EB 病毒编码的 miR-BART5-5p 通过 PIAS3/pSTAT3 调节上调 PD-L1，使 PD-L1 阳性胃癌的临床预后恶化。

Epstein-Barr virus-encoded miR-BART5-5p upregulates PD-L1 through PIAS3/pSTAT3 modulation, worsening clinical outcomes of PD-L1-positive gastric carcinomas.

机构信息

Department of Infectious Diseases, Asan Medical Center, Asan Institute for Life Sciences, University of Ulsan College of Medicine, 88 Olympic-ro 43-gil, Songpa-gu, Seoul, 05505, Republic of Korea.

Department of Pathology, Seoul National University Boramae Hospital, Seoul National University College of Medicine, 20 Boramae-ro 5-gil, Dongjak-gu, Seoul, 07061, Republic of Korea.

出版信息

Gastric Cancer. 2020 Sep;23(5):780-795. doi: 10.1007/s10120-020-01059-3. Epub 2020 Mar 23.

DOI:10.1007/s10120-020-01059-3

PMID:32206940

Abstract

BACKGROUND

Epstein-Barr virus (EBV) is etiologically associated with ~ 10% of all gastric carcinomas. However, the molecular mechanisms and roles of EBV miRNAs in gastric carcinoma oncogenesis are yet to be elucidated.

METHODS

MicroRNA microarray and TaqMan quantitative real-time RT-PCR were conducted. RT-PCR and luciferase reporter assay for PIAS3, western blotting for 20 proteins, immunofluorescence for STAT3, transfection with miRBART5-5p-plasmid, STAT3-plasmid, miRBART5-5p mimic, or PIAS3-siRNA, and in vitro assays for biological effects of PD-L1 were implemented. In situ hybridization for EBV-encoded small RNAs and immunohistochemistry were performed on gastric carcinoma tissues.

RESULTS

Transfecting miR-BART5-5p into EBV(-) gastric carcinoma cell lines caused a decrease in PIAS3 3'-UTR reporter activity, PIAS3 downregulation, and subsequent STAT3 activation followed by PIAS3/pSTAT3-dependent PD-L1 upregulation. Interestingly, due to PD-L1 knockdown, apoptosis was increased, while the rate of cell proliferation, invasion capacity, and migration were decreased in miR-BART5-5p-transfected cells. In EBV(+) gastric carcinoma cells, anti-miR-BART5-5p reduced PD-L1 levels through PIAS3/pSTAT3 control. Among 103 patients with EBV-associated gastric carcinomas, overall survival was significantly shortened for those with PD-L1(+) tumors compared to those with PD-L1(-) tumors (P = 0.049).

CONCLUSIONS

Our findings imply that miR-BART5-5p directly targets PIAS3 and augments PD-L1 through miR-BART5/PIAS3/pSTAT3/PD-L1 axis control. This contributes to antiapoptosis, tumor cell proliferation, invasion and migration, as well as immune escape, furthering gastric carcinoma progression and worsening the clinical outcome, especially in the PD-L1(+) group of patients with EBV-associated gastric carcinomas. miR-BART5-5p may, therefore, be amenable to PD-1/PD-L1 immune checkpoint inhibitor therapy.

摘要

背景

Epstein-Barr 病毒（EBV）与约 10%的所有胃癌有关。然而，EBV miRNAs 在胃癌发生中的分子机制和作用仍有待阐明。

方法

进行 microRNA 微阵列和 TaqMan 定量实时 RT-PCR。进行 PIAS3 的 RT-PCR 和荧光素酶报告基因检测、20 种蛋白质的 Western blot 分析、STAT3 的免疫荧光、miRBART5-5p-质粒、STAT3-质粒、miRBART5-5p 模拟物或 PIAS3-siRNA 的转染以及 PD-L1 的体外生物效应检测。对胃癌组织进行 EBV 编码小 RNA 的原位杂交和免疫组化分析。

结果

将 miR-BART5-5p 转染到 EBV(-)胃癌细胞系中，导致 PIAS3 3'-UTR 报告活性、PIAS3 下调以及随后的 STAT3 激活降低，进而导致 PIAS3/pSTAT3 依赖性 PD-L1 上调。有趣的是，由于 PD-L1 的下调，凋亡增加，而 miR-BART5-5p 转染细胞的增殖、侵袭能力和迁移率降低。在 EBV(+)胃癌细胞中，抗 miR-BART5-5p 通过 PIAS3/pSTAT3 控制降低 PD-L1 水平。在 103 例 EBV 相关胃癌患者中，与 PD-L1(-)肿瘤患者相比，PD-L1(+)肿瘤患者的总生存期明显缩短（P=0.049）。

结论

我们的研究结果表明，miR-BART5-5p 通过 miR-BART5/PIAS3/pSTAT3/PD-L1 轴的直接靶向 PIAS3 并增强 PD-L1。这有助于抗凋亡、肿瘤细胞增殖、侵袭和迁移以及免疫逃避，进一步促进胃癌的进展并恶化临床结局，尤其是在 EBV 相关胃癌的 PD-L1(+)患者中。miR-BART5-5p 可能适合 PD-1/PD-L1 免疫检查点抑制剂治疗。