Objective Myeloid-derived suppressor cells (MDSCs) contribute to tumour immunosuppressive microenvironment and immune-checkpoint blockade resistance. Emerging evidence highlights the pivotal functions of cyclin-dependent kinases (CDKs) in tumour immunity. Here we elucidated the role of tumour-intrinsic CDK20, or cell cycle-related kinase (CCRK) on immunosuppression in hepatocellular carcinoma (HCC). Design Immunosuppression of MDSCs derived from patients with HCC and relationship with CCRK were determined by flow cytometry, expression analyses and co-culture systems. Mechanistic studies were also conducted in liver-specific CCRK-inducible transgenic (TG) mice and Hepa1–6 orthotopic HCC models using CRISPR/Cas9-mediated Ccrk depletion and liver-targeted nanoparticles for interleukin (IL) 6 trapping. Tumorigenicity and immunophenotype were assessed on single or combined antiprogrammed death-1-ligand 1 (PD-L1) therapy. Results Tumour-infiltrating CD11b+CD33+HLA-DR− MDSCs from patients with HCC potently inhibited autologous CD8+T cell proliferation. Concordant overexpression of CCRK and MDSC markers (CD11b/CD33) positively correlated with poorer survival rates. Hepatocellular CCRK stimulated immunosuppressive CD11b+CD33+HLA-DR− MDSC expansion from human peripheral blood mononuclear cells through upregulating IL-6. Mechanistically, CCRK activated nuclear factor-κB (NF-κB) via enhancer of zeste homolog 2 (EZH2) and facilitated NF-κB-EZH2 co-binding to IL-6 promoter. Hepatic CCRK induction in TG mice activated the EZH2/NF-κB/IL-6 cascade, leading to accumulation of polymorphonuclear (PMN) MDSCs with potent T cell suppressive activity. In contrast, inhibiting tumorous Ccrk or hepatic IL-6 increased interferon γ+tumour necrosis factor-α+CD8+ T cell infiltration and impaired tumorigenicity, which was rescued by restoring PMN-MDSCs. Notably, tumorous Ccrk depletion upregulated PD-L1 expression and increased intratumorous CD8+ T cells, thus enhancing PD-L1 blockade efficacy to eradicate HCC. Conclusion Our results delineate an immunosuppressive mechanism of the hepatoma-intrinsic CCRK signalling and highlight an overexpressed kinase target whose inhibition might empower HCC immunotherapy.

中文翻译：

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肝癌内源性 CCRK 抑制减少髓源性抑制细胞免疫抑制并增强免疫检查点阻断功效

目的髓源性抑制细胞（MDSCs）有助于肿瘤免疫抑制微环境和免疫检查点阻断抵抗。新出现的证据强调了细胞周期蛋白依赖性激酶 (CDK) 在肿瘤免疫中的关键功能。在这里，我们阐明了肿瘤内在 CDK20 或细胞周期相关激酶 (CCRK) 对肝细胞癌 (HCC) 免疫抑制的作用。通过流式细胞术、表达分析和共培养系统确定源自 HCC 患者的 MDSC 的设计免疫抑制及其与 CCRK 的关系。还在肝脏特异性 CCRK 诱导型转基因 (TG) 小鼠和 Hepa1-6 原位 HCC 模型中进行了机制研究，使用 CRISPR/Cas9 介导的 Ccrk 耗竭和肝脏靶向纳米粒子用于白细胞介素 (IL) 6 捕获。在单一或联合抗程序性死亡 1-配体 1 (PD-L1) 治疗中评估致瘤性和免疫表型。结果 来自 HCC 患者的肿瘤浸润 CD11b+CD33+HLA-DR-MDSC 有效抑制自体 CD8+T 细胞增殖。CCRK 和 MDSC 标志物 (CD11b/CD33) 的一致过度表达与较差的存活率呈正相关。肝细胞 CCRK 通过上调 IL-6 刺激免疫抑制性 CD11b+CD33+HLA-DR-MDSC 从人外周血单核细胞中扩增。从机制上讲，CCRK 通过 zeste 同源物 2 (EZH2) 的增强子激活核因子-κB (NF-κB)，并促进 NF-κB-EZH2 与 IL-6 启动子的共结合。TG 小鼠的肝 CCRK 诱导激活了 EZH2/NF-κB/IL-6 级联，导致具有强效 T 细胞抑制活性的多形核 (PMN) MDSC 的积累。相比之下，抑制肿瘤性 Ccrk 或肝脏 IL-6 增加了干扰素 γ+肿瘤坏死因子-α+CD8+ T 细胞浸润和致瘤性受损，这通过恢复 PMN-MDSCs 得以挽救。值得注意的是，肿瘤 Ccrk 耗竭上调 PD-L1 表达并增加肿瘤内 CD8+ T 细胞，从而增强 PD-L1 阻断功效以根除 HCC。结论我们的结果描绘了肝癌内在 CCRK 信号传导的免疫抑制机制，并突出了一个过度表达的激酶靶点，其抑制可能会增强 HCC 免疫治疗。从而增强 PD-L1 阻断功效以根除 HCC。结论我们的结果描绘了肝癌内在 CCRK 信号传导的免疫抑制机制，并突出了一个过度表达的激酶靶点，其抑制可能会增强 HCC 免疫治疗。从而增强 PD-L1 阻断功效以根除 HCC。结论我们的结果描绘了肝癌内在 CCRK 信号传导的免疫抑制机制，并突出了一个过度表达的激酶靶点，其抑制可能会增强 HCC 免疫治疗。