BackgroundAs the most powerful T cell agonists known, superantigens (SAgs) have enormous potential for cancer immunotherapy. Their development has languished due to high incidence (60%–80%) of seroreactive neutralizing antibodies in humans and tumor necrosis factor-α (TNFα)-mediated cardiopulmonary toxicity. Such toxicity has narrowed their therapeutic index while neutralizing antibodies have nullified their therapeutic effects.MethodsFemale HLA-DQ8 (DQA*0301/DQB*0302) tg mice expressing the human major histocompatibility complex II (MHCII) HLA-DQ8 allele on a high proportion of PBL, spleen and lymph node cells were used. In the established tumor model, staphylococcal enterotoxin G and staphylococcal enterotoxin I (SEG/ SEI) (50 µg each) were injected on days 6 and 9 following tumor inoculation. Lymphoid, myeloid cells and tumor cell digests from tumor tissue were assayed using flow cytometry or quantitated using a cytometric bead array. Tumor density, necrotic and viable areas were quantitated using the ImageJ software.ResultsIn a discovery-driven effort to address these problems we introduce a heretofore unrecognized binary complex comprizing SEG/SEI SAgs linked to the endogenous human MHCII HLA-DQ8 allele in humanized mice. By contrast to staphylococcal enterotoxin A (SEA) and staphylococcal enterotoxin B (SEB) deployed previously in clinical trials, SEG and SEI does not exhibit neutralizing antibodies in humans or TNFα-mediated toxicity in humanized HLA-DQ8 mice. In the latter model wherein SAg behavior is known to be ‘human-like’, SEG/SEI induced a powerful tumoricidal response and long-term survival against established melanoma in 82% of mice. Other SAgs deployed in the same model displayed toxic shock. Initially, HLA-DQ8 mediated melanoma antigen priming, after which SEG/SEI unleashed a broad CD4+ and CD8+ antitumor network marked by expansion of melanoma reactive T cells and interferon-γ (IFNy) in the tumor microenvironment (TME). SEG/SEI further initiated chemotactic recruitment of tumor reactive T cells to the TME converting the tumor from ‘cold’ to a ‘hot’. Long-term survivors displayed remarkable resistance to parental tumor rechallenge along with the appearance of tumor specific memory and tumor reactive T memory cells.ConclusionsCollectively, these findings show for the first time that the SEG/SEI-(HLA-DQ8) empowers priming, expansion and recruitment of a population of tumor reactive T cells culminating in tumor specific memory and long-term survival devoid of toxicity. These properties distinguish SEG/SEI from other SAgs used previously in human tumor immunotherapy. Consolidation of these principles within the SEG/SEI-(HLA-DQ8) complex constitutes a conceptually new therapeutic weapon with compelling translational potential.

Staphylococcal enterotoxins (SE) induce powerful T cell stimulation with specificity for the variable beta (Vβ) region of the T cell receptor (TCR). Additionally, MHC in mice (H2) and humans (human leukocyte antigen; HLA) differ in binding superantigens and, in turn, provide different T cell activating potentials. We have previously demonstrated that SEG and SEI, administered together, elicited progression free survival of B16–F10 challenged HLA-DQ8 (DQ8; DQA1*0301 and DQB1*0302) transgenic mice. C57BL/6 (B6) mice did not benefit from SEG/SEI given post B16–F10 challenge, as expected, given a markedly reduced T cell proliferation in the B6 mice compared to the DQ8 mice. We provide evidence here of the potential of dendritic cells (DCs) to be the vector for a SEG and SEI immunotherapy. Primary antigen presenting cells, DCs and monocytes, were isolated from the bone marrow of DQ8 or HLA-DR3 (DRB1*0301) transgenic mice. SEG/SEI loaded DQ8 and DR3 DCs drove a strong B6 CD8 T cell response at low T cell:DC ratios; whereas both DQ8 and DR3 DCs at a high T cell:DC ratios activated CD4 T cells. Collectively, these data support incorporating SEG and SEI into the current DC based immunotherapeutic strategies to induce a stronger and direct anti-cancer T cell response.