Radiation therapy is a typical treatment for esophageal squamous cell carcinoma (ESCC), especially middle and upper segment esophagus, and inoperable patients. However, how to promote radiation sensitivity in radio-resistant cancer cells is a conundrum. Here, our study investigated the radiosensitizing effect of bortezomib, a specific and reversible dipeptide boronic acid analog, in ESCC cells. Human esophageal squamous carcinoma cell lines Eca109 and TE-13 were exposed to hypoxia and/or ionizing radiation (IR) with or without treatment of bortezomib. Cell proliferation assay was performed with CCK8. Cell apoptosis and cell cycle assay were performed with flow cytometry. The radiosensitization effect of was assessed by clonogenic survival and progression of tumor xenograft. The expression of HIF-1α, VEGF, and apoptosis proteins was evaluated by Western blot. Radiation-induced DNA double strand break and homologous recombination repair were assessed by immunofluorescence. Our results show that bortezomib efficiently radiosensitizes ESCC cells by decreasing the expression of HIF- 1α and VEGF, inducing apoptosis by activating caspase, and delaying DNA damage repair after radiation.