RFC5在子宫颈癌中的表达及其对预后、免疫调控的影响和相关机制

Expression of RFC5 in cervical cancer and its effect on the prognosis and immune regulation as well as its related mechanism

目的探讨复制因子C5(RFC5)在子宫颈癌中的表达及其对预后、免疫调控的影响和相关机制。方法2023年5月从癌症基因组图谱(TCGA)数据库下载280例子宫颈癌患者RFC5 mRNA表达数据及临床数据。比较子宫颈癌组织和癌旁正常组织中RFC5 mRNA表达水平差异,分析不同临床病理特征患者RFC5 mRNA表达水平;根据子宫颈癌组织RFC5 mRNA中位表达水平将280例患者分为高表达组和低表达组,采用Kaplan-Meier法对两组进行生存分析,比较采用log-rank检验;利用基因表达谱交互分析(GEPIA)2在线工具验证子宫颈癌RFC5基因表达情况及与预后的关系。采用单因素、多因素Cox比例风险模型分析TCGA数据库子宫颈癌患者总生存(OS)的影响因素。利用LinkedOmics数据库筛选子宫颈癌中RFC5相关的共同差异表达基因(DEG),基于DAVID数据库对RFC5相关DEG进行基因本体(GO)分析和京都基因与基因组百科全书(KEGG)通路富集分析。基于肿瘤免疫评估资源(TIMER)数据库,通过Spearman法分析RFC5表达与子宫颈癌肿瘤浸润免疫细胞的关系;基于肿瘤免疫系统相互作用数据库(TISIDB)分析子宫颈癌RFC5表达与免疫调节因子的关系。基于人类蛋白图谱(HPA)数据库分析子宫颈癌组织中RFC5蛋白表达情况。基于GEPIA2在线工具分析RFC5在泛癌中的表达及其与OS的关系。结果TCGA数据库中RFC5 mRNA在子宫颈癌组织(277例)中相对表达水平高于癌旁正常组织(3例),差异有统计学意义(P<0.001);不同年龄、病理类型、临床分期子宫颈癌患者间癌组织中RFC5 mRNA相对表达水平差异均无统计学意义(均P>0.05);RFC5高表达组(139例)子宫颈癌患者OS优于RFC5低表达患者(138例),差异有统计学意义(P=0.027)。GEPIA工具验证RFC5在子宫颈癌中的表达及其与OS的关系,得到相同结果。GO和KEGG富集分析显示,RFC5相关基因主要参与DNA复制、细胞周期、范可尼贫血、错配修复、碱基切除修复、核苷酸切除修复等信号通路。多因素Cox回归分析显示,年龄≥65岁、临床分期Ⅳ期、RFC5表达低水平是子宫颈癌患者OS不良的独立危险因素(均P<0.05)。TIMER数据库分析显示,RFC5的表达与肿瘤纯度呈正相关(rho=0.198,P<0.001),而与B细胞(rho=0.062,P=0.306)、CD8+T细胞(rho=0.168,P=0.005)、CD4+T细胞(rho=-0.049,P=0.418)、巨噬细胞(rho=0.034,P=0.577)、中性粒细胞(rho=0.169,P=0.005)、骨髓树突细胞(rho=0.026,P=0.667)的浸润水平呈弱相关性或无相关性。对TISIDB数据分析显示,RFC5表达与免疫抑制因子和免疫刺激因子大多呈负相关性。HPA数据库分析显示,子宫颈癌组织RFC5蛋白的表达水平高于正常子宫颈组织。GEPIA在线工具分析显示,RFC5在多种恶性肿瘤中表达上调,RFC5高表达胸腺瘤患者OS优于其低表达患者,而RFC5高表达急性髓系白血病患者OS较其低表达患者差,差异均有统计学意义(均P<0.05)。结论RFC5在子宫颈癌组织中高表达,且其水平与子宫颈癌患者预后相关。RFC5过表达可能抑制子宫颈癌免疫调节因子的表达,可能是通过DNA复制、细胞周期、错配修复、碱基切除修复等相关通路调控子宫颈癌的发生与发展的。

ObjectiveTo investigate the expression of replication factor C5(RFC5)in cervical cancer,and its effect on the prognosis and immune regulation as well as its related mechanism.MethodsRFC5 mRNA expression data and the clinical data of 280 cervical cancer patients were downloaded from the Cancer Genome Atlas(TCGA)database in May 2023.The difference of RFC5 mRNA expression between cervical cancer tissues and paracancerous normal tissues was compared and the expression of RFC5 mRNA in patients with different clinicopathological characteristics was analyzed.According to the median expression level of RFC5 mRNA in cancer tissues,280 patients were divided into the high expression group and the low expression group.Kaplan-Meier method was used for survival analysis,and log-rank test was used for comparison.Gene Expression Profiling Interactive Analysis(GEPIA)2 online tool was used to verify the relationship between RFC5 gene expression and the prognosis of cervical cancer.Univariate and multivariate Cox proportional risk models were used to analyze the factors influencing the overall survival(OS)of patients with cervival cancer in TCGA database.LinkedOmics database was used to screen the co-differentially expressed genes(DEG)related to RFC5 in cervical cancer.Gene Ontology(GO)and Kyoto Encyclopedia of Genesand Genomes(KEGG)pathway enrichment analysis of DEG related to RFC5 were performed based on the DAVID database.Based on the Tumor Immune Estimation Resource(TIMER)database,the relationship between RFC5 expression and tumor infiltrating immune cells in cervival cancer was analyzed by using Spearman method.The relationship between RFC5 expression and immunomodulatory factors in cervical cancer was analyzed based on tumor-immune system interactions database(TISIDB).The expression of RFC5 protein in cervical cancer tissues was analyzed based on Human Protein Atlas(HPA)database.The expression of RFC5 in pan-cancer and its correlation with OS were analyzed based on GEPIA2 online tool.ResultsIn TCGA database,the relative expression level of RFC5 mRNA in cervical cancer tissues(277 cases)was higher than that in paracancerous normal tissues(3 cases),and the difference was statistically significant(P<0.001),while there were no significant differences in the relative expression level of RFC5 mRNA in cancer tissues of patients with different age,pathological type and clinical staging(all P>0.05).The OS of cervival cancer patients in high RFC5 expression group(139 cases)was better than that of patients in low RFC5 expression group(138 cases),and the difference was statistically significant(P=0.027).GEPIA tool verification indicated that expression of RFC5 in cervical cancer and its relationship with OS showed the same results.GO and KEGG enrichment analysis showed that RFC5-related genes were mainly involved in DNA replication,cell cycle,Fanconi anemia,mismatch repair,base excision repair,nucleotide excision repair and other signaling pathways.Multivariate Cox regression analysis showed that age≥65 years,clinical stagingⅣand the low expression of RFC5 were independent risk factors of poor OS in patients with cervical cancer(all P<0.05).TIMER database analysis showed that the expression of RFC5 was positively correlated with tumor purity(rho=0.198,P<0.001),while weakly correlated or not correlated with the infiltration levels of B cells(rho=0.062,P=0.306),CD8+T cells(rho=0.168,P=0.005),CD4+T cells(rho=-0.049,P=0.418),macrophages(rho=0.034,P=0.577),neutrophils(rho=0.169,P=0.005)and bone marrow dendritic cells(rho=0.026,P=0.667).Analysis of TISIDB data showed that RFC5 expression was mostly negatively correlated with immunosuppressive factors and immunostimulatory factors.HPA database showed that the expression level of RFC5 in cervical cancer tissues was higher than that in normal cervical tissues.GEPIA online tool database analysis showed that RFC5 expression was up-regulated in a variety of malignant tumors.The OS of thymoma patients with high RFC5 expression was better than that of those with low RFC5 expression,while the OS of acute myeloid leukemia patients with high RFC5 expression was worse than those with low RFC5 expression,and the differences in OS were statistically significant(both P<0.05).ConclusionsRFC5 is highly expressed in cervical cancer and its expression is associated with prognosis of patients with cervival cancer.Overexpression of RFC5 may inhibit the expression of immunomodulatory factors,and it may regulate the development and progression of cervical cancer through DNA replication,cell cycle,mismatch repair,base excision repair and other related pathways.