Immunotherapy alongside IL-15 promote intratumoral cytotoxic CD4+T cell activation against MHC-II-expressing tumors

By killing tumor cells, cytotoxic CD8+ T cells are the major effectors in antitumor immunity. A subset of CD4+ T cells also possesses cytotoxic activity by expressing perforin and granzymes and is associated with immunotherapy efficacy. Despite the progress made in characterizing cytotoxic CD4+ T cells in various diseases, the status of cytotoxic CD4+ T cells in non-small cell lung cancer (NSCLC) and the driving mechanisms and forces involved in reviving intratumoral cytotoxic CD4+ T cells remain unclear. Here, we showed that CD4+GzmB+ T cells obtained from patients with NSCLC expressed increased levels of SLC7A5 compared with their counterparts. Upregulation of SLC7A5 was essential for the differentiation of CD4+GzmB+ T cells from naïve CD4+ T cells stimulated with TCR and IL-2. Both T-bet and Eomes are required for the differentiation of CD4+GzmB+ T cells. Interestingly, IL-15 can further increase SLC7A5 expression in differentiated CD4+GzmB+ T cells. Moreover, through activation of the AKT-FOXO1-T-bet axis, IL-15 increased the effector function of intratumoral CD4+GzmB+ T cells. In addition to IL-15, owing to the unique expression profile of PD-1 and CD85j in tumor-infiltrating CD4+GzmB+ T cells, using patient-derived lung cancer explants, we showed that simultaneous blockade of PD-1 and CD85j promoted the effector function of CD4+GzmB+ T cells by activating the AKT pathway. Importantly, using a mouse model of lung cancer, we demonstrated that intrinsic MHC II expression in cancer cells determines the significance of CD4+GzmB+ T-cell-mediated antitumor immunity in response to immunotherapy. Purified CD4+T cells were enriched from peripheral blood of four healthy donors.Then CD4+T cells were treated with anti-CD3/28 plus IL-15(-com group) or not（-b group) to analyse for RNA-seq.

能够杀伤肿瘤细胞的细胞毒性CD8+ T细胞是抗肿瘤免疫的核心效应细胞。部分CD4+ T细胞可通过表达穿孔素（perforin）和颗粒酶（granzymes）获得细胞毒性活性，且与免疫治疗疗效密切相关。尽管目前已在多种疾病中完成了细胞毒性CD4+ T细胞的特征解析，但非小细胞肺癌（non-small cell lung cancer, NSCLC）中细胞毒性CD4+ T细胞的具体状态，以及唤醒肿瘤内浸润性细胞毒性CD4+ T细胞的驱动机制与调控因素仍未明确。
本研究证实，从非小细胞肺癌患者体内分离得到的CD4+GzmB+ T细胞，相较于对照群体，其SLC7A5的表达水平显著上调。SLC7A5的上调对于经T细胞受体（TCR）与IL-2刺激的初始CD4+ T细胞向CD4+GzmB+ T细胞分化过程至关重要。转录因子T-bet与Eomes均为CD4+GzmB+ T细胞分化所必需的调控因子。值得注意的是，IL-15可进一步提升已分化CD4+GzmB+ T细胞中SLC7A5的表达水平。此外，IL-15可通过激活AKT-FOXO1-T-bet信号轴，增强肿瘤内CD4+GzmB+ T细胞的效应功能。
除IL-15外，鉴于肿瘤浸润性CD4+GzmB+ T细胞具有独特的PD-1与CD85j表达谱，本研究通过患者来源肺癌外植体模型证实，同时阻断PD-1与CD85j可通过激活AKT通路，强化CD4+GzmB+ T细胞的效应功能。尤为关键的是，通过肺癌小鼠模型，本研究证明肿瘤细胞固有的MHC II表达，决定了CD4+GzmB+ T细胞介导的抗肿瘤免疫在免疫治疗响应中的作用效力。
研究人员从4名健康志愿者的外周血中富集纯化CD4+ T细胞，随后将CD4+ T细胞分别用抗CD3/28抗体联合IL-15处理（-com组）与不做该处理（-b组），以开展RNA测序分析。