EDDM3A Down-regulates MTA3 Expression And Promotes NSCLC Cell Epithelial-mesenchymal Transition,invasion And Metastasis

Background and Objective:Primary lung cancer is one of the most common malignant tumors within the scope of the world,with a high incidence in our country.In the next few decades,lung cancer will continue to be the important content of cancer prevention in China.Because of the long incubation period and high malignant degree,most of the lung cancer cases had metastasized at the time of diagnosis,missed the best treatment opportunity.Thus,studies on potential proliferation and metastatic mechanisms are of great importance to improve the therapeutic effect and prognosis of lung cancer.Metastasis is one of the main causes of death to patients,studies found that metastasis of malignant tumor is closely related with epithelial to mesenchymal transition.MTA3 inhibits the expression of Snail,and Snail as a transcription factor,can inhibit the epithelial marker E-cadherin transcription and activity,promote epithelial to mesenchymal transition,lead to cell polarity disappears,adhesion force drops,migration and proliferation ability enhancement.EDDM3A is a protein coding gene and locates at 14q11.2.In recent years,more and more evidence found that loci on 14q11.2 is associated with risk of tumor diseases.However,research about EDDM3A in mechanism of promoting proliferation,invasion and metastasis of lung cancer has not been reported.We propose that EDDM3A may down-regulate MTA3 expression,thus play a role.In the current study,we test the expression of EDDM3A in cancer database,lung cancer tissue and cell lines,analyze the correlation of EDDM3A and proliferation of lung cancer,explore the mechanism of metastasis in lung cancer,in order to provide new ideas of improving the therapeutic effect and prognosis of lung cancer.Methods:1.Expression of EDDM3A in NSCLC and its clinical significance.Datasets from the Cancer Genome Atlas were utilized for EDDM3A expression detection,differential analysis in NSCLC.EDDM3 A protein level was evaluated in surgical samples from 30 patients via immunohistochemistry staining,and analyzed the relationship between EDDM3A and tumor differentiated degree,T,N,clinic stages.2.EDDM3 A promotes proliferation of NSCLC.Lentivirus-mediated short hairpin RNA(shRNA)was used to knockdown EDDM3 A expression in the human A549 and 95D NSCLC cell lines and assessed by RT-qPCR and Western blot.Cell proliferation was evaluated by MTT assay and Celigo imaging cytometry.Cell apoptosis were detected by Annexin V staining.3.EDDM3A down-regulates MTA3 expression and promotes NSCLC cell EMT,invasion and metastasis.In order to explore the mechanism of EDDM3A in proliferation of non-small cell lung carcinoma,we tested the cell migration through up-regulation and down-regulation to EDDM3A.Confirm the ability of EDDM3A in regulating the invasion and metastasis in A549 and H1299 cell by Transwell and Matrigel test.Down-regulated EDDM3A in A549 and up-regulated in H1299,detected the expression of MTA3 protein by Western blot.Results:1.Data from the Cancer Genome Atlas containing 57 paired normal lung tissue samples respectively was retrieved,together with immunohistochemistry of surgical samples,we found that EDDM3A expression was significantly increased in lung carcinoma compared with adjacent non-tumor tissues.High expression of EDDM3A was related to low differential grade,advanced T,N and clinical stages.2.After lentivirus-shEDDM3A being used to infect lung cancer cell lines,EDDM3A expression was reduced observably both in mRNA and protein level.At the same time,the proliferation was significantly inhibited,the apoptosis was promoted,and colony-forming ability significantly attenuated.3.Down-regulated EDDM3A expression suppressed A549 cells migration,up-regulated EDDM3A expression promoted H1299 cells migration.Microscopic morphology observed that,down-regulated EDDM3 A expression induced H446 cells with mesenchymal cells phenotype transition to epithelial cells phenotype,over-expressed EDDM3A induced SPC-A1 cells with epithelial cells phenotype transition to mesenchymal cells phenotype.Western blot revealed that down-regulated EDDM3A enhanced MTA3 protein expression observably,and up-regulated EDDM3A suppressed MTA3 protein expression.Conclusions:1.EDDM3 A was upregulated in NSCLC and correlated with poorer clinical outcome.EDDM3 A will provide a potential new clinical stage and metastasis target for lung cancer.2.Down-regulated EDDM3A expression in the human lung cancer cell lines restrains cell proliferation.EDDM3 A promotes proliferation in NSCLC.3.Up-regulated EDDM3A expression in lung cancer cells promoted invasion,migration and epithelial-mesenchymal transition,and down-regulated EDDM3 A expression could suppressed these functions.EDDM3A could inhibit the expression of MTA3,then activate Snail,at last promote NSCLC proliferation and epithelial-mesenchymal transition.The conclusion provided a new method for prevention and treatment of NSCLC.