Regulatory T cells converted from T helper 1 cells in tumors suppress cancer immunity via CD39. 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-γ-expressing T helper (Th) 1 cells developed into Treg cells in tumors, in response to TGF-β 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细胞（regulatory T cells, Treg）被广泛认为会阻碍抗肿瘤免疫，但其调控机制与功能通路仍未被充分阐明。本研究通过对多种癌症模型开展表征分析，在肿瘤微环境中发现了大量外周诱导型Treg细胞（periphery-induced Treg, pTreg）；耗竭这群细胞可通过提升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（IFN-γ）的辅助性T细胞1（Th1）可在肿瘤微环境中响应转化生长因子β（TGF-β）信号，分化为Treg细胞。因此，肿瘤驻留型Treg细胞高表达T-bet，该分子对于这类细胞的分化与功能至关重要。在小鼠与人类肿瘤中，T-bet阳性Treg细胞均高表达CD39，而该分子是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细胞的分化通路，并为癌症患者靶向这群细胞提供了全新策略。

实验设计：Treg细胞存在多种功能亚群，与CD4+辅助性T细胞（Th）类似。因此，我们通过基因表达分析，探究肿瘤内外周诱导型Treg细胞的基因表达谱是否与某类Th亚群相似。我们采用RNA测序技术，对来自Hepa1-6与E.G7两种小鼠肿瘤模型的肿瘤内Treg细胞进行转录组分析。鉴于肿瘤内Treg细胞主要由外周诱导型Treg细胞构成，而淋巴结（lymph nodes, LN）来源的Treg细胞则以胸腺来源的Treg（thymus-derived Treg, tTreg）为主，因此我们将从无瘤小鼠淋巴结中纯化得到的Treg细胞作为对照。我们分别于接种后第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，并将从无瘤小鼠淋巴结中分选得到的Treg细胞纳入对照实验。