Synergistic Combinations of Chimeric Anti-Seb: Engineering Anti-Toxins

Engineering anti-toxins Staphylococcal enterotoxin B (SEB) is a superantigen that causes food poisoning and toxic shock syndrome. The biological activity of SEB arises from its ability to nonspecifically activate large numbers of T cells by simultaneously binding to the MHC-II on antigen presenting cells and TCRs of T cells belonging to a subset of V2 subfamilies. Antibodies can neutralize these toxins by blockading either of these sites. <P> These studies arise from the hypothesis that combinations of antibodies targeting different and spatially separated neutralizing epitopes will act synergistically. The antibodies for this study will be obtained by screening a library of anti-SEB hybridomas generated from Balb/C mice immunized with native SEB. <P> The screening strategy will identify two groups of neutralizing antibodies, one of which recognizes epitopes in the TCR- binding site of SEB and a second group that reacts with determinants in the MHC-II binding site of the toxin. Combinations of noncrossreactive members of each of these groups will be tested to determine which act synergistically. The VH and VL regions of each of the members of the most potent neutralizing combination will be cloned and grafted to sequences encoding the human IgG1CH (mouse VH ) and human Ig:CL (mouse VL ). The chimerized human-mouse antibodies which would be expected to produce much less HAMA reaction in clinical applications will be evaluated by comparing the neutralization activity of the chimeric set and its members with that of its mouse counterparts. <P> The proposal has the following specific aims: <br> 1. Generate neutralizing antibodies against the MHC binding site of SEB and derive neutralizing antibodies specific for the TCR beta binding site. <P> 2. Identify neutralizing antibodies specific for different epitopes of SEB and determine if combinations of these antibodies synergistically inhibit SEB. <P> 3: Graft the VH and VL domains of each member of the most potent sets of synergistically acting neutralizing antibodies onto human CH and CL frameworks. <P> Combinations of antibodies that synergistically inhibit SEB may have two important practical advantages should they see application in a clinical setting. First, it should be less expensive because much smaller amounts would be needed than with single monoclonal antibodies. Second, the ability to achieve an equivalent therapeutic result with smaller amounts of antibody should decrease the incidence and intensity of side effects. <P> Staphylococcal enterotoxin B (SEB) can cause food poisoning and toxic shock syndrome. Antibodies can neutralize SEB and this study is guided by the hypothesis that combinations of mouse monoclonal antibodies targeting different neutralizing sites will act synergistically. Chimeric human-mouse forms of the most effective combinations will be engineered to produce antibodies that will be better tolerated in humans.