Recently, dendritic cells (DC) transfected with tumor RNA have been used as a cancer vaccine. The efficacy of a cancer vaccine using DC transfected tumor RNA was examined. Of particular interest was whether a vaccine using DC transfected with recrudescent tumor RNA is effective for the treatment of a regrowing tumor after prior immunotherapy. In addition, the usefulness of co-transfection of granulocyte macrophage colony-stimulating factor (GM-CSF) mRNA to augment the DC vaccine was examined. CT26 tumor-bearing mice were immunized by s.c. injection with DC transfected with CT26 mRNA (DC-CT26). The cytotoxic activity against CT26 in mice immunized with DC-CT26 was significantly higher than that in the control group (P < 0.001) and was augmented by GM-CSF mRNA co-transfection (P < 0.05), resulting in remarkable therapeutic efficacy in CT26 s.c. tumor models. Cytotoxic T lymphocytes induced by the vaccination using DC transfected with mRNA from the recrudescent tumor showed a potent cytotoxicity against the recrudescent CT26 tumor cells, which was significantly higher than the cytotoxicity induced by the vaccination using DC-CT26 (P < 0.05). In addition, in a recrudescent tumor model, this vaccination suppressed the regrowing s.c. tumors, and was augmented by GM-CSF mRNA co-transfection (P < 0.05). These results suggested that vaccination therapy using DC simultaneously transfected with whole tumor RNA and GM-CSF mRNA could generate therapeutic immune responses even against recrudescent tumor after prior vaccination.