transcriptomic and phenotypic profiles of GPC3-CAR T cells, cancer cells, and non- neoplastic stromal cells. We will identify survival programs in CAR T cells and define immune escape mechanisms in the TME using single-cell RNA sequencing, flow cytometry, and COdetection-by-inDEXing. Through the proposed research, we will determine the safety, in vivo expansion and persistence, and antitumor activity of our therapeutic cells. The results will identify survival programs in CAR T cells and immune escape mechanisms in the human TME to provide a blueprint for developing effective measures that boost the efficacy of CAR-based and other immunotherapies for patients with solid tumors.