The abnormal upregulation of programmed death ligand-1 (PD-L1) on tumor cells impedes T-cell mediated cytotoxicity through PD-1 engagement, and further exploring the mechanisms regulation of PD-L1 in cancers may enhance the clinical efficacy of PD-L1 blockade. Here, using single-guide RNAs (sgRNAs) screening system, we identify ubiquitin-specific processing protease 2 (USP2) as a novel regulator of PD-L1 stabilization for tumor immune evasion. USP2 directly interacts with and increases PD-L1 abundance in colorectal and prostate cancer cells. Our results show that Thr288, Arg292 and Asp293 at USP2 control its binding to PD-L1 through deconjugating the K48-linked polyubiquitination at lysine 270 of PD-L1. Depletion of USP2 causes endoplasmic reticulum (ER)-associated degradation of PD-L1, thus attenuates PD-L1/PD-1 interaction and sensitizes cancer cells to T cell-mediated killing. Meanwhile, USP2 ablation-induced PD-L1 clearance enhances antitumor immunity in mice via increasing CD8⁺ T cells infiltration and reducing immunosuppressive infiltration of myeloid-derived suppressor cells (MDSCs) and regulatory T cells (Tregs), whereas PD-L1 overexpression reverses the tumor growth suppression by USP2 silencing. USP2-depletion combination with anti-PD-1 also exhibits a synergistic anti-tumor effect. Furthermore, analysis of clinical tissue samples indicates that USP2 is positively associated with PD-L1 expression in cancer. Collectively, our data reveal a crucial role of USP2 for controlling PD-L1 stabilization in tumor cells, and highlight USP2 as a potential therapeutic target for cancer immunotherapy.

肿瘤细胞上程序性死亡配体 1 （PD-L1） 的异常上调阻碍了 T 细胞通过 PD-1 参与介导的细胞毒性，进一步探索 PD-L1 在癌症中的调节机制可能会增强 PD-L1 阻断的临床疗效。在这里，使用单向导 RNA （sgRNA） 筛选系统，我们将泛素特异性加工蛋白酶 2 （USP2） 确定为肿瘤免疫逃避 PD-L1 稳定的新型调节因子。USP2 直接与结直肠癌和前列腺癌细胞相互作用并增加 PD-L1 的丰度。我们的结果表明，USP2 的 Thr288 、 Arg292 和 Asp293 通过解解 PD-L1 赖氨酸 270 位点的 K48 连接多泛素化来控制其与 PD-L1 的结合。USP2 的耗竭导致 PD-L1 的内质网 （ER） 相关降解，从而减弱 PD-L1/PD-1 相互作用并使癌细胞对 T 细胞介导的杀伤敏感。同时，USP2 消融诱导的 PD-L1 清除通过增加 CD8⁺ T 细胞浸润和减少髓源性抑制细胞 （MDSC） 和调节性 T 细胞 （Treg） 的免疫抑制浸润来增强小鼠的抗肿瘤免疫力，而 PD-L1 过表达通过 USP2 沉默逆转肿瘤生长抑制。USP2 耗竭与抗 PD-1 的组合也表现出协同抗肿瘤作用。此外，临床组织样本分析表明 USP2 与癌症中 PD-L1 表达呈正相关。总的来说，我们的数据揭示了 USP2 在控制肿瘤细胞中 PD-L1 稳定的关键作用，并强调了 USP2 作为癌症免疫治疗的潜在治疗靶点。