A Preliminary Study On The Role And Mechanism Of CLIC4 In The Development Of Esophageal Squamous Cell Carcinoma

Background China is one of the regions with the highest incidence of esophageal cancer in the world,esophageal cancer is the fifth leading cause of cancer death,with an average of about 150,000 deaths each year.The main form of esophageal cancer in China is esophageal squamous cell carcinoma（ESCC）.Despite continuous advances in diagnostic techniques and therapies,the five-year overall survival rate is still not high and the prognosis is poor^[2].Therefore,the study of ESCC biomarkers or drug targets is very important,especially for China,where the incidence of esophageal squamous cell carcinoma is high.The chloride intracellular channels（CLICs）protein family has different functions,such as dielectric transport,ion homeostasis,cellular p H balance,and cell volume regulation.They are involved in regulating tumor cell proliferation,migration and non-tumor tissue invasion,so may be potential targets for tumor treatment.CLIC4,in particular,is not only overexpressed in specific tumor types or their corresponding stroma,but also changes localization and function from hydrophilic cytosolic to integral transmembrane proteins as active ionic channels or signal transducers during cell cycle progression.In certain cases,it can target tumor cells without affecting the physiological functions of normal cells.Objective To investigate the expression of CLIC4 in cancerous tissues of patients with ESCC.To investigate the function of CLIC4 in regulating the proliferation,migration or invasion of ESCC cells.This study aims to further reveal the role of CLIC4 in ESCC and its regulatory mechanism,and provides new ideas for the development of drugs for the treatment of ESCC.Methods Immunohistochemistry and Western blot were used to analyze the expression of CLIC4 in normal and cancer tissues.CLIC4 stable low-expressing cell lines with lentiviral sh RNA were constructed.Colony formation assays and CCK-8 assays were used to detect the proliferation of ESCC cells after CLIC4 knockdown.Western blot was used to identify the changes of migration-related proteins such as E-cadherin and N-cadherin in ESCC cells after CLIC4 knockdown,and potential downstream target molecules of CLIC4 including TRIM25,NDRG1 and MCM2.Results CLIC4 is highly expressed in ESCC patient tissues and cell lines,which is consistent with RNA-seq results.A stable CLIC4 low-expressing ESCC cell line was successfully constructed.CLIC4 does not affect the proliferation of ESCC cells,and it promotes cancer progression by promoting the migration of ESCC cells.Western blot results showed that CLIC4 knockdown,up-regulated TRIM25 expression and down-regulated NDRG1 and MCM2 expression.Conclusion CLIC4 does not affect the proliferation of ESCC cells.It may promote cancer progression by promoting the migration of ESCC cells.CLIC4 may function by down-regulating the expression of TRIM25 and up-regulating the expression of NDRG1 and MCM2.