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Abstract Background Caspase 3 is not only involved in apoptosis, but also participates in the nonapoptotic functions. Previously, we found that caspase 3 gene knockout mice displayed decreased number of dendritic cells (DCs). However, whether caspase 3 participate in the function of DCs is unclear. Thus, the present study aims to investigate the role of caspase 3 in the maturation and antitumor function of DCs. Methods Caspase 3 gene was overexpressed in DC2.4 cell line through Lentivirus system. The impact of caspase 3 gene overexpression on the biological behavior of DC2.4 cells was determined by CCK-8, colony formation and apoptosis analysis. The impact of caspase 3 gene overexpression on the antigen uptake, maturation, migration, T cell activation of DC2.4 cells was analyzed with phagocytosis, transwell and mixed lymphocyte reaction assay. Tumor growth and tumor infiltrated T cells were also investigated through tumor bearing model. Results Caspase 3 gene overexpression could slightly increase the apoptosis of DC2.4 cells. Antigen uptake capability and maturation of DC2.4 cells were significantly promoted through caspases 3 gene overexpression. However, CXCR4 expression on DC2.4 cells and migration of DC2.4 cells were not influenced. Caspase 3 gene overexpression also enhanced the T cell activation and cytotoxicity of activated T cells. Finally, overexpression of caspase 3 gene significantly increased the tumor suppression of DC2.4 cells, accompanied by increased infiltration of CD4+ and CD8+ Cells in tumor tissue. Conclusion Caspase 3 gene overexpression could promote maturation and enhance antitumor capability of DC2.4 cells. Highlights • Caspase 3 overexpression only slightly influenced apoptosis of DC2.4 cells. • Antigen uptake, maturation and T cell activation of DC2.4 cells were enhanced. • Finally, antitumor immunity of DC2.4 cells were enhanced. [ABSTRACT FROM AUTHOR]