We constructed an adeno-associated virus (AAV) vector containing the human interferon-beta (HuIFN-b ) gene (AAV-IFN-beta) and investigated its antitumor effect against human glioma cells (U251-SP) inoculated into the brain of nude mice. Prior to this, we examined human glioma cells transduced with AAV-IFN-beta using video-enhanced contrast differential interference contrast (VEC-DIC) microscopy. Infection of AAV-IFN-beta induced apoptosis and secondary necrosis in human glioma cells. In in vivo experiments, we confirmed production of HuIFN-beta and induction of heat-shock protein (HSP) in glioma cells transduced with AAV-IFN-beta. Growth of the experimental gliomas was completely inhibited by six injections of AAV-IFN-beta, starting 7 days after transplantation of glioma cells. In addition, the survival of mice treated with AAV-IFN-beta was remarkably prolonged. These results indicate that AAV-IFN-beta induces apoptosis of glioma cells and has a strong antitumor effect in this experimental glioma model.