Identification of Key Gene and Functional Pathways in Radioresistance of non-small cell lung cancer

For better understanding of radiotherapy resistance and its potential mechanism, we established radioresistance cell lines of non-small cell lung cancer (NSCLC) followed by microarray analysis. 529 differentially expressed genes (DEGs) were then screened between radiation resistant cell lines compared with the sensitive cell lines. The biological functions and enrichment pathways of the above DEGs were identified using Kyoto Encyclopedia of Genes and Genomes (KEGG) and gene ontology (GO) enrichment analyses. Gene Set Enrichment Analysis (GSEA) revealed that the radiation resistance group had the most gene sets enriched in altered immune response, such as TNF signaling pathway, when compared to the radiation sensitive group. Protein-protein interaction (PPI) network was carried out from the STRING database. Following that, by means of Cytoscape software, five hub genes (CXCL10, IFIH1, DDX58, CXCL11, RSAD2) were identified, and qRT-PCR confirmed the expression of the above hub genes. ChIP-X Enrichment Analysis showed that STAT1 might be the transcription factor of the above hub genes. Our results suggested that although immune system activation occurs followed by radiation resistance, the unregulated expression of PD-L1 ultimately leads to the exhaustion of anti-tumor immunity, which may be the possible mechanisms of tumor resistance to radiotherapy. The lung cancer cell line A549 in logarithmic growth phase was irradiated with 4 Gy for the first time and cultured for subculture. After cell adherence growth, the cells were irradiated again with 4 Gy, with a total dose of 60Gy. The radiotherapy resistance cell line A549-RR was obtained by further culture. Total RNA of cultured cells was isolated using TRIzol reagent (Invitrogen), there were three samples in each group and the samples were sent to Shanghai Kangcheng Biological Co LTD for RNA microarray analysis.