接受富含记忆T细胞的嵌合抗原受体（CAR-T）细胞治疗的病人，在CAR-T细胞扩增和持续性增加的结果下，表现出更好的疾病控制。人类记忆T细胞包括干细胞样CD8+记忆T细胞前体，可以成为功能性干细胞样T（TSTEM）细胞或非功能性的T前体耗竭（TPEX）细胞。为此，我们证明在测试卢易斯Y-CAR-T细胞（NCT03851146）临床试验中，静脉注入的CAR-T细胞制品中TSTEM细胞数量较少，并且在患者中，注入的CAR-T细胞显示持续性较差。为了解决这个问题，我们开发了生产方案，以生成TSTEM样CAR-T细胞，富含细胞复制途径基因的表达。相比传统的CAR-T细胞，TSTEM样CAR-T细胞具有增强的增殖能力和CAR刺激后的细胞因子分泌，包括体外慢性CAR刺激后的细胞因子分泌。这些反应取决于TSTEM样CAR-T细胞生成过程中CD4+ T细胞的存在。TSTEM样CAR-T细胞的移植诱导在临床前模型中建立的肿瘤得到更好的控制和对肿瘤再挑战的抵抗力。这些更有利的结果与TSTEM样CAR-T细胞的持续性增加和记忆T细胞数量增加有关。最后，TSTEM样CAR-T细胞和抗程序性细胞死亡蛋白1（PD-1）治疗消灭了建立的肿瘤，并且这与产生干扰素γ的肿瘤浸润CD8+CAR+ T细胞数量的增加有关。总之，我们的CAR-T细胞方案生成了TSTEM样CAR-T细胞，在体内具有增强的治疗效果，结果包括增加的增殖能力和持续性。

Patients who receive chimeric antigen receptor (CAR)-T cells that are enriched in memory T cells exhibit better disease control as a result of increased expansion and persistence of the CAR-T cells. Human memory T cells include stem-like CD8+ memory T cell progenitors that can become either functional stem-like T (TSTEM) cells or dysfunctional T progenitor exhausted (TPEX) cells. To that end, we demonstrated that TSTEM cells were less abundant in infused CAR-T cell products in a phase 1 clinical trial testing Lewis Y-CAR-T cells (NCT03851146), and the infused CAR-T cells displayed poor persistence in patients. To address this issue, we developed a production protocol to generate TSTEM-like CAR-T cells enriched for expression of genes in cell replication pathways. Compared with conventional CAR-T cells, TSTEM-like CAR-T cells had enhanced proliferative capacity and increased cytokine secretion after CAR stimulation, including after chronic CAR stimulation in vitro. These responses were dependent on the presence of CD4+ T cells during TSTEM-like CAR-T cell production. Adoptive transfer of TSTEM-like CAR-T cells induced better control of established tumors and resistance to tumor rechallenge in preclinical models. These more favorable outcomes were associated with increased persistence of TSTEM-like CAR-T cells and an increased memory T cell pool. Last, TSTEM-like CAR-T cells and anti-programmed cell death protein 1 (PD-1) treatment eradicated established tumors, and this was associated with increased tumor-infiltrating CD8+CAR+ T cells producing interferon-γ. In conclusion, our CAR-T cell protocol generated TSTEM-like CAR-T cells with enhanced therapeutic efficacy, resulting in increased proliferative capacity and persistence in vivo.