口腔鳞状细胞癌患者预后相关基因标志物的生物信息学分析

doi:10.12016/j.issn.2096-1456.2021.01.004

摘要/Abstract

摘要：

目的通过生物信息分析方法筛选口腔鳞状细胞癌与癌旁组织之间的差异表达基因,初步确定潜在生物标志物。方法从基因表达数据库（Gene Expression Omnibus,GEO）中下载口腔鳞状细胞癌相关芯片数据集GSE23558、GSE37991、GSE30784,利用GEO在线分析工具筛选差异表达基因,并对其进行基因本体、京都基因与基因组百科全书分析。利用STRING在线数据库,构建差异基因蛋白互作（protein-protein interaction,PPI）网络,并通过Cytoscape筛选关键基因。进而使用Kaplan Meier在线软件对差异表达基因进行预后分析,并使用GEPIA（http://gepia.cancer-pku.cn/）验证其表达情况。结果共筛选出212个差异表达基因,进一步分析发现16个核心基因,其中与预后相关的核心基因包括:极光激酶A（aurora kinase A,AURKA）、极光激酶B（aurora kinase B,AURKB）、细胞凋亡抑制因子5（baculoviral IAP repeat containing 5,BIRC5）、细胞分裂周期蛋白6（cell division cycle 6,CDC6）、E2F转录因子7（E2F transcription factor 7,E2F7）、泛素样含PHD和环指域蛋白1（ubiquitin-like with PHD and ring finger domains 1,UHRF1）,这些关键基因与癌旁组织相比在口腔鳞状细胞癌组织呈高表达并且患者生存时间均较短,差异均具有统计学意义（P < 0.05）。结论 AURKA、AURKB、BIRC5、CDC6、E2F7和UHRF1可作为口腔鳞状细胞癌患者预后潜在的标志物。

关键词: 口腔鳞状细胞癌, 生物信息学, 生物学标志物, 差异表达基因, 基因芯片, 极光激酶, 细胞凋亡抑制因子5, 细胞分裂周期蛋白6, E2F转录因子7, 泛素样含PHD和环指域蛋白1

Abstract:

Objective To analyze the differentially expressed genes of patients with oral squamous cell carcinoma (OSCC) from paracarcinoma through biological information analysis to preliminarily identify OSCC-associated genes. Methods GSE23558, GSE37991 and GSE30784 were downloaded from the Gene Expression Omnibus (GEO), which is the mRNA expression profile dataset. The differentially expressed genes (DEGs) were identified based on the gene ontology and the Kyoto Encyclopedia of Genes and Genomes. Then, the protein-protein interaction (PPI) network was constructed using the STRING online tool, and Cytoscape was used to filter the critical genes. Furthermore, key genes involved in the survival of patients with OSCC were analyzed using Kaplan-Meier analysis. The expression of hub genes was validated based on GEPIA（http://gepia.cancer-pku.cn/）. Results A total of 212 DEGs were screened, and further analysis revealed 16 core genes, among which the core genes associated with prognosis included aurora kinase A (AURKA), aurora kinase B (AURKB), apoptosis inhibiting factor 5 (BIRC5), cell division cycle 6 (CDC6), E2F transcription factor 7 (E2F7), ubiquitin-like with PHD and ring finger domains 1 (UHRF1). These key genes were highly expressed in patients with oral squamous cell carcinoma, and the survival time of patients was short; the difference was statistically significant (P < 0.05). Conclusion AURKA, AURKB, BIRC5, CDC6, E2F7 and UHRF1 may be useful as potential biomarkers for OSCC prognosis prediction.

Key words: oral squamous cell carcinoma, bioinformatics, biomarker, differentially expressed genes, microarray, aurora kinase, baculoviral IAP repeat containing 5, cell division cycle 6, E2F transcription factor 7, ubiquitin-like with PHD and ring finger domains 1

中图分类号:

R78
R739.81

王力业,高莺,田淳. 口腔鳞状细胞癌患者预后相关基因标志物的生物信息学分析[J]. 口腔疾病防治, 2021, 29(1): 27-33.

WANG Liye,GAO Ying,TIAN Chun. Identification of potential genetic markers in prognosis of oral squamous cell carcinoma patients by bioinformatic analysis[J]. Journal of Prevention and Treatment for Stomatological Diseases, 2021, 29(1): 27-33.

