gene expression profile data.This study included four GEO data sets, including a total of 297 ovarian cancer samples and 32 healthy control samples (Table 2). They were standardized by the limma software package in the R/Bioconductor software (Fig. 1). 812, 2820, 1495 and 536 DEGs were screened out respectively (P < 0.05, |logFC|>1.5). The differentially expressed genes in the sample data of the 4 data sets are shown in Fig. 2 (Fig. 2). Use VennDiagram package to perform gene integration of DEG that meets the standard. In conclusion, compared with normal OV tissue, a total of 135 (33 up-regulated genes and 102 down-regulated genes) in OC tissue samples were identified as DEG (Table 3).
enrichment analysis.The cluster profiler package was used in R software to biologically annotate 33 up-regulated DEGs and 102 down-regulated DEGs after integration, and the GO function enrichment with P-value < 0.05 was obtained. The significant results of GO enrichment analysis showed that: 1. In the cell composition, the up-regulated DEG is mainly enriched in the double-strain tight junction, late promotion complex, apical junction complex, tight junction, and nuclear ubiquitin junction complex. Down-regulated DEG is mainly enriched in the extracellular matrix, collagen-containing extracellular matrix, and blood particles; 2. In biological processes, up-regulated DEG is enriched in mitotic spindle assembly checkpoints, chromosome separation and regulation, and cell cycle The regulation of later transitions, the positive regulation of ubiquitin-protein ligase activity, the involvement of signal transduction in gene expression regulation and chromosome separation, etc. The down-regulated DEG is obviously enriched in the regulation of neurotransmitter levels, the regulation of blood coagulation, protein serine/thereon The regulation of amino acid kinase activity, the process of mucopolysaccharide metabolism, and the Wnt signaling pathway; 3. In the molecular function group, the down-regulated DEG is mainly enriched in heparin-binding and frizzled binding, while the up-regulated DEG is not significantly enriched in compliance with the standard. (Table 4 & Fig. 3)
PPI network and module analysis.The STRING database was used to establish a PPI network, and 152 protein pairs were obtained. The PPI network was constructed after the comprehensive score > 0.4 and the removal of 29 individual nodes (Fig. 4). The gene data was input into Cytoscape software, and a PPI network diagram containing 29 up-regulated DEG and 77 down-regulated DEG was further obtained. MCODE detected a total of 4 modules, and we chose the module with a higher score for the next analysis (Fig. 5). Use the MCC algorithm in Cytohubba to get the top 20 hub genes, which are: KDR, SOX9, EPCAM, WNT5A, FGF13, PDGFRA, CP, ALDH1A1, KLF4, CDC20, UBE2C, FGF9, SOX17, TTK, TRIP13, CKS2, RACGAP1, CD24, CHGB, LAMB1.
gene enrichment through KEGG pathway.In order to understand the functions of the modules, we have performed KEGG enrichment analysis for each module. The results are shown in(Table 5). TRIP13, RACGAP1, CKS2, UBE2C, TTK, and CDC20 in module 1 all up-regulate DEG, which is mainly enriched in the cell cycle and ubiquitin-mediated proteolysis pathways. There are four genes ALDH1A1, CD24, EPCAM, and SOX9 in module 2. Except for ALDH1A1 which is down-regulated DEG, the other 3 genes are all up-regulated DEG. There is no obvious pathway enrichment in this module. There are four genes CP, LAMB1, CHRDL1, and CHGB in module 3. Except for CP which up-regulates DEG, the rest are down-regulated DEG. After enrichment, CP exists in iron death, porphyrin, and chlorophyll metabolism pathways, and LAMB1 is in ECM receptor Interaction, small cell lung cancer, and other pathways exist.
survival analysis and expression level analysis of hub gene.We used the Kaplan Meier Plotter online website to analyze the survival of 20 hub genes and found that 13 genes associated with ovarian cancer have a poor prognosis (P < 0.05, Fig. 6). Then use the GEPIA online database to mine the expression levels of 13 genes between ovarian cancer patients and normal people. The results showed that compared with normal ovarian samples, among the 13 prognostic-related genes in ovarian cancer samples, SOX9, EPCAM, CP, UBE2C, TTK, RACGAP1, and CD24 7 genes reflected high expression (P < 0.01, Fig. 7 ).
clinicopathological characteristics and TTK expression.In this study, the average age of all patients at surgery was 52 years, and the median age was 53 years. Among them, 59.1% of patients with epithelial ovarian cancer have lymph node metastasis. The proportion of middle-high-middle-differentiated cancer was 43.0%. 56.9% of patients had distant metastases. We found that the expression of TTK was negative in normal ovarian tissues. In tumor tissues, all specimens had positive cytoplasmic staining, and the expression of a benign group, borderline group, and malignant group increased in turn. We calculated the H score of TTK expression in tumor tissues. Among them, the H score is 180 (10–220).(Table 6&Figure 8)
analysis of the correlation between TTK expression and clinicopathological factors.We found that there was no significant correlation between TTK expression and age and fertility level. However, there is a significant positive correlation between TTK expression and tumor differentiation, CA125 level, HE4 level, clinical stage, lymphatic metastasis, and distant metastasis. Compared with patients with normal CA125, HE4, moderately well-differentiated, stage I-II, no lymph node metastasis, and no distant metastasis, CA125 elevated, HE4 elevated, poorly differentiated, stage III-IV, lymph node metastasis, and distant metastasis patients, TTK expression rate is higher, multivariate logistic regression analysis with statistically significant clinical-pathological factors in the univariate analysis as independent variables, the results show: distant metastasis, lymph node metastasis, clinical stage (III-IV), Poor differentiation is an independent risk factor for high TTK expression (P < 0.05).(Table 7&Table 8)
the ROC curve analysis of TTK, CA125, and HE4 alone and combined detection for diagnosis of ovarian cancer. Draw the ROC curve with the benign ovarian tumor group as the reference, and calculate the AUC, the AUC of TTK, CA125, and HE4 in the joint monitoring of ovarian cancer are 0.927, 0.899, and 0.882, respectively, which are significantly higher than when each index is tested separately. The three biological indicators of TTK, CA125, and HE4 show excellent diagnostic value in the joint monitoring of ovarian cancer.(Fig. 9)