EXO1 Regulates Malignant Phenotype And Immunological Effects Of Radiotherapy In Lung Adenocarcinoma

Background: Non-small cell lung cancer(NSCLC)is the most common malignant tumor with the highest incidence and mortality rates in China,and requires more in-depth research.Radiotherapy combined with immunotherapy has shown good clinical efficacy in NSCLC,but only a small number of patients currently benefit from it.DNA repair proteins participate in a variety of damage repair responses after radiotherapy,thereby affecting the immunogenicity induced by radiotherapy,which has become a hot topic in the field of radiotherapy and immunotherapy research.Objective: This study aims to explore the effect of DNA repair protein exonuclease I(EXO1)on the malignant biological behavior of lung adenocarcinoma,and to verify its role as a novel prognostic biomarker for lung adenocarcinoma.The study also investigates the impact of EXO1 on radiotherapy-induced immunogenicity in lung adenocarcinoma,providing a theoretical basis and new target for radiotherapy-induced immunogenicity.Methods: Firstly,in this study,the transcriptome data and clinical data of lung adenocarcinoma patients were downloaded from TCGA,and correlation analysis was used to explore the correlation between the expression level of EXO1 and clinical characteristics in lung adenocarcinoma.Survival analysis was used to further investigate the relationship between EXO1 expression levels and prognosis of lung adenocarcinoma patients.The correlation between EXO1 expression and the immune microenvironment was explored using algorithms such as ESTIMATE and ss GSEA.A549 lung cancer cells were used as the cell model,and CRISPR-Cas9 technology was used to regulate the expression of EXO1 in cells.The effects of EXO1 on the malignant biological behavior of lung adenocarcinoma cells were explored through cell proliferation experiments,clone formation assays,wound-healing experiments,and flow cytometry.Finally,the effects of EXO1 on radiation-induced immune-related molecules and pathways were validated through real-time fluorescent quantitative PCR and protein blotting.Immunofluorescence was used to detect changes in the amount of ds DNA in the cytoplasm after radiotherapy.Results:(1)Bioinformatics analysis showed that the expression of EXO1 was significantly higher in lung cancer tissues.Clinical correlation analysis revealed that the expression of EXO1 was correlated with worse staging and grading in lung adenocarcinoma patients.Survival analysis showed that high expression of EXO1 was significantly associated with poor clinical outcomes in lung adenocarcinoma.On the other hand,immune-related analysis showed that EXO1 was significantly associated with immune cell infiltration in lung adenocarcinoma patients.Drug sensitivity analysis results suggested that patients with high expression of EXO1 were more sensitive to multiple chemotherapy drugs.(2)Cloning formation and CCK8 experiments showed that knocking out EXO1 inhibited the proliferation of A549 cells.The wound-healing experiment showed that knocking out EXO1 inhibited the migration ability of A549 cells.Flow cytometry showed that knocking out EXO1 resulted in A549 cells being blocked in the G1 phase.(3)In A549 cells after radiotherapy,the interferon pathway-related molecules were significantly downregulated in the EXO1 knockout group.(4)Immunofluorescence results showed that knocking out EXO1 significantly reduced the content of ds DNA in the cytoplasm of A549 cells after radiotherapy.Conclusion: High expression of EXO1 is associated with poor prognosis in patients with lung adenocarcinoma,and enhances the proliferation and migration abilities of lung adenocarcinoma cells.On the other hand,EXO1 may enhance the radiation-induced immune response by regulating the release of cytoplasmic ds DNA after radiation.Therefore,radiotherapy combined with immunotherapy may be a potential clinical treatment option for patients with high EXO1 expression and poor prognosis in non-small cell lung cancer.