In this study, 311 DEGs were identified including 74 up-regulated and 238 down-regulated genes in five datasets from the GEO database. Furthermore, through Enricher online web tool, we visualized the outcomes derived from Gene Ontology and KEGG pathway enrichment analysis. As for the biological processes, these DEGs were enriched in extracellular matrix organization, regulation of angiogenesis, negative regulation of blood vessel morphogenesis, and negative regulation of angiogenesis and so on. As for the molecular functions, the DEGs were mainly enriched in amyloid-beta binding, protein homodimerization activity, metalloendopeptidase inhibitor activity, low-density lipoprotein particle binding and calcium ion binding. For the cellular components, the DEGs showed enrichment in the integral component of plasma membrane. Angiogenesis is closely related to the occurrence and progression of cancers [18] and the formation of extracellular matrix is also associated with tumor metastasis and invasion [19, 20]. According to KEGG pathway enrichment analysis, tyrosine metabolism was found to be significant in LUAD. Li et al. revealed that activation of tyrosine metabolism in CD13+ cancer stem cells may drive relapse in hepatocellular carcinoma by means of generating nuclear acetyl-CoA to acetylate and stabilize Foxd3, and allowing CD13+ cancer stem cells to sustain quiescence and resistance to chemotherapeutic agents[21]. Nitration of protein tyrosine has proved to be involved in a variety of biological processes, including signal transduction, protein degradation, energy metabolism, mitochondrial dysfunction, enzyme inactivation, immunogenic response, cell apoptosis and cell death, and plays an important role in the occurrence and metastasis of lung cancer[22]. Therefore, the signaling pathway of tyrosine metabolism was expected to be a potential drug therapy target for LUAD.
Next, DEGs PPI network was constructed via the STRING online database and Cytoscape software. By virtue of “CytoHubba” plug-in, the top ten hub genes, CDC20, CENPF, TPX2, TOP2A, KIAA0101, CDCA7, ASPM, ECT2, UBE2T, COL1A1 were identified and they were all found up-regulated in LUAD. In addition, miRWalk, an online analysis tool, was used to construct a network of miRNAs associated with the regulation of these genes, and finally four genes (TOP2A, CDCA7, TPX2 and COL1A1) were found to show strong associations with each other, which could be considered as new effective targets to improve the prognosis of LUAD patients.
Topoisomerase II (TOP2) has been clarified to have crucial functions, including DNA replication, transcription and chromosome segregation, and more and more active anticancer drugs targeted it[23]. TOP2 contains two types of isozymes: TOP2A and topoisomerase II beta (TOP2B)[24], TOP2A is the only enzyme able to cleave and re-ligate the double-strand backbone of DNA, which is indispensable for DNA replication, transcription, and repair[25, 26]. Ejlertsen et al.[25] showed that TOP2A was a direct molecular target of anthracyclines that can improve the sensitivity of anthracycline-containing chemotherapy in high-risk breast cancer patients. In malignant peripheral-nerve sheath tumor, TOP2A was the most overexpressed gene compared with benign neurofibromas[27]. High expression of TOP2A was found to be correlated to worse overall survival (OS) in all non-small-cell lung cancer and lung adenocarcinoma patients, but not in lung squamous cell carcinoma patients[28, 29]. It has also been reported that miRNA’s being associated with TOP2A plays an important role in lung cancer, for example, down-regulation of miRNA-144-3p whose potential target was TOP2A, was highly enriched in various key pathways like the protein digestion and absorption and the thyroid hormone signaling pathways in non-small cell lung cancer from the comprehensive meta-analysis[30].
Cell division cycle associated 7, CDCA7, was identified as a c-MYC responsive gene, and behaved as a direct c-MYC target gene. Overexpression of CDCA7 was found to enhance the transformation of lymphoblastoid cells, and it complements a transformation-defective MYC Box II mutant, suggesting its involvement in c-MYC-mediated cell transformation[31]. In quite a number of tumors, such as hepatocellular carcinoma[32], colorectal cancer[33], lymphoma[34], breast cancer[35], CDCA7 was all reported up-regulated and might be a potential prognostic factor and therapeutic target. Wang et al. have found that CDCA7 could promote lung adenocarcinoma proliferation via regulating the cell cycle and silencing CDCA7 inhibited cell proliferation in LUAD through G1 phase arrest and induction of apoptosis, which implied that CDCA7 might be identified as a potential therapeutic target for new biomarkers and LUAD[36].
TPX2, which is also known as DIL2 or p100, uses two flexibly linked elements ('ridge' and 'wedge') in a novel interaction mode to simultaneously bind across longitudinal and lateral tubulin interfaces[37, 38]. In ovarian cancer, it can promote the proliferation and migration of human ovarian cancer cells by regulating PLK1 expression[39]. Except for these function, in various cancers can TPX2 also control bladder cancer cell’s proliferation and invasion via TPX2-p53-GLIPR1 regulatory circuitry[40], regulate the PI3K/AKT signaling pathway to facilitate hepatocellular carcinoma[41], interactive with miRNA such as miR-485-3p[42], miR-361-5p[43], miR-335-5p[44], miR-216b[45] and so on. Zhou et al. have verified that TPX2 can activate the epithelial-mesenchymal transition process and promote both the expression and activities of matrix metalloproteinase (MMP)2 and MMP9 in non-small cell lung cancer (NSCLC), which means TPX2 promotes the metastasis and malignant progression of NSCLC and could thus serve as a marker of poor prognosis in NSCLC[46].
Some studies have confirmed that COL1A1-related miRNAs were involved in the regulation of different kinds of tumors, including LUAD[17, 47]. Zhang et al. found that COL1A1 was positively correlated with NOTCH3 expression, and the miR-150 /NOTCH3/COL1A1 axis might be involved in EGFR-TKI resistance in LUAD, which provided a potential target for LUAD therapy and could be used as a prognostic target[48]. Hsa-let-7d-5p has been proved that it can regulate cell cycle including both G1/S and G2/M cell cycle phase transitions and telomere maintenance in human lung fibroblasts and is related to cellular senescence [17]. Hsa-miR-483-3p can also play an important role in lung cancer caused by radon exposure by regulating certain signaling pathways such as proliferative protein kinases (MAPK) and reactive oxygen species (ROS) [49].
Followed by, ROC curve analysis of these 10 hub genes was conducted based on dataset GSE43458, and we found that they all had certain diagnostic efficacy, which TOP2A ranked first. The results of Kaplan-Meier survival analysis also showed that these hub genes were significantly correlated with the prognosis of LUAD patients, suggesting that they may be potential prognostic biomarkers of LUAD.