通过microRNA-mRNA网络分析结合免疫相关基因筛选鉴定鼻咽癌预后生物标志物

文献信息

中文摘要

背景： 鼻咽癌（NPC）是一种常见的头颈部癌症，放疗是其主要的治疗手段。复发和转移是治疗失败的主要原因。尽管免疫治疗可以延长患者的生存期，但NPC发生和进展的分子机制仍不清楚。微小RNA（miRNA）及其靶基因在许多肿瘤中异常表达，并与肿瘤发展密切相关。因此，本研究旨在鉴定NPC中的关键miRNA和免疫相关靶基因，构建miRNA-mRNA调控网络，并筛选和验证核心免疫基因，以识别NPC的预后分子生物标志物。
方法： 我们从基因表达综合数据库（GEO）下载了NPC相关的miRNA（GSE32960）和mRNA（GSE12452、GSE13597、GSE64634）微阵列数据集，并使用R鉴定差异表达miRNA（DEM）和mRNA（DEG）。使用TargetScan、miRDB和miRWalk预测DEM的靶基因，然后与DEG取交集以鉴定核心基因。在Cytoscape中构建miRNA-mRNA调控网络。将核心基因进一步与免疫相关基因集取交集，随后进行功能富集分析。使用LASSO-Cox回归建立预后风险模型，并使用CIBERSORT评估免疫细胞浸润。还进行了模型验证、免疫组化分析和药物-基因相互作用分析。在体外，于C666-1和NP69细胞系中进行RT-qPCR以验证关键基因的表达。
结果： 我们鉴定了关键miRNA的213个潜在靶基因。三个miRNA（hsa-miR-30e-5p、hsa-miR-30a-5p和hsa-miR-20a-5p）与NPC预后相关，17个核心免疫相关基因富集于细胞因子介导的信号传导、趋化因子反应和JAK-STAT通路。五个基因（CXCL11、SOCS1、ROBO1、LGR5和RORB）对NPC显示出有前景的诊断价值；其表达与CD8⁺T细胞、巨噬细胞和中性粒细胞浸润呈正相关（P < 0.05）。药物-基因分析预测5-氟尿嘧啶靶向LGR5，阿地白介素靶向SOCS1。RT-qPCR证明SOCS1在NPC细胞中上调约3.7倍（P < 0.01）。
结论： 本研究鉴定了鼻咽癌中潜在的新关键基因，为该疾病的治疗提供了新的治疗靶点。

英文摘要

BACKGROUND: Nasopharyngeal carcinoma (NPC) is a common head and neck cancer, and radiotherapy is its main treatment. Recurrence and metastasis are the main causes of treatment failure. Although immunotherapy can prolong patient survival, the molecular mechanisms underlying NPC development and progression remain unclear. MicroRNAs (miRNAs) and their target genes are abnormally expressed in many tumors and are closely involved in tumor development. Therefore, this study aimed to identify key miRNAs and immune-related target genes in NPC, construct miRNA-mRNA regulatory networks, and screen for and validate core immune genes to identify prognostic molecular biomarkers for NPC.
METHODS: We downloaded NPC-related miRNA (GSE32960) and mRNA (GSE12452, GSE13597, GSE64634) microarray datasets from the Gene Expression Omnibus (GEO) and identified differentially expressed miRNAs (DEMs) and mRNAs (DEGs) using R. Target genes of DEMs were predicted with TargetScan, miRDB, and miRWalk, and then intersected with DEGs to identify core genes. A miRNA-mRNA regulatory network was constructed in Cytoscape. The core genes were further overlapped with immune-related gene sets, followed by functional enrichment analysis. A prognostic risk model was established using LASSO-Cox regression, and immune cell infiltration was evaluated with CIBERSORT. Model validation, immunohistochemical analysis, and drug-gene interaction analysis were also performed. In vitro, RT-qPCR was conducted in C666-1 and NP69 cell lines to verify the expression of key genes.
RESULTS: We identified 213 potential target genes of key miRNAs. Three miRNAs (hsa-miR-30e-5p, hsa-miR-30a-5p, and hsa-miR-20a-5p) were associated with NPC prognosis, and 17 core immune-related genes were enriched in cytokine-mediated signaling, chemokine response, and JAK-STAT pathways. Five genes (CXCL11, SOCS1, ROBO1, LGR5, and RORB) showed promising diagnostic value for NPC; their expression was positively correlated (P < 0.05) with CD8 ⁺ T-cell, macrophage, and neutrophil infiltration. Drug-gene analysis predicted that 5-fluorouracil targeted LGR5 and aldesleukin targeted SOCS1. RT-qPCR demonstrated that SOCS1 was upregulated approximately 3.7-fold in NPC cells (P < 0.01).
CONCLUSION: This study identified potential novel key genes in nasopharyngeal carcinoma, providing new therapeutic targets for the treatment of this disease.