EGFR-P38 MAPK 轴通过 miR-675-5p 上调 PD-L1，并通过己糖激酶-2 下调 HLA-ABC 在肝癌细胞中。
The EGFR-P38 MAPK axis up-regulates PD-L1 through miR-675-5p and down-regulates HLA-ABC via hexokinase-2 in hepatocellular carcinoma cells.
机构信息
Laboratory of Endocrinology and Metabolism, Guangzhou Women and Children's Medical Center, Guangzhou Medical University, Guangzhou, Guangdong, 510623, P. R. China.
Laboratory of Uterine Vascular Biology, Guangzhou Institute of Pediatrics, Guangzhou Women and Children's Medical Center, Guangzhou Medical University, Guangzhou, Guangdong, 510623, P. R. China.
出版信息
Cancer Commun (Lond). 2021 Jan;41(1):62-78. doi: 10.1002/cac2.12117. Epub 2021 Jan 1.
BACKGROUND
Immunotherapy has been shown to be a promising strategy against human cancers. A better understanding of the immune regulation in hepatocellular carcinoma (HCC) could help the development of immunotherapy against HCC. The epidermal growth factor receptor (EGFR) signaling is frequently activated in HCC and plays important roles in tumorigenesis. However, its role in HCC immunity is still largely unknown. This study aimed to investigate the impact of EGFR signaling on programmed death-ligand 1 (PD-L1) and human leukocyte antigen class-I (HLA-I) expression in HCC cells and its underlying mechanisms.
METHODS
The expression of phosphorylated EGFR (p-EGFR), PD-L1, and HLA-I (HLA-ABC) in HCC specimens was detected by immunohistochemistry, and their correlations were analyzed. PD-L1 and HLA-ABC expression in EGFR-activated HCC cells were detected by quantitative real-time PCR, Western blotting, and flow cytometry, and T cell-mediated lysis was performed to test the immunosuppressive effects of PD-L1 and HLA-ABC alterations in HCC cells. Furthermore, the underlying mechanisms of EGFR activation-induced PD-L1 up-regulation and HLA-ABC down-regulation were explored by animal experiments, luciferase reporter assay, and gene gain- and loss-of-function studies.
RESULTS
p-EGFR was positively correlated with PD-L1 and negatively correlated with HLA-ABC expression in HCCs. EGFR activation by its ligand EGF up-regulated PD-L1 and down-regulated HLA-ABC in HCC cells, which was functionally important and could be abolished by the EGFR inhibitor, gefitinib, both in vitro and in vivo. Mechanistically, enhanced P38 mitogen-activated protein kinase (MAPK) activation down-regulated microRNA-675-5p (miR-675-5p) and up-regulated glycolysis-related enzyme hexokinase 2 (HK2); miR-675-5p down-regulation enhanced the stability of PD-L1 mRNA probably via the 3'-untranslated region (3'-UTR) of PD-L1 and thereby caused PD-L1 accumulation, and HK2 up-regulation enhanced aerobic glycolysis and mediated a decrease in HLA-ABC.
CONCLUSIONS
The EGFR-P38 MAPK axis could up-regulate PD-L1 through miR-675-5p and down-regulate HLA-ABC via HK2 in HCC cells. Our study reveals a novel signaling network that may cause immune suppression in HCC and suggests that EGFR signaling can be targeted for HCC immunotherapy.
背景
免疫疗法已被证明是对抗人类癌症的一种有前途的策略。更好地了解肝细胞癌（HCC）中的免疫调控可能有助于开发针对 HCC 的免疫疗法。表皮生长因子受体（EGFR）信号在 HCC 中经常被激活，并在肿瘤发生中发挥重要作用。然而，其在 HCC 免疫中的作用在很大程度上仍然未知。本研究旨在探讨 EGFR 信号对 HCC 细胞中程序性死亡配体 1（PD-L1）和人类白细胞抗原 I 类（HLA-I）表达的影响及其潜在机制。
方法
通过免疫组织化学检测 HCC 标本中磷酸化 EGFR（p-EGFR）、PD-L1 和 HLA-I（HLA-ABC）的表达，并分析它们之间的相关性。通过定量实时 PCR、Western blot 和流式细胞术检测 EGFR 激活的 HCC 细胞中 PD-L1 和 HLA-ABC 的表达，并进行 T 细胞介导的裂解实验以测试 HCC 细胞中 PD-L1 和 HLA-ABC 改变的免疫抑制作用。此外，通过动物实验、荧光素酶报告基因检测和基因增益和缺失功能研究探索 EGFR 激活诱导的 PD-L1 上调和 HLA-ABC 下调的潜在机制。
结果
p-EGFR 与 HCC 中的 PD-L1 呈正相关，与 HLA-ABC 表达呈负相关。其配体 EGF 激活 EGFR 可上调 HCC 细胞中的 PD-L1 并下调 HLA-ABC，这在体外和体内均可被 EGFR 抑制剂吉非替尼所抑制。从机制上讲，增强的 P38 丝裂原活化蛋白激酶（MAPK）激活下调 microRNA-675-5p（miR-675-5p）并上调糖酵解相关酶己糖激酶 2（HK2）；miR-675-5p 的下调可能通过 PD-L1 mRNA 的 3'-非翻译区（3'-UTR）增强 PD-L1 mRNA 的稳定性，从而导致 PD-L1 积累，而 HK2 的上调增强有氧糖酵解并介导 HLA-ABC 的减少。
结论
EGFR-P38 MAPK 轴可通过 miR-675-5p 上调 PD-L1，并通过 HCC 细胞中的 HK2 下调 HLA-ABC。我们的研究揭示了一个新的信号网络，它可能导致 HCC 中的免疫抑制，并表明 EGFR 信号可以作为 HCC 免疫治疗的靶点。
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