Optimization of mAbs to staphylococcal enterotoxin B for treatment

针对葡萄球菌肠毒素 B 的单克隆抗体的优化治疗

• 批准号: 7670783
• 负责人: Bettina Fries
• 金额: $ 52.95万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2014-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7670783
• 关键词: Affinity; Animal Model; Arts; B-Lymphocytes; Binding; Biological Assay; C-terminal; Collaborations; Dose; Engineering; Enzyme-Linked Immunosorbent Assay; Epitopes; Funding; Goals; