Objective To observe the effect of different expression levels of USP28 on the radiosensitivity of ECA109 cells by gene transfection method,aiming to provide theoretical basis for comprehensive treatment of esophageal cancer.Methods The expression levels of USP28 and c-Myc in the esophageal epithelial cells Het-1A,ECA109 and ECA109R were quantitatively measured by qRT-PCR.The specific siRNA sequences were designed according to the USP28 and c-Myc genes.The pcDNA-USP28 and pcDNA-c-Myc plasmids were constructed.The esophageal cancer cell ECA109 was transfected with Lipofectamine 2000 to observe the transfection effect and related protein expression.ECA109 and ECA109R cells were exposed to 6 Gy X-ray radiation.The cell apoptosis in each group was detected by flow cytometry.The radiosensitivity was evaluated by clone formation assay.Results The expression levels of USP28 and c-Myc in ECA109 were significantly higher than those in Het-1A (both P＜0.05),and the expression levels of USP28 and c-Myc in ECA109R were remarkably higher than those in ECA109(both P＜0.05).The pcDNA-USP28 and pcDNA-c-Myc recombinant plasmids were successfully constructed.Compared with the negative control group,the expression of USP28 at the protein and mRNA levels in the si-USP28 group was significantly down-regulated,whereas those in the pcDNA-USP28 group were remarkably up-regulated.Similar results were obtained in terms of c-Myc.Compared with the control group,the expression level of c-Myc protein was significantly up-regulated in the pcDNA-USP28 group,whereas considerably down-regulated in the si-USP28 group.After 6 Gy irradiation,the apoptosis rate and radiosensitivity of ECA109 cells were significantly declined.The apoptosis rate and radiosensitivity of ECA109R cells were increased in the si-USP28 group.Conclusions The expression of USP28 protein is closely correlated with the radiosensitivity of esophageal cancer cells.The underlying mechanism may be related to the regulation of c-Myc expression by USP28.