摘要目的 筛选乳腺癌放射治疗(放疗)抵抗相关微小RNA(miRNAs),为乳腺癌患者放疗抵抗的基础研究与临床应用研究提供实验基础.方法 从基因表达综合数据库(GEO)中下载乳腺癌放疗患者相关miRNAs微阵列数据集GSE 107743,利用GEO2R分析工具筛选放疗后局部复发患者的差异表达miRNAs,mirDIP数据库预测差异表达miRNAs的靶基因,DAVID数据库对靶基因分别进行基因本体论(GO)富集分析和京都基因与基因组百科全书(KEGG)通路富集分析.最后采用实时荧光定量PCR在人乳腺癌细胞MCF-7中进行差异表达验证.结果 采用GEO2R分析工具筛选出9种与放疗抵抗相关的差异表达miRNAs,其中3种miRNAs(hsa-miR-600、hsa-miR-525-3p、hsa-miR-591)表达上调,6种miRNAs(hsa-miR-488-5p、hsa-miR-582-3p、hsa-miR-520h、hsa-miR-488-3p、hsa-miR-744-3p、hsa-miR-103b)表达下调.靶基因预测结果显示,9种差异表达miRNAs的潜在靶基因共134个.靶基因显著富集在细胞周期、细胞凋亡、干细胞分化等相关生物学过程(均P＜0.05)以及转化生长因子-β、磷脂酰肌醇3-激酶\蛋白激酶B信号通路等信号通路中(均P＜0.05).实时荧光定量PCR检测结果表明,6种miRNAs (hsa-miR-600、hsa-miR-525-3p、hsa-miR-591、hsa-miR488-5p、hsa-miR-582-3p、hsa-miR-520h)在经5 Gy 137Csγ射线辐照后的MCF-7细胞中表达差异改变与GEO2R分析结果完全一致.结论 利用生物信息学从乳腺癌放疗患者局部复发临床样本中筛选出的差异表达miRNAs可能与乳腺癌患者的放疗抵抗密切相关,有望成为放疗抵抗治疗的新靶点.

abstractsObjective To screen radiotherapy resistance related microRNAs (miRNAs) in breast cancer and provide experimental basis for basic researches and clinical solutions of radiotherapy resistance in breast cancer patients.Methods The miRNA microarray dataset GSE107743 related to breast cancer radiotherapy patients was downloaded from the Gene Expression Omnibus (GEO).The GEO2R analysis tool was used to screen differentially expressed miRNAs in patients with local recurrence after radiotherapy.The target genes of differentially expressed miRNAs were predicted by the mirDIP database.GO enrichment analysis and KEGG pathway analysis on the target genes were performed by DAVID dataset.Finally,differential expression verification was performed in human breast cancer cell line MCF-7 by real-time fluorescent quantitative PCR.Results A total of 9 differentially expressed miRNAs related to radiotherapy resistance were screened by the GEO2R analysis tool,in which three miRNAs (hsa-miR-600,hsa-miR-525-3p and hsa-miR-591) were up-regulated and 6 miRNAs (hsa-miR-488-5p,hsa-miR-582-3p,hsa-miR-520h,hsa-miR-488-3p,hsa-miR-744-3p and hsa-miR-103b) were down-regulated.Target gene prediction results showed that there were 134 potential target genes in these nine differentially expressed miRNAs.These target genes were significantly enriched in related biological processes such as apoptosis and stem cell differentiation (all P＜0.05) and signal transduction pathways such as transforming growth factor-β and phosphatidylinositol 3-kinase-protein kinase B signaling pathway (all P＜0.05).The results of real-time PCR showed that the differential expression of six miRNAs,i.e.hsa-miR-600,hsa-miR-525-3p,hsa-miR-591,hsa-miR488-5p,hsa-miR-582-3p and hsa-miR-520h,was detected in the MCF-7 cells irradiated by 5 Gy 137Cs γ-rays,and this result was consistent with the results of GEO2R analysis.Conclusion The differentially expressed miRNAs screened from clinical samples of breast cancer patients with local recurrence using bioinformatics may be closely associated with the radiotherapy resistance of these patients.These miRNAs are expected to become new biomarkers for the therapy of radiotherapy resistance.