核糖体蛋白L4基因表达水平与结直肠癌预后相关性

Correlation between the gene expression of ribosome protein L4 and the prognosis of colorectal cancer

目的探讨核糖体蛋白L4(RPL4)在结直肠癌患者中的预后价值及生物学意义。方法从癌症基因组图谱(TCGA)数据库下载542例结直肠癌患者的转录组数据和417例具有完整的总生存期和生存状态资料患者的临床数据。经limma差异基因分析,比较RPL4在结直肠癌组织和正常组织的表达情况,并采用Kaplan-Meier生存分析,评估RPL4表达与结直肠癌患者预后的关系。采用TISCH数据库在单细胞测序水平上验证RPL4在肿瘤微环境中表达的主要细胞类型。经GSEA富集分析探究RPL4表达水平上调相关的肿瘤特征情况。采用Cell Miner数据库中癌NCI-60细胞系抗癌药物测试数据,探讨与RPL4基因相关药物的药物敏感性分析。采用HPA数据库和实时荧光定量聚合酶链反应(PCR)实验分别对组织中RPL4蛋白和mRNA的表达水平进行验证。结果在TCGA结直肠癌的转录组数据中,结直肠癌组织样本中RPL4的表达水平明显高于正常组织,差异有统计学意义(P<0.001)。经患者配对差异分析发现,结直肠癌组织样本中RPL4的表达水平大多高于正常组织。经Kaplan-Meier生存分析发现,RPL4低表达组的总生存期明显长于高表达组,差异有统计学意义(P<0.05)。在单细胞测序水平分析中,RPL4主要在恶性细胞和大多数免疫细胞高表达,而在上皮细胞几乎不表达。经GSEA富集分析发现,在Hallmark基因集中,主要富集于MYC和氧化磷酸化通路;在KEGG基因集中,主要富集于细胞周期和DNA复制以及氨基酸代谢相关通路,这些通路主要参与调控肿瘤的增殖与分化。药物敏感性分析发现,贝利司他、帕博西尼和小分子XR-11576等8种药物的IC50值在RPL4高表达组中显著降低,差异有统计学意义(P<0.05)。在HPA数据库中,RPL4在结直肠癌组织的蛋白表达水平明显高于正常组织。经实时荧光定量PCR实验验证,在结直肠癌组织中,RPL4的mRNA表达水平明显高于正常组织,差异有统计学意义(P<0.05)。结论核糖体基因RPL4有潜力成为用于结直肠癌诊断和预后评估的新型生物学标志物。

Objective To investigate the prognostic value and biological significance of ribosomal protein L4(RPL4)in patients with colorectal cancer.Methods Transcriptome data for 542 colorectal cancer patients and clinical data for 417 patients with complete overall survival and survival status were downloaded from the cancer genome atlas(TCGA)database.Limma differential gene analysis was used to compare the expression of RPL4 in colorectal cancer tissues and normal tissues,and Kaplan-Meier survival analysis was used to evaluate the relationship between RPL4 expression and prognosis of colorectal cancer patients.TISCH database was used to verify the main cell types of RPL4 expression in the tumor microenvironment at the level of single cell sequencing.GSEA enrichment analysis was used to explore the tumor characteristics related to the up-regulation of RPL4 expression.Based on the test data of NCI-60 cell line anticancer drugs in Cell Miner database,the drug sensitivity analysis of drugs related to RPLA gene was discussed.HPA database and real-time fluorescence quantitative polymerase chain reaction(PCR)were used to verify the expression levels of RPL4 protein and mRNA in tissues,respectively.Results In the transcriptome data of TCGA colorectal cancer,the expression leyel of RPL4 in colorectal cancer tissue samples was significantly higher than that in normal tissues,with statistical significance(P<0.001).The expression level of RPL4 in colorectal cancer tissue samples was higher than that in normal tissues.Kaplan-Meier survival analysis showed that the overall survival of low RPL4 expression group was significantly longer than that of high RPL4 expression group,and the difference was statistically significant(P<0.05).In single-cell sequencing level analysis,RPL4 was highly expressed mainly in malignant cells and most immune cells,but hardly expressed in epithelial cells.GSEA enrichment analysis showed that the Hallmark gene concentration was mainly concentrated in MYC and oxidative phosphorylation pathway.KECG gene concentration was mainly concentrated in cell cycle,DNA replication and amino acid metabolism related pathways,which were mainly involved in the regulation of tumor proliferation and diferentiation.Drug sensitivity analysis showed that IC50 values of Bellistat,Palbocinib and small molecule XR-11576 were significantly decreased in RPL4 high expression group,the difference was statistically significant(P<0.05).In the HPA database,RPL4 protein levels were significantly higher in colorectal cancer tissue than in normal tissue.Real-time fluorescence quantitative PCR test confirmed that the mRNA expression level of RPL4 in colorectal cancer tissues was significantly higher than that in normal tissues,and the difference was statistically significant(P<0.05).Conclusion The RPL4 gene has the potential to become a new biological marker for the diagnosis and prognosis evaluation of colorectal cancer.