DBCCR1 Inhibits The Development Of Lung Cancer Via Regulating DNMT1

Background: Lung cancer has become the highest morbidity and mortality in the world,and has become a serious public health problem.In 2017,the latest statistics showed that the number of new lung cancer reached 800 thousand,which is the first cancer in China.Genetic alterations such as gene mutation and DNA methylation and other epigenetic abnormalities are considered to play a crucial role in the development of lung cancer.Epigenetic mutation is considered to play a key role in the development of tumors.For example,DNA aberrant methylation leads to the development of human tumors.Studies have revealed that DNA hypermethylation in the promoter region of tumor suppressor genes in tumor tissues can lead to its silence and further promote tumor progression.Therefore,the study of the function and mechanism of hypermethylation inactivation of tumor related gene promoter is beneficial to reveal the pathogenesis of lung cancer and provide a theoretical basis for the comprehensive prevention and treatment of lung cancer.DBCCR1 is a tumor suppressor gene with low expression in human bladder cancer tissue.It is revealed that low expression of DBCCR1 can lead to gene silencing through hypermethylation.At present,DBCCR1 related hypermethylation has been reported in oral squamous cell carcinoma,hepatocellular carcinoma and glioma.However,there is still a lack of information about DBCCR1 in lung cancer. Therefore,DBCCR1 may play an important role in the activation of tumor suppressor genes in different cancer species.In order to study the potential mechanism of DBCCR1 in lung cancer,we analyzed the expression of DBCCR1 and DNA methyltransferase DNMT1 in the tissues of lung cancer patients and human lung cancer cell lines,and used the strategy of gene function acquisition or deletion to explore the effect of DBCCR1 on the development of lung cancer by DBCCR1.This study provides experimental basis for the role of DBCCR1 in lung cancer,and provides a theoretical basis for exploring new molecular markers of lung cancer and intervention targets.Objective: To explore the role and potential mechanism of DBCCR1 in the occurrence and development of lung cancer,and to provide a theoretical basis for exploring new molecular markers of lung cancer and intervention targets.Methods: 1.RT-QPCR Detection of Expression Levels of DBCCR1 and DNMT1 in Lung Cancer Tissues and Peripheral Normal Tissues 2.Establishment of DBCCR1 stable and knockdown lung cancer cell lines by using RNA interference and over expression technology using lentivirus system.3.CCK-8 test was used to detect the effect of DBCCR1 on the proliferation of lung cancer cells.4.Cell scratch test was used to analyze the effect of DBCCR1 on the migration of lung cancer cells.5.Transwell cell invasion assay was used to analyze the effect of DBCCR1 on the invasion of lung cancer cells.6.RT-PCR and Western blot analysis of DBCCR1 regulating DNMT1.7.Using SPSS18.0 statistical software.According to the type of data distribution,we choose the expression method and the statistical test method between groups.Chi square test,t test,F test and unconditional Logistic regression were used to analyze the results.Results: 1.RT-QPCR was used to detect the expression level of DBCCR1 in the normal tissues surrounding lung cancer tissues and adjacent tissues.The results showed that the expression of DBCCR1 in lung cancer tissues was significantly lower than that of normal tissues surrounding the cancer.Further analysis of the correlation between the expression level of DBCCR1 and the characteristics of clinical cases revealed that low expression of DBCCR1 was positively correlated with tumor progression;the later the tumor stage,the lower the expression of DBCCR1.In patients with low expression of DBCCR1,Ki-67,a proliferation marker,was highly expressed(p = 0.000),and tumor metastasis(p = 0.001)and tumor invasion(p = 0.000)were significantly increased.Kaplan-Meier survival analysis showed that patients with high expression of DBCCR1 had longer overall survival.Further Cox regression analysis of a variety of potential prognostic factors affecting survival revealed that Ki-67 and DBCRR1 expression levels,tumor stage,tumor metastasis,and tumor invasion are relatively independent prognostic markers of overall survival.2.Effect of DBCCR1 on the growth of lung cancer cells: Using lentivirus system to apply RNA interference and overexpression technology to establish a stable knockdown and overexpression of DBCCR1 lung cancer cell lines;cell proliferation was detected by CCK-8 experiments,the results showed that overexpression of DBCCR1 significantly reduced the proliferation of lung cancer cell line A549;and knocked down DBCCR1 significantly enhances A549 cell proliferation.3.Effect of DBCCR1 on lung cancer cell migration: The effect of DBCCR1 on migration of lung cancer cells was analyzed by cell scratch assay.It was found that overexpression of DBCCR1 significantly decreased A549 cell migration;knocking down DBCCR1 significantly enhanced A549 cell migration.4.Effect of DBCCR1 on lung cancer cell invasion: Transwell cell invasion assays showed that overexpression of DBCCR1 significantly decreased the invasion of A549 cells;knocking down DBCCR1 significantly enhanced the invasion of A549 cells.5.DBCCR1 regulates DNMT1 expression: The expression of DNMT1 in tumor tissues and normal tissues around the tumor was compared by RT-QPCR.The results showed that the expression of DNMT1 in tumor tissues was significantly higher than that of the surrounding normal tissues,and the expression of DNMT1 was negatively correlated with the expression of DBCCR1;The expression of DNMT1 was significantly inhibited in A549 cells expressing DBCCR1,while the protein and RNA levels of DNMT1 were significantly increased in A549 cells silenced by DBCCR1.Conclusion: 1.Low expression of DBCCR1 is associated with progression of lung cancer and is a poor prognostic factor for lung cancer 2.DBCCR1 can inhibit the proliferation,migration and invasion of lung cancer cell A549 cells,and inhibit the occurrence and development of lung cancer by regulating DNMT1.