顺铂耐药相关基因的筛选、验证及生物信息学分析
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摘要
目的筛选并验证卵巢上皮癌和肺腺癌顺铂耐药的共同差异基因并进行生物信息学分析。方法从基因表达数据库GEO中下载卵巢上皮癌和肺腺癌顺铂耐药芯片数据GSE33482和GSE108214,用R语言软件筛选差异基因并分析得到共同差异基因。运用Metascape进行富集,运用STRING构建蛋白互作网络并使用Cytoscape筛选关键基因。运用实时荧光定量PCR(qRT-PCR)验证关键基因的mRNA表达水平。运用Kaplan-Meier Plotter绘制关键基因的生存曲线。结果筛选得到295个表达上调和176个表达下调的共同差异基因。共同差异基因主要富集于细胞损伤修复、辅助性T淋巴细胞17(Th17)细胞分化、磷脂酰肌醇-3-激酶/丝苏氨酸蛋白激酶(PI3K-Akt)信号通路、细胞黏附、轴突导向和细胞因子间受体相互作用。蛋白互作网络筛选得到10个关键差异基因,qRT-PCR结果显示除HSPB1外其余9个关键基因表达情况与芯片结果一致。生存曲线显示GNG4和PRKCA基因的高表达导致卵巢癌和肺腺癌患者的生存率均明显降低。结论GNG4和PRKCA基因的表达与卵巢上皮癌及肺腺癌细胞顺铂耐药及患者的总生存率均密切相关,有望成为逆转顺铂耐药和改善患者预后的生物学新靶标。
Objective To screen out and verify the common differentially expressed genes related to cisplatin resistance in epithelial ovarian cancer cell lines and lung adenocarcinoma cell lines by bioinformatics analysis.Methods Two different gene expression profiles including the cisplatin-resistant cells and their parental sensitive cell lines of epithelial ovarian cancer(GSE33482)and lung adenocarcinoma cell lines(GSE108214)were obtained from Gene Expression Omnibus(GEO).R language software was used to screen the differentially expressed genes and analyze the common differentially expressed genes.Metascape was used for enrichment.STRING was used to construct the protein interaction network.Cytoscape was used to screen the key genes.Levels of mRNA expression of key genes were verified by qRT-PCR.Kaplan-meier Plotter was used to plot the survival curve of key genes.Results After intersection,295 up-regulated and 176 down-regulated common differentially expressed genes were selected.These common differentially expressed genes were mainly enriched in response to wounding,T helper lymphocytes cell differentiation,phosphatidylinositol 3-kinases-serine/threorine protein kinase signaling pathway,cell adhesion,axon guidance and cytokine-cytokine receptor interaction based on enrichment analysis and qRT-PCR showed that the mRNA expression level of these 10 key differentially genes except for HSPB1 were consistent with the gene expression profiles.KaplanMeier survival curve analysis revealed that high expression of GNG4 and PRKCA resulted in a significant decrease in overall survival in patients with epithelial ovarian cancer and lung adenocarcinoma cell lines.Conclusion GNG4 and PRKCA played an important role in cisplatin resistance and overall survival of epithelial ovarian cancer and lung adenocarcinoma cell lines patients,which might provide new biological targets for reversing cisplatin resistance and improving the prognosis of patients.
Objective To screen out and verify the common differentially expressed genes related to cisplatin resistance in epithelial ovarian cancer cell lines and lung adenocarcinoma cell lines by bioinformatics analysis.Methods Two different gene expression profiles including the cisplatin-resistant cells and their parental sensitive cell lines of epithelial ovarian cancer(GSE33482)and lung adenocarcinoma cell lines(GSE108214)were obtained from Gene Expression Omnibus(GEO).R language software was used to screen the differentially expressed genes and analyze the common differentially expressed genes.Metascape was used for enrichment.STRING was used to construct the protein interaction network.Cytoscape was used to screen the key genes.Levels of mRNA expression of key genes were verified by qRT-PCR.Kaplan-meier Plotter was used to plot the survival curve of key genes.Results After intersection,295 up-regulated and 176 down-regulated common differentially expressed genes were selected.These common differentially expressed genes were mainly enriched in response to wounding,T helper lymphocytes cell differentiation,phosphatidylinositol 3-kinases-serine/threorine protein kinase signaling pathway,cell adhesion,axon guidance and cytokine-cytokine receptor interaction based on enrichment analysis and qRT-PCR showed that the mRNA expression level of these 10 key differentially genes except for HSPB1 were consistent with the gene expression profiles.KaplanMeier survival curve analysis revealed that high expression of GNG4 and PRKCA resulted in a significant decrease in overall survival in patients with epithelial ovarian cancer and lung adenocarcinoma cell lines.Conclusion GNG4 and PRKCA played an important role in cisplatin resistance and overall survival of epithelial ovarian cancer and lung adenocarcinoma cell lines patients,which might provide new biological targets for reversing cisplatin resistance and improving the prognosis of patients.
引文
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[16]AN H,KWAK S,YOO J,et al.Novel miR-5582-5p functions as a tumor suppressor by inducing apoptosis and cell cycle arrest in cancer cells through direct targeting of GAB1,SHC1,and CDK2[J].Biochim Biophys Acta,2016,1862(10):1926-1937.
[17]ACHARYYA S,OSKARSSON T,VANHARANTA S,et al.A CXCL1paracrine network links cancer chemoresistance and metastasis[J].Cell,2012,150(1):165-178.
[2]CHEN W,SUN K,ZHENG R,et al.Cancer incidence and mortality in China,2014[J].Chin J Cancer Res,2018,30(1):1-12.
[3]MEN W,LI W,ZHAO J,et al.TIFA promotes cell survival and migration in lung adenocarcinoma[J].Cell Physiol Biochem,2018,47(5):2097-2108.
[4]THIBAULT B,CASTELLS M,DELORD J,et al.Ovarian cancer microenvironment:implications for cancer dissemination and chemoresistance acquisition[J].Cancer Metastasis Rev,2014,33(1):17-39.
[5]毛楠,何关生,饶进军,等.沉默Bmi-1表达可逆转肺癌细胞对顺铂的耐药性[J].南方医科大学学报,2014,34(7):1000-1004.
[6]GALLUZZI L,SENOVILLA L,VITALE I,et al.Molecular mechanisms of cisplatin resistance[J].Oncogene,2012,31(15):1869-1883.
[7]郑积富,陈丽娜,华建媛,等.TCF3-PBX1阳性急性淋巴细胞白血病相关基因的筛选及生物信息学分析[J].重庆医学,2017,46(3):376-379.
[8]HUANG X,MCGANN J C,LIU B Y,et al.Phosphorylation of dishevelled by protein kinase RIPK4regulates Wnt signaling[J].Science,2013,339(6126):1441-1445.
[9]LOPEZ-BERGAMI P,LAU E,RONAI Z.Emerging roles of ATF2and the dynamic AP1network in cancer[J].Nat Rev Cancer,2010,10(1):65-76.
[10]FU Q,SONG X,LIU Z,et al.miRomics and proteomics reveal a miR-296-3p/PRKCA/FAK/Ras/c-Myc feedback loop modulated by HDGF/DDX5/beta-catenin complex in lung adenocarcinoma[J].Clin Cancer Res,2017,23(20):6336-6350.
[11]DAS R,BHATTACHARYA K,SAMANTA S K,et al.Improved chemosensitivity in cervical cancer to cisplatin:synergistic activity of mahanine through STAT3inhibition[J].Cancer Lett,2014,351(1):81-90.
[12]TRACZ-GASZEWSKA Z,KLIMCZAK M,BIECEK P,et al.Molecular chaperones in the acquisition of cancer cell chemoresistance with mutated TP53and MDM2up-regulation[J].Oncotarget,2017,8(47):82123-82143.
[13]SENGELAUB C A,NAVRAZHINA K,ROSS J B,et al.PTPRN2and PLCβ1 promote metastatic breast cancer cell migration through PI(4,5)P2-dependent actin remodeling[J].EMBO J,2016,35(1):62-76.
[14]PAL J,PATIL V,MONDAL B,et al.Epigenetically silenced GNG4inhibits SDF1α/CXCR4signaling in mesenchymal glioblastoma[J].Genes Cancer,2016,7(3):136-147.
[15]WACLAW B,BOZIC I,PITTMAN M E,et al.A spatial model predicts that dispersal and cell turnover limit intratumour heterogeneity[J].Nature,2015,525(7568):261-264.
[16]AN H,KWAK S,YOO J,et al.Novel miR-5582-5p functions as a tumor suppressor by inducing apoptosis and cell cycle arrest in cancer cells through direct targeting of GAB1,SHC1,and CDK2[J].Biochim Biophys Acta,2016,1862(10):1926-1937.
[17]ACHARYYA S,OSKARSSON T,VANHARANTA S,et al.A CXCL1paracrine network links cancer chemoresistance and metastasis[J].Cell,2012,150(1):165-178.