Department of Molecular oncology and immunology, Netherlands Cancer Institute, Oncode Institute, Amsterdam, The Netherlands.
Cancer Data Science Laboratory, National Cancer Institute Center for Cancer Research, Bethesda, Maryland, USA.
J Immunother Cancer. 2024 Nov 7;12(11):e010145. doi: 10.1136/jitc-2024-010145.
Blockade of the programmed cell death protein 1 (PD-1) immune checkpoint (ICB) is revolutionizing cancer therapy, but little is known about the mechanisms governing its expression on CD8 T cells. Because PD-1 is induced during activation of T cells, we set out to uncover regulators whose inhibition suppresses PD-1 abundance without adversely impacting on T cell activation.
To identify PD-1 regulators in an unbiased fashion, we performed a whole-genome, fluorescence-activated cell sorting (FACS)-based CRISPR-Cas9 screen in primary murine CD8 T cells. A dual-readout design using the activation marker CD137 allowed us to uncouple genes involved in PD-1 regulation from those governing general T cell activation.
We found that the inactivation of one of several members of the TMED/EMP24/GP25L/p24 family of transport proteins, most prominently TMED10, reduced PD-1 cell surface abundance, thereby augmenting T cell activity. Another client protein was cytotoxic T lymphocyte-associated protein 4 (CTLA-4), which was also suppressed by TMED inactivation. Treatment with TMED inhibitor AGN192403 led to lysosomal degradation of the TMED-PD-1 complex and reduced PD-1 abundance in tumor-infiltrating CD8 T cells (TIL) in mice, thus reversing T cell dysfunction. Clinically corroborating these findings, single-cell RNA analyses revealed a positive correlation between TMED expression in CD8 TIL, and both a T cell dysfunction signature and lack of ICB response. Similarly, patients receiving a TIL product with high TMED expression had a shorter overall survival.
Our results uncover a novel mechanism of PD-1 regulation, and identify a pharmacologically tractable target whose inhibition suppresses PD-1 abundance and T cell dysfunction.
程序性细胞死亡蛋白 1（PD-1）免疫检查点（ICB）的阻断正在彻底改变癌症治疗方法，但对于其在 CD8 T 细胞上表达的调控机制知之甚少。由于 PD-1 在 T 细胞激活过程中诱导产生，我们旨在揭示抑制其表达而不影响 T 细胞激活的调控因子。
为了以无偏倚的方式识别 PD-1 的调控因子，我们在原代小鼠 CD8 T 细胞中进行了基于全基因组、荧光激活细胞分选（FACS）的 CRISPR-Cas9 筛选。使用激活标志物 CD137 的双读数设计使我们能够将参与 PD-1 调控的基因与那些调控一般 T 细胞激活的基因区分开来。
我们发现，几种跨膜蛋白/EMP24/糖蛋白 25L/p24 家族（TMED/EMP24/GP25L/p24）转运蛋白成员中的一个或多个（最显著的是 TMED10）的失活会降低 PD-1 细胞表面丰度，从而增强 T 细胞活性。另一个客户蛋白是细胞毒性 T 淋巴细胞相关蛋白 4（CTLA-4），它也被 TMED 失活抑制。用 TMED 抑制剂 AGN192403 处理会导致 TMED-PD-1 复合物的溶酶体降解，并降低小鼠肿瘤浸润性 CD8 T 细胞（TIL）中的 PD-1 丰度，从而逆转 T 细胞功能障碍。这些发现得到了临床验证，单细胞 RNA 分析显示 TMED 在 CD8 TIL 中的表达与 T 细胞功能障碍特征和缺乏 ICB 反应之间呈正相关。同样，接受 TMED 表达水平较高的 TIL 产品的患者总生存期较短。
我们的研究结果揭示了 PD-1 调控的新机制，并确定了一种具有治疗潜力的药物靶点，其抑制作用可降低 PD-1 丰度和 T 细胞功能障碍。
