Adoptive transfer of tumor-sensitized T lymphocytes has demonstrated therapeutic efficacy in animal tumor models and in some patients with melanoma and renal cell cancers. In experimental settings, T lymphocytes derived from lymph nodes (LNs) draining progressively growing tumors can be activated ex vivo to generate tumor-reactive lymphocytes with therapeutic efficacy. Despite the theoretical concern regarding inaccessibility of the central nervous system to systemically transferred T cells, our recent experiments demonstrated that anti-CD3-activated tumor-draining LN cells are capable of mediating the regression of established intracerebral tumors. In this study, several staphylococcal enterotoxins (SEs), including SEA, SEC2, and SEE, and exfoliating toxin, known to be superantigens, were tested for their ability to stimulate tumor-draining LN cells to acquire antitumor reactivity for the treatment of intracerebral tumors. SEs bind to the MHC class II molecule and provide an activating signal to T cells bearing particular T-cell receptor Vbeta chains. Tumor-draining LN cells activated with SEs demonstrated selective Vbeta T-cell expansion. In adoptive immunotherapy of intracranial (IC) tumors, SEA-and SEC2-activated cells had the highest efficacy, whereas SEE-activated cells were not therapeutic. Despite the antigen independence of SE activation, the T cells retained immunological specificity for the tumor, which provided the initial in vivo sensitization of the LN. During the ex vivo stimulation with superantigens, both CD4+ and CD8+ T cells proliferated, and both subsets were required to mediate regression of IC tumors. In contrast to the adoptive immunotherapy of visceral tumors, the systemic administration of exogenous interleukin 2 failed to support the antitumor reactivity in mice depleted of CD4 cells, and, in fact, it inhibited the therapeutic efficacy. Furthermore, mice cured of intracerebral tumors by the adoptive transfer of T cells were resistant to an IC tumor rechallenge. However, in contrast to the immunological specificity demonstrated during the primary adoptive transfer, cured mice were able to reject challenge with several immunologically distinct fibrosarcomas but not a melanoma. These results indicate that superantigen-activated LN cells can circulate to and interact with intracerebral tumors mediating tumor regression in an immunologically specific manner. Although conditions that optimize the treatment of intracerebral tumors appear to be different from those for visceral tumors, analysis of T-cell receptor Vbeta expression among cells activated with several superantigens does not reflect a preferential usage of Vbeta gene segments in the immune response to autochthonous tumors.