Expression And Significance Of P42.3 Gene In Non-small Cell Lung Cancer

Background Lung cancer is the most malignant tumor with the highest morbidity and mortality in China,accounting for about a quarter of the death of malignant tumors in China.About 780,000 cases died of lung cancer each year.Primary lung cancer is divided into non-small cell lung cancer and small cell lung cancer,among which non-small cell lung cancer(NSCLC)accounts for 80 ～ 85%.Due to the insidious onset of lung cancer,more than 80% of the patients were already in the advanced stage(stage IIIB-IV)at the time of diagnosis.Systemic treatment is the main treatment method.The overall 5-year survival rate was only 15% to 17%.In recent years,targeted therapy and immunotherapy has made rapid progress,but chemotherapy is still an important treatment for advanced NSCLC.In view of the low effective rate of chemotherapy,most of the efficacy is unsatisfying,and even with serious toxic and side effects,it has great potential for clinical transformation to open up more therapeutic targets and efficacy predictors for NSCLC to achieve accurate treatment,improve the prognosis and survival of patients.Based on the current published research results,p42.3 gene has the characteristics of being only expressed in embryonic period,not expressed in adult normal tissues,but re-expressed in tumor tissues.It is specifically expressed in a variety of human tumors,including gastric cancer,colorectal cancer and lung cancer.However,the research of p42.3 in the field of lung cancer is still lacking.Therefore,this study through the in-depth study of bioinformatics of p42.3 gene and the verification of the expression of p42.3 gene in NSCLC with real world samples,in combination with the clinical pathological characteristics of patients to explore the clinical significance of the gene.Furthermore,by constructing lung cancer cell lines with high and low expression of p42.3,cytological experiments were carried out to explore the influence of p42.3 on the biological characteristics of lung cancer cells and the sensitivity of chemotherapy drugs.The aim of this study was to construct a stable,efficient,repeatable,economic and practical prognostic indicator for NSCLC.Method In this study(1)m RNA-seq data(1evel 3)and corresponding clinical information of1017 NSCLC patients were downloaded from The Cancer Genome Atlas(TCGA)public database,including 502 lung squamous cell carcinoma(LUSC)cases and 515 lung adenocarcinoma(LUAD)cases.Bioinformatics analysis tools such as Fire Browse(http://firebrowse.org/),UCSC Xena browser(https://xenabrowser.net/),c Bio Portal(http://www.cbioportal.org/)were used to analyze the expression of p42.3gene in LUSC and LUAD,DNA methylation level,mutation type and prognostic significance.(2)Tumor and normal tissue specimens of 142 patients with NSCLC(72 cases of LUSC and70 cases of LUAD,all of whom had not received anti-tumor treatment)admitted to Beijing Hospital from 2005 to 2009 were retrospectively collected for verification.After paraffin-embedded tissue was used and tissue microarray was made,the expression of p42.3 gene in the real world was detected by immunohistochemistry,and the correlation analysis was carried out based on the clinicopathological characteristics of the patients.(3)Firstly,p42.3 overexpression plasmid and small interfering RNA(si RNA)were constructed and transfected into human lung squamous cancer cell line NCI-H520 and human lung adenocarcinoma cell line A549 by liposomal lipo3000 membrane penetration technology to establish different histological types of lung cancer cell lines with p42.3 expression was up-regulated or down-regulated.Subsequently,Western Blot was used to verify whether there was difference in the expression level of p42.3 protein.Then CCK-8(cell counting kit-8)Assay was used to detect the change of cell proliferation ability after overexpression or silencing of p42.3 protein.Scratch Test and Transwell were used to determine the changes of p42.3 expression on the migration and invasion of lung cancer cell lines.Finally,CCK-8 in vitro drug sensitivity test was used to detect the sensitivity of lung cancer cell lines with high or low expression of p42.3 to common used chemotherapy drug carboplatin at different concentrations,and to preliminarily determine whether the expression level of p42.3 is related to the efficacy of chemotherapy.Result(1)TCGA data set analysis results showed that compared with normal lung tissues,the expression of p42.3 gene in both lung squamous cell carcinoma and lung adenocarcinoma was significantly up-regulated.In lung squamous cell carcinoma,the expression level of p42.3 m RNA was significantly higher than that of lung adenocarcinoma,which was 17.8 and 13.5 times higher than that of normal lung tissue,respectively.The p42.3 DNA methylation level of lung squamous cell carcinoma is lower than that of lung adenocarcinoma.Mutations were found in 1.1%of lung squamous cell carcinoma and 0.4% of lung adenocarcinoma samples,respectively.(2)Statistical analysis of clinical sample results showed that: in the tissue samples of lung squamous cell carcinoma,the expression rate of p42.3 protein was significantly higher than that of lung adenocarcinoma(31/72,43.1% VS 1/70,1.4%,p < 0.001).Combined with the clinicopathological features of the patients,the results showed that there was no correlation between p42.3 protein expression and gender and tumor stage.p42.3 protein expression was correlated with age(p < 0.05),and p42.3 protein expression rate decreased in patients aged 70 years and over.The expression of p42.3 protein was correlated with the pathological grade(p < 0.05).The expression of p42.3 protein was high and the histological grade was higher.(3)Compared with the control group,up-regulation of p42.3 gene expression increased the proliferation of NCI-H520 cell line(p < 0.05),decreased the sensitivity of NCI-H520 and A549 cell lines to carboplatin(p < 0.001,p < 0.05).After silencing p42.3 gene expression,nci-h520 was more sensitive to carboplatin(p < 0.05).ConclusionBoth genetic and epigenetic changes can lead to the expression disorder of p42.3gene in NSCLC.The up-regulated expression of p42.3 protein in lung squamous cell carcinoma was significantly higher than that in lung adenocarcinoma.In addition,the high expression of p42.3 was significantly correlated with the low differentiation of lung squamous cell carcinoma.Therefore,the prognostic value of p42.3 in lung squamous cell carcinoma is worthy of further study.