图/表 6

表1 口腔鳞状细胞癌的差异表达基因

Table 1 Venn diagrams of differentially expressed genes of OSCC

DEGs

Genes Name

Up-regulated

ADAM12 HOXA1 LY6K ITGA3 BIRC5 FOXM1 PCDH7 DENND1A IL24 CDC6 AURKA SH2D2A AURKB SPP1 MELK TMEM132A GTSE1 E2F7 LAMA3 NAA40 MMP1 CD109 BNC1 CDH3 MMP3 LPAR3 CD276 AGRN CYP27B1 GALNT6 DCBLD1 PDE7A ARPC1B MCM2 COL4A6 CXCL10 LAMC2 GBP5 KIAA0101 MMP10 PLAU SLC28A3 HOMER3 ITGB4 FBLIM1 PTHLH TENM2 DSG2 CXCL11 SERINC2 PTK7 SPHK1 TNFRSF12A OASL TRIP13 TLDC1 HMGA2 ISG15 PDPN TK1 SHCBP1 DPP3 SERPINH1 PLEK2 INHBA CDCA3 CXCL9 CXCL5 KIF20A AIM2 UHRF1 SCG5 PPP4R4 WDR66

Down-regulated

MITF DKK2 HNMT GDF10 BDH2 MYO5C KRT222 HLF TLE2 LONRF2 PBX1 VIT SYTL4 IL17D OMD PDGFD BEX2 ARHGEF26 DCT C1orf115 SH3BGRL2 C1QTNF7 NEGR1 RBP7 CAND2 NRXN1 PHYH GPD1L HRASLS CTTNBP2 ANGPTL1 ARHGAP20 NR3C2 SLC27A6 IL17RD OGN GIPC2 TCEAL2 NTRK2 PLCB4 NBEA MIR99AHG APOD F10 PID1 ZNF415 SUSD4 SCN7A PEG3 MYZAP TGFBR3 RORC BOC PGM5 FXYD1 GSTM5 PRELP CKMT2 PLP1 C2orf40 AOX1 GULP1 CFD RNASE4 CYP4B1 PTN ZSCAN18 CHRDL1 CILP SYNPO2 FAM150B RERG SLC16A7 SOD3 CRYAB TSPAN8 CD36 DCLK1 COL21A1 SLC47A1 TM7SF2 FAM149A ABCA8 HSPB2 ASPA MAN1C1 TNMD AMPD1 MYH11 GAMT ABI3BP C8orf22 ANG HPGD NTRK3 BEX4 TMEM100 PLIN1 CNTN3 DPT SORCS1 MFAP4 MGLL SCARA5 MAOB ATP1A2 ACADSB IGSF10 ACKR1 SORBS1 FGF7 LIFR STXBP6 PMP2 LGI1 2-Mar PPM1L PDZRN4 COX7A1 FAM189A2 FMO2 LMO3 ARHGAP6 FAM221A MYRIP CAB39L FAM107A CCDC85A PPP1R3C RRAGD TCEA3 MMRN1 GKAP1 GGTA1P MAMDC2 AR GATM SELENBP1

图1 口腔鳞状细胞癌的差异表达基因的基因本体论分析结果

Figure 1 Gene ontology enrichment analysis results of differentially expressed genes in OSCC Red represents biological processes（BP）, green represents cellular components（CC）, and blue represents molecular function （MF）; OSCC: oral squamous cell carcinoma; GO: gene ontology

图2 口腔鳞状细胞癌的差异表达基因参与的代谢途径

Figure 2 Differentially expressed genes in OSCC are involved in metabolic pathways The X-axis represents the ratio of gene number, and the Y-axis represents each signalling pathway; OSCC: oral squamous cell carcinoma

图3 口腔鳞状细胞癌的差异基因蛋白质互作网络图

Figure 3 Constructed protein-protein interaction network of differentially expressed genes in OSCC Module analysis was performed via Cytoscape software. Finally, 16 core genes were obtained: blue represents downregulated DEGs and red represents upregulated DEGs; OSCC: oral squamous cell carcinoma; MELK: maternal embryonic leucine zipper kinase; GTSE1: G2 and S phase-expressed protein 1; TK1: thymidine kinase 1; KIF20A: kinesin family member 20A; SHCBP1: SHCSH2-binding protein 1; MCM2: minichromosome maintenance protein 2; CDCA3: cell division cycle A3; TRIP13: thyroid hormone receptor interactor 13; FOXM1: Forkhead box protein M1; AURKA: aurora kinase A; AURKB: aurora kinase B; BIRC5: baculoviral IAP repeat containing 5; CDC6: cell division cycle 6; E2F7: E2F transcription factor 7; UHRF1: ubiquitin-like with PHD and ring finger domains 1

