Regulatory T cells converted from T helper 1 cells in tumors suppress cancer immunity via CD39

Although Foxp3-expressing regulatory T (Treg) cells are widely believed to impede anti-tumor immunity, their regulation and functional mechanisms are not well understood. Here, through characterization of multiple cancer models, we identified substantial periphery-induced Treg cells in the tumor microenvironment, depletion of which provoked anti-tumor responses and conferred potent therapeutic effects by increasing CD8+ T cell numbers and cytolytic function. Through fate-mapping and transfer experiments, we found IFN-?-expressing T helper (Th) 1 cells developed into Treg cells in tumors, in response to TGF-ß signaling. Hence, tumor-resident Treg cells highly expressed T-bet, which was essential for their development and function. CD39, highly expressed by T-bet+ Treg cells in both mouse and human tumors, is required for Treg suppression of CD8+ T cell response. In summary, our study has elucidated a developmental pathway of intra-tumoral Treg cells and implicated new ways of targeting them in cancer patients. Overall design: Treg cells contain several functional subsets analogous to CD4+ helper T (Th) cells. Thus, we analyzed if intra-tumoral pTreg cells were similar to any of the Th subsets in their gene expression. We performed transcriptomic analyses on intra-tumoral Treg cells derived from both Hepa1-6 and E.G7 murine tumor models by RNA sequencing. Given that intra-tumoral Treg cells were composed of a substantial proportion of pTreg cells whereas Treg cells derived from lymph nodes (LN) were mainly composed of tTreg cells, we included Treg cells purified from tumor-free LNs as a control. Intra-tumoral Treg cells were sorted from E.G7 and Hepa1-6 tumor-bearing Foxp3YFP-cre mice on day 21 and day 23, respectively. Treg cells sorted from LNs of tumor-free mice were included as a control.

尽管表达Foxp3的调节性T（regulatory T, Treg）细胞被广泛认为会阻碍抗肿瘤免疫，但其调控机制与功能通路尚未得到充分阐释。本研究通过对多种癌症模型进行表征分析，在肿瘤微环境中鉴定出大量外周诱导型调节性T（periphery-induced Treg, pTreg）细胞；耗竭该类细胞可通过提升CD8+ T细胞的数量与溶细胞活性，触发抗肿瘤免疫应答并产生强效治疗效果。通过命运图谱分析与过继转移实验，我们发现表达干扰素-γ（interferon-γ, IFN-γ）的T辅助（T helper, Th）1细胞可在肿瘤内响应转化生长因子-β（transforming growth factor-β, TGF-β）信号，分化为Treg细胞。因此，肿瘤驻留Treg细胞高表达转录因子T-bet，而该分子对于这类细胞的分化与功能至关重要。CD39在小鼠与人类肿瘤中的T-bet+ Treg细胞表面均呈高表达，且该分子是Treg细胞抑制CD8+ T细胞应答所必需的。综上，本研究阐明了肿瘤内Treg细胞的一条发育通路，并为癌症患者靶向该类细胞提供了全新的治疗策略。

实验整体设计：调节性T细胞存在多个功能亚群，其分类模式与CD4+辅助T（Th）细胞类似。因此，我们分析了肿瘤内pTreg细胞的基因表达谱是否与某一类Th亚群相似。我们通过RNA测序（RNA sequencing）对来自Hepa1-6与E.G7两种小鼠肿瘤模型的肿瘤内Treg细胞进行了转录组分析。鉴于肿瘤内Treg细胞主要由pTreg细胞构成，而淋巴结（lymph node, LN）来源的Treg细胞则主要为天然胸腺来源Treg（thymic Treg, tTreg），我们同时纳入了来自无瘤小鼠淋巴结的Treg细胞作为对照。分别于造模后第21天与第23天，从携带E.G7与Hepa1-6肿瘤的Foxp3YFP-cre小鼠体内分选肿瘤内Treg细胞；同时收集无瘤小鼠淋巴结来源的Treg细胞作为对照样本。