Synergistic combinations of chimeric anti-SEB: Engineering anti-toxins

描述（由申请人提供）：嵌合抗SEB的协同组合：工程抗毒素葡萄球菌肠毒素B （SEB）是一种引起食物中毒和中毒性休克综合征的超抗原。SEB的生物活性源于其通过同时结合抗原提呈细胞和属于V2亚家族的T细胞的tcr上的MHC-II而非特异性激活大量T细胞的能力。抗体可以通过阻断这两个位点来中和这些毒素。这些研究源于这样的假设，即针对不同和空间分离的中和表位的抗体组合将协同作用。本研究的抗体将通过筛选经天然SEB免疫的Balb/C小鼠产生的抗SEB杂交瘤文库获得。筛选策略将确定两组中和抗体，其中一组识别SEB TCR结合位点的表位，另一组与毒素MHC-II结合位点的决定因子发生反应。这些组的非交叉反应的成员的组合将被测试，以确定哪些协同作用。最有效的中和组合的每个成员的VH和VL区域将被克隆并移植到编码人类IgG1CH（小鼠VH）和人类Ig:CL（小鼠VL）的序列上。嵌合的人-鼠抗体在临床应用中产生较少的HAMA反应，将通过比较嵌合集及其成员与小鼠对应物的中和活性来评估。该建议有以下具体目的：产生针对SEB MHC结合位点的中和抗体，并获得针对TCR β结合位点的特异性中和抗体。2. 鉴定针对SEB不同表位的特异性中和抗体，并确定这些抗体的组合是否协同抑制SEB。3：将最有效的协同作用中和抗体的每个成员的VH和VL结构域移植到人CH和CL框架上。协同抑制SEB的抗体组合在临床应用中可能有两个重要的实际优势。首先，它应该更便宜，因为与单克隆抗体相比，它所需的数量要少得多。其次，使用更少量的抗体也能达到相同的治疗效果，这应该会降低副作用的发生率和强度。葡萄球菌肠毒素B （SEB）可引起食物中毒和中毒性休克综合征。抗体可以中和SEB，本研究假设针对不同中和位点的小鼠单克隆抗体组合可以协同作用。最有效的人-鼠嵌合形式的组合将被改造，以产生在人类中耐受性更好的抗体。

DESCRIPTION (provided by applicant): Synergistic combinations of chimeric anti-SEB: 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. 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. 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. The proposal has the following specific aims: 1. Generate neutralizing antibodies against the MHC binding site of SEB and derive neutralizing antibodies specific for the TCR beta binding site. 2. Identify neutralizing antibodies specific for different epitopes of SEB and determine if combinations of these antibodies synergistically inhibit SEB. 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. 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. 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.