PD-1 is an immune checkpoint on T cells, and interventions to block this receptor result in T cell activation and enhanced immune response to tumors and pathogens. Reciprocally, despite a decade of research, approaches to treat autoimmunity with PD-1 agonists have only had limited successful. To resolve this, new methods must be developed to augment PD-1 function beyond engaging the receptor. We conducted a flow cytometry analysis of T cells isolated from the peripheral blood and synovial fluid of patients with rheumatoid arthritis. In addition, we performed a genome-wide CRISPR/Cas9 screen to identify genes associated with PD-1 signaling. We further analyzed genes involved in PD-1 signaling using publicly available bulk and single-cell RNA sequencing datasets. Our screen confirmed known regulators in proximal PD-1 signaling and, importantly, identified an additional 1112 unique genes related to PD-1 ability to inhibit T cell functions. These genes were strongly associated with the response of cancer patients to PD-1 blockades and with high tumor immune dysfunction and exclusion scores, confirming their role downstream of PD-1. Functional annotation revealed that the most significant genes uncovered were those associated with known immune regulation processes. Remarkably, these genes were considerably downregulated in T cells isolated from patients with inflammatory arthritis, supporting their overall inhibitory functions. A study of rheumatoid arthritis single-cell RNA sequencing data demonstrated that five genes, KLRG1, CRTAM, SLAMF7, PTPN2, and KLRD1, were downregulated in activated and effector T cells isolated from synovial fluids. Backgating these genes to canonical cytotoxic T cell signatures revealed PD-1+ HLA-DRHIGH KLRG1LOW T cells as a novel inflammatory subset of T cells. We concluded that PD-1+ HLA-DRHIGH KLRG1LOW T cells are a potential target for future PD-1 agonists to treat inflammatory diseases. Our study uncovers new genes associated with PD-1 downstream functions and, therefore, provides a comprehensive resource for additional studies that are much needed to characterize the role of PD-1 in the synovial subset of T cells.

中文翻译：

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PD-1 信号传导揭示了炎症性关节炎中 T 细胞的致病亚群

PD-1 是 T 细胞上的免疫检查点，阻断该受体的干预措施会导致 T 细胞激活并增强对肿瘤和病原体的免疫反应。相反，尽管经过了十年的研究，用 PD-1 激动剂治疗自身免疫的方法只取得了有限的成功。为了解决这个问题，必须开发新的方法来增强 PD-1 的功能，而不仅仅是与受体结合。我们对从类风湿性关节炎患者的外周血和滑液中分离的 T 细胞进行了流式细胞术分析。此外，我们还进行了全基因组 CRISPR/Cas9 筛选，以鉴定与 PD-1 信号传导相关的基因。我们使用公开的批量和单细胞 RNA 测序数据集进一步分析了参与 PD-1 信号传导的基因。我们的筛选证实了近端 PD-1 信号传导中的已知调节因子，重要的是，还鉴定了另外 1112 个与 PD-1 抑制 T 细胞功能的能力相关的独特基因。这些基因与癌症患者对 PD-1 阻断的反应以及高肿瘤免疫功能障碍和排除评分密切相关，证实了它们在 PD-1 下游的作用。功能注释显示，发现的最重要的基因是那些与已知的免疫调节过程相关的基因。值得注意的是，这些基因在从炎性关节炎患者分离的 T 细胞中显着下调，支持其整体抑制功能。类风湿性关节炎单细胞 RNA 测序数据的研究表明，从滑液中分离的激活 T 细胞和效应 T 细胞中，五个基因 KLRG1、CRTAM、SLAMF7、PTPN2 和 KLRD1 下调。将这些基因与典型的细胞毒性 T 细胞特征进行反向门控，发现 PD-1+ HLA-DRHIGH KLRG1LOW T 细胞是 T 细胞的一种新型炎症子集。我们的结论是，PD-1+ HLA-DRHIGH KLRG1LOW T 细胞是未来 PD-1 激动剂治疗炎症性疾病的潜在靶点。我们的研究发现了与 PD-1 下游功能相关的新基因，因此为表征 PD-1 在 T 细胞滑膜亚群中的作用所急需的其他研究提供了全面的资源。