图4 口腔鳞状细胞癌核心基因的预后信息

Figure 4 The prognostic information of the core genes in OSCC The core genes had a significantly worse survival rate (P < 0.05); AURKA: aurora kinase A; AURKB: aurora kinase B; CDC6: cell division cycle 6; BIRC5: baculoviral IAP repeat containing 5; E2F7: E2F transcription factor 7; UHRF1: ubiquitin-like with PHD and ring finger domains 1; OSCC: oral squamous cell carcinoma

图5 验证口腔鳞状细胞癌核心基因表达水平

Figure 5 Validation of the expression levels of critical genes in OSCC Six genes related to poor prognosis were analyzed using the GEPIA website; six genes had significant expression levels in OSCC specimens compared to normal specimens （*P < 0.05）; red represents tumor tissues, and gray represents normal tissues; AURKA: aurora kinase A; AURKB: aurora kinase B; CDC6: cell division cycle 6;BIRC5: baculoviral IAP repeat containing 5; E2F7: E2F transcription factor 7; UHRF1: ubiquitin-like with PHD and ring finger domains 1; OSCC: oral squamous cell carcinoma

参考文献 23

[1]	Zhao X, Sun S, Zeng X , et al. Expression profiles analysis identifies a novel three-mRNA signature to predict overall survival in oral squamous cell carcinoma[J]. Am J Cancer Res, 2018,8(3):450-461. URL PMID
[2]	赵思语, 欧阳少波, 王军 , 等. 口腔鳞状细胞癌组织中环状RNA的差异表达谱分析[J]. 口腔疾病防治, 2018,26(2):83-89. doi: 10.12016/j.issn.2096-1456.2018.02.003.
	Zhao SY, Ouyang SB, Wang J , et al. Differential expression of circular RNA in oral squamous cell carcinoma[J]. J Prev Treat Stomatol Dis, 2018,26(2):83-89. doi: 10.12016/j.issn.2096-1456.2018.02.003.
[3]	Deb B, Sengupta P, Sambath J , et al. Bioinformatics analysis of global proteomic and phosphoproteomic data sets revealed activation of NEK2 and AURKA in cancers[J]. Biomolecules, 2020,10(2):237-252. doi: 10.3390/biom10020237. DOI URL
[4]	Zheng JW, Yang Y, Yang S , et al. Gene microarray analysis revealed a potential crucial gene RACK1 in oral squamous cell carcinoma (OSCC)[J]. Anim Cells Syst (Seoul), 2018,22(2):82-91. doi: 10.1080/19768354.2018.1443493. DOI URL
[5]	Zhang X, Feng H, Li D , et al. Identification of differentially expressed genes induced by aberrant methylation in oral squamous cell carcinomas using integrated bioinformatic analysis[J]. International journal of molecular sciences, 2018,19(6):1698-1713. doi: 10.3390/ijms19061698. DOI URL
[6]	Jeyaprakash AA, Klein UR, Lindner D , et al. Structure of a Survivin-Borealin-INCENP core complex reveals how chromosomal passengers travel together[J]. Cell, 2007,131(2):271-285. doi: 10.1016/j.cell.2007.07.045. DOI URL PMID
[7]	Li DY, Hu CH, Li HB . Survivin as a novel target protein for reducing the proliferation of cancer cells[J]. Biomed Rep, 2018,8(5):399-406. doi: 10.3892/br.2018.1077. DOI URL PMID
[8]	Troiano G, Guida A, Aquino G , et al. Integrative histologic and bioinformatics analysis of birc5/survivin expression in oral squamous cell carcinoma[J]. Int J Mol Sci, 2018,19(9):2664-2677. doi: 10.3390/ijms19092664. DOI URL
[9]	Polepalli S, George SM, Vidya RVS , et al. Role of UHRF1 in malignancy and its function as a therapeutic target for molecular docking towards the SRA domain[J]. Int J Biochem Cell Biol, 2019,114:105558. doi: 10.1016/j.biocel.2019.06.006. DOI URL PMID
[10]	Zhou L, Shang Y, Jin Z , et al. UHRF1 promotes proliferation of gastric cancer via mediating tumor suppressor gene hypermethylation[J]. Cancer Biol Ther, 2015,16(8):1241-1251. doi: 10.1080/15384047.2015.1056411. DOI URL PMID
[11]	Baroudi ME, Machiels JP, Schmitz S . Expression of SESN1, UHRF1BP1, and miR-377-3p as prognostic markers in mutated TP53 squamous cell carcinoma of the head and neck[J]. Cancer Biol Ther, 2017,18(10):775-782. doi: 10.1080/15384047.2017. 1373212. DOI URL PMID
[12]	Guo H, Zhang L . MicroRNA-30a suppresses papillary thyroid cancer cell proliferation, migration and invasion by directly targeting E2F7[J]. Exp Ther Med, 2019,18(1):209-215. doi: 10.3892/etm.2019.7532. DOI URL PMID
[13]	Hazar-Rethinam M, Long LM, Gannon MO , et al. RacGAP1 is a novel downstream effector of E2F7-dependent resistance to doxorubicin and is prognostic for overall survival in squamous cell carcinoma[J]. Mol Cancer Ther, 2015,14(8):1939-1950. doi: 10.1158/1535-7163.MCT-15-0076. DOI URL PMID
[14]	Saen-Ponce N, Pillay R, De Long LM , et al. Targeting the XPO1-dependent nuclear export of E2F7 reverses anthracycline resistance in head and neck squamous cell carcinomas[J]. Sci Transl Med, 2018, 10(447): eaar7223. doi: 10.1126/scitranslmed.aar7223. DOI URL PMID
[15]	Lim N, Townsend PA . Cdc6 as a novel target in cancer: oncogenic potential, senescence and subcellular localisation[J]. Int J Cancer, 2020,147(6):1528-1534. doi: 10.1126/scitranslmed.aar7223. DOI URL PMID
[16]	Feng CJ, Lu XW, Luo DY , et al. Knockdown of Cdc6 inhibits proliferation of tongue squamous cell carcinoma Tca8113 cells[J]. Technol Cancer Res Treat, 2013,12(2):173-181. doi: 10.7785/tcrt.2012.500302. DOI URL PMID
[17]	Feng CJ, Li HJ, Li JN , et al. Expression of Mcm7 and Cdc6 in oral squamous cell carcinoma and precancerous lesions[J]. Anticancer Res, 2008,28(6A):3763-3769. URL PMID
[18]	Huang C, Wang L, Song H , et al. Interactive effects of AURKA polymorphisms with smoking on the susceptibility of oral cancer[J]. Artif Cells Nanomed Biotechnol, 2019,47(1):2333-2337. doi: 10.1080/21691401.2019.1601101. DOI URL PMID
[19]	Yang Y, Ding L, Zhou Q , et al. Silencing of AURKA augments the antitumor efficacy of the AURKA inhibitor MLN8237 on neuroblastoma cells[J]. Cancer Cell Int, 2020,20:9. doi: 10.1186/s12935-019-1072-y. DOI URL PMID
[20]	Mehra R, Serebriiskii IG, Burtness B , et al. Aurora kinases in head and neck cancer[J]. Lancet Oncol, 2013,14(10):e425-e435. doi: 10.1016/S1470-2045(13)70128-1.
[21]	Bertran-Alamillo J, Cattan V, Schoumacher M , et al. AURKB as a target in non-small cell lung cancer with acquired resistance to anti-EGFR therapy[J]. Nat Commun, 2019,10(1):1812. doi: 10.1038/s41467-019-09734-5. DOI URL PMID
[22]	Togar T, Desai S, Mishra R , et al. Identifying cancer driver genes from functional genomics screens[J]. Swiss Med Wkly, 2020,150:w20195. doi: 10.4414/smw.2020.20195. DOI URL PMID
[23]	Boeckx C, Op de Beeck K, Wouters A , et al. Overcoming cetuximab resistance in HNSCC: the role of AURKB and DUSP proteins[J]. Cancer Lett, 2014,354(2):365-377. doi: 10.1016/j.canlet.2014.08.039. DOI URL