Interleukin-21 (IL-21) is a recently characterized T cell-derived cytokine with a significant homology to IL-2,IL-4 and IL-15. To determine whether IL-21 has broad immunoregulatory activity and can stimulate durable antitumour responses, we constructed mouse IL-21 (mIL-21) recombinant plasmid and evaluated its antitumor efficacy. Mouse IL-21 cDNA was amplified from Con A-activated mouse T cells by RT-PCR. Recombinant pcDNA3.1/mIL-21 was constructed and transfected into Sp2/0 cells. Mouse IL-21 expression was analyzed by West blotting and its activities were detected by 3H-TdR incorporation and MTT assay. The recombinant pcDNA3.1/mIL-21 was injected s.c. into tumor lump. Tumor size, weight, the activities of CTLs, NK cells and LAK cells and serum IFN-γlevel were measured for evaluating mIL-21 mediated antitumor responses. The results indicated that mIL-21 was correctly expressed in Sp2/0 cells and it can improve the proliferation of T cells and B cells, and enhance NK cytotoxic activity in vitro. The activities of CTL and NK cells, and serum IFN-γlevel were significantly improved, furthermore the tumor growth was obviously suppressed in pcDNA3.1/mIL-21 treated mice. However, the LAK activity did not alter significantly. Taken together, this study suggests that the injection with recombinant plasmid containing mIL-21 is a potential approach for tumor gene therapy. Cellular & Molecular Immunology. 2004;1(6):461-466.