Tilahun, Mulualem E.

Abstract Staphylococcal Enterotoxin B (SEB) is a superantigen made by S. aureus. SEB can cause food poisoning and shock and its stability and ease of preparation make it a potential agent of bioterrorism. SEB-mediated activation induces T cell proliferation and secretion of cytokines. Antibodies that block the interaction between SEB and MHC or SEB and TCR, can neutralize the activating effects of SEB and provide protection against its debilitating effects. Human IVIG, a therapy for SEB intoxication is of low potency and sometimes, short supply. To surmount we have derived of mouse monoclonal antibodies that block the binding of SEB to MHC at separate and distinct sites. Combinations of these antibodies synergistically neutralize SEB mediated T cell activation. Genes encoding the VL and VH genes of synergistically neutralizing pairs of cell lines were cloned and grafted onto genes encoding either the constant region of human IgH or Igκ and transfected into SP2/0 cells. Chimeric human-mouse antibodies expressed by the transfectants were purified. Tests of these chimeric antibodies demonstrated that they were neutralizing and in combination with each other, still capable of synergistic neutralization of SEB. Supported by NIH grant # AI057652

ABSTRACTStaphylococcal enterotoxin B (SEB), a shock-inducing exotoxin synthesized byStaphylococcus aureus, is an important cause of food poisoning and is a class B bioterrorism agent. SEB mediates antigen-independent activation of a major subset of the T-cell population by cross-linking T-cell receptors (TCRs) with class II major histocompatibility complex (MHC-II) molecules of antigen-presenting cells, resulting in the induction of antigen independent proliferation and cytokine secretion by a significant fraction of the T-cell population. Neutralizing antibodies inhibit SEB-mediated T-cell activation by blocking the toxin's interaction with the TCR or MHC-II and provide protection against the debilitating effects of this superantigen. We derived and searched a set of monoclonal mouse anti-SEB antibodies to identify neutralizing anti-SEB antibodies that bind to different sites on the toxin. A pair of non-cross-reactive, neutralizing anti-SEB monoclonal antibodies (MAbs) was found, and a combination of these antibodies inhibited SEB-induced T-cell proliferation in a synergistic rather than merely additive manner. In order to engineer antibodies more suitable than mouse MAbs for use in humans, the genes encoding the VL and VH gene segments of a synergistically acting pair of mouse MAbs were grafted, respectively, onto genes encoding the constant regions of human Igκ and human IgG1, transfected into mammalian cells, and used to generate chimeric versions of these antibodies that had affinity and neutralization profiles essentially identical to their mouse counterparts. When tested in cultures of human peripheral blood mononuclear cells or splenocytes derived from HLA-DR3 transgenic mice, the chimeric human-mouse antibodies synergistically neutralized SEB-induced T-cell activation and cytokine production.