Hepatocellular carcinoma (HCC) is the fourth most common cause of cancer-related death in the world, and curative systemic therapies are lacking. Chimeric antigen receptor (CAR)-expressing T cells induce robust antitumor responses in patients with hematologic malignancies but have limited efficacy in patients with solid tumors, including HCC. IL15 and IL21 promote T-cell expansion, survival, and function and can improve the antitumor properties of T cells. We explored whether transgenic expression of IL15 and/or IL21 enhanced glypican-3-CAR (GPC3-CAR) T cells' antitumor properties against HCC. We previously optimized the costimulation in GPC3-CARs and selected a second-generation GPC3-CAR incorporating a 4-1BB costimulatory endodomain (GBBz) for development. Here, we generated constructs encoding IL15, IL21, or both with GBBz (15.GBBz, 21.GBBz, and 21.15.GBBz, respectively) and examined the ability of transduced T cells to kill, produce effector cytokines, and expand in an antigen-dependent manner. We performed gene-expression and phenotypic analyses of GPC3-CAR T cells and CRISPR-Cas9 knockout of the TCF7 gene. Finally, we measured GPC3-CAR T-cell antitumor activity in murine xenograft models of GPC3+ tumors. The increased proliferation of 21.15.GBBz T cells was at least in part dependent on the upregulation and maintenance of TCF-1 (encoded by TCF7) and associated with a higher percentage of stem cell memory and central memory populations after manufacturing. T cells expressing 21.15.GBBz had superior in vitro and in vivo expansion and persistence, and the most robust antitumor activity in vivo These results provided preclinical evidence to support the clinical evaluation of 21.15.GPC3-CAR T cells in patients with HCC.

中文翻译：

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共表达 IL15 和 IL21 的 Glypican-3 特异性 CAR T 细胞对肝细胞癌具有卓越的扩增和抗肿瘤活性。


肝细胞癌（HCC）是世界上第四大癌症相关死亡原因，目前缺乏有效的全身治疗方法。表达嵌合抗原受体 (CAR) 的 T 细胞在血液恶性肿瘤患者中诱导强烈的抗肿瘤反应，但在包括 HCC 在内的实体瘤患者中疗效有限。 IL15 和 IL21 促进 T 细胞扩增、存活和功能，并可以提高 T 细胞的抗肿瘤特性。我们探讨了 IL15 和/或 IL21 的转基因表达是否增强了磷脂酰肌醇蛋白聚糖-3-CAR (GPC3-CAR) T 细胞对抗 HCC 的抗肿瘤特性。我们之前优化了 GPC3-CAR 中的共刺激，并选择了包含 4-1BB 共刺激内域 (GBBz) 的第二代 GPC3-CAR 进行开发。在这里，我们生成了编码 IL15、IL21 或两者与 GBBz（分别为 15.GBBz、21.GBBz 和 21.15.GBBz）的构建体，并检查了转导 T 细胞杀伤、产生效应细胞因子和在抗原中扩增的能力依赖方式。我们对 GPC3-CAR T 细胞进行了基因表达和表型分析，并对 TCF7 基因进行了 CRISPR-Cas9 敲除。最后，我们测量了 GPC3+ 肿瘤的小鼠异种移植模型中的 GPC3-CAR T 细胞抗肿瘤活性。 21.15.GBBz T 细胞增殖的增加至少部分依赖于 TCF-1（由 TCF7 编码）的上调和维持，并与制造后较高比例的干细胞记忆和中央记忆群体相关。表达21.15.GBBz的T细胞具有优异的体外和体内扩增和持久性，以及最强的体内抗肿瘤活性。这些结果为支持21.15.GPC3-CAR T细胞在HCC患者中的临床评估提供了临床前证据。