基于TCGA数据库分析、筛选并验证前列腺癌诊断或预后标志物

目的:综合分析表达谱数据与DNA甲基化数据，探索新的前列腺癌诊断或预后的标志物。方法:在TCGA公共数据库下载前列腺癌的甲基化水平数据以及mRNA表达数据；利用R软件中的DESeq2包对表达谱数据进行差异表达分析，利用R软件中的limma包对DNA甲基化芯片进行差异甲基化位点分析；采用美国国立卫生研究院的DAVID v6.7在线软件平台进行功能和通路的富集分析；对筛选出的基因进行Kaplan-Meier生存分析以及分子生物学实验验证。结果:差异表达分析得到1 130个差异表达基因，其中759个表达下调，371个表达上调；差异甲基化位点分析获得375个差异甲基化位点，其中8个低甲基化位点，367个高甲基化位点；综合分析发现CBX5、NBPF13P、RARB、RCN1、SRSF5基因的异常甲基化与肿瘤复发风险密切相关；定量PCR显示前列腺癌患者SRSF5 mRNA水平升高（P<0.05），甲基化特异性PCR显示前列腺癌患者SRSF5甲基化水平降低（均P<0.05）。结论:SRSF5在前列腺癌患者中甲基化水平降低且mRNA表达水平升高，且其低甲基化水平与复发风险密切相关。

Analysis,screening and validation of diagnostic or prognostic markers for prostate cancer based on TCGA database

Objective: To comprehensively analyze the expression profile data and DNA methylation data in order to explore new molecular targets or potential therapeutic targets for prostate cancer. Methods: The methylation data and mRNA expression data of prostate cancer from TCGA public database were downloaded. Differential expression analysis was performed using DESeq2 package in R software and differential methylation site analysis was performed using limma package in R software. Then，enrichment analysis of functions and pathways were performed using the DAVID v6.7 online software platform. Finally，Kaplan-Meier survival analysis and molecular biological experiments were performed on the selected genes. Results: Differential expression analysis revealed 1 130 differentially expressed genes，of which 759 were down-regulated and 371 were up-regulated. Differential methylation sites analysis obtained 375 differential methylation sites，including 8 hypomethylation sites and 367 hypermethylation sites. Comprehensive analysis found that abnormal methylation of CBX5，NBPF13P，RARB，RCN1，and SRSF5 genes were closely related to the risk of recurrence. Quantitative PCR showed that SRSF5 mRNA were increased（all P<0.05）， and methylation-specific PCR successfully showed that the methylation was decreaed in patients with prostate cancer. Conclusion: The methylation level of SRSF5 is decreased and mRNA expression level is increased in patients with prostate cancer，and its low methylation level is closely related to the risk of recurrence.