4-1BB is an inducible receptor-like protein expressed rapidly by both CD4 and CD8 T-cells after activation. 4-1BB cross-linking, either by binding to 4-1BBL or by antibody ligation, delivers a costimulatory signal to enhance T-cell activation and proliferation. Previous studies have demonstrated that the administration of 4-1BB monoclonal antibodies (mAbs) induces antitumor immune responses. In the current study using several murine tumors, we examined the systemic effects of 4-1BB mAb on the growth of s.c., intracranial (i.c.), and pulmonary metastases. In addition, the effects of 4-1BB mAb on the generation of antitumor effector T cells were examined. Treatment of 3-day i.c. MCA 205 sarcoma and GL261 glioma with the antibody resulted in prolongation of survival and cure of disease in some mice, whereas only minimal therapeutic effects were observed in established s.c. and pulmonary tumors. No antitumor effects against the poorly immunogenic B16/D5 melanoma were observed. Interestingly, successful treatment of i.c. tumors induced concomitant regression of s.c. tumors. Experiments using severe combined immunodeficient mice and mice depleted of either CD4 or CD8 T cells demonstrated T-cell dependence of the antitumor effects. For generation of effector T cells in the tumor-draining lymph nodes (LNs), administration of 4-1BB mAb had adverse effects, despite the apparent hypertrophy of the LNs. During in vitro activation of tumor-draining LN T cells with anti-CD3 and interleukin 2, the 4-1BB mAb augmented proliferation, resulting in an increase in CD8 T cells. However, they were less therapeutic than not treated LN cells. In adoptive immunotherapy, the coadministration of 4-1BB mAb enhanced the therapeutic efficacy. These results thus demonstrate the limits and potential advantages of 4-1BB antibody interactions with antitumor T cells in vivo and in vitro and suggest that therapeutic interactions of the antibody may be used in a variety of immunotherapeutic approaches.