The role of radiotherapy-related autophagy genes in the prognosis and immune infiltration in lung adenocarcinoma

Background: There is a close relationship between radiotherapy and autophagy in tumors, but the prognostic role of radiotherapy-related autophagy genes (RRAGs) in lung adenocarcinoma (LUAD) remains unclear.

Methods: Data used in the current study were extracted from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. Weighted gene co-expression network analysis (WGCNA) was executed to recognize module genes associated with radiotherapy. The differentially expressed genes (DEGs) between different radiotherapy response groups were filtered via edgeR package. The differentially expressed radiotherapy-related autophagy genes (DERRAGs) were obtained by overlapping the module genes, DEGs, and autophagy genes (ATGs). Then, prognostic autophagy genes were selected by Cox analyses, and a risk model and nomogram were subsequently built. Gene Set Enrichment Analysis (GSEA) and single-sample Gene Set Enrichment Analysis (ssGSEA) were performed to investigate potential mechanisms through which prognostic autophagy signatures regulate LUAD. Radiotherapy-resistant cell lines (A549IR and PC9IR) were established after exposure to hypo-fractionated irradiation. Ultimately, mRNA expression was validated by quantitative real-time PCR (qRT-PCR), and relative protein levels were measured in different cell lines by western blot.

Results: A total of 11 DERRAGs were identified in LUAD. After Cox analyses, SHC1, NAPSA, and AURKA were filtered as prognostic signatures in LUAD. Then, the risk score model was constructed using the prognostic signatures, which had a good performance in predicting the prognosis, as evidenced by receiver operating characteristics curves. Furthermore, Cox regression analyses demonstrated that risk score was deemed as an independent prognostic factor in LUAD. Moreover, GSEA and ssGSEA results revealed that prognostic RRAGs may regulate LUAD by modulating the immune microenvironment and affecting cell proliferation. The colony formation assay showed that the radiosensitivity of radiation-resistant cell lines was lower than that of primary cells. The western blot assay found that the levels of autophagy were elevated in the radiotherapy-resistant cell lines. Moreover, the expression of DERRAGs (SHC1, AURKA) was higher in the radiotherapy-resistant cells than in primary cells.

Conclusion: Our study explored the role of RRAGs in the prognosis of LUAD and identified three biomarkers. The findings enhanced the understanding of the relationship between radiotherapy, autophagy, and prognosis in LUAD and provided potential therapeutic targets for LUAD patients.