The Screening And Functional Study Of Genes With Copy Number Amplification In Esophageal Squamous Cell Carcinoma And Gastric Cardia Adenocarcinoma

Background&Aims:Esophageal squamous-cell carcinoma(ESCC)is one of the most frequently diagnosed malignant tumor derived from esophageal squamous epithelium.China is one of the countries with high incidence and mortality rate of ESCC.Our previous study of ESCC has identified 23 recurrent amplified focal regions containing 1,591 genes.Further integrative analysis of copy number and mRNA expression level showed 149 copy-number amplified genes overexpressed in ESCC simultaneously,suggesting important roles of these genes in ESCC progression.Therefore,the present study aims to investigate the functional roles of these genes and their potential as therapeutic targets for ESCC.Methods:Select target gene VAV2 from candidate genes screened by high-throughput technology.Levels of mRNAs and proteins in tissue samples were measured by RNA-sequencing and immunohistochemistry,respectively.The potential mechanisms were explored using molecular and cell biological assays.Candidate gene was knocked down or overexpressed in ESCC cells(KYSE30 and KYSE150)using CRISPER/Cas9 and lentiviral vectors.We analyzed its effects on migration in vitro and in vivo,and analyzed effects on invasion in vitro.Results:We found that VAV2 affected both proliferation and invasiveness abilities of ESCC cells.We investigated mRNA and protein levels of VAV2 in ESCC and adjacent normal tissues.The results showed that VAV2 was up-regulated in tumor tissues,suggesting that VAV2 may act as an oncogene in ESCC.By constructing stable VAV2-knockdown and-overexpressed ESCC cell lines,we found that down-regulation of VAV2 significantly inhibited the proliferation,colony formation and migration abilities of ESCC cells,whereas up-regulation of VAV2 resulted in opposite results.In vivo experiments showed the similar results with in vitro studies.Conclusion:By analyzing in ESCC samples and cell lines,we associated copy number amplification and elevated expression of VAV2 with tumor proliferation and invasiveness abilities for the first time.Further investigation of the biological mechanism of VAV2 in cancer development and progression is of great significance for the early diagnosis and treatment of ESCC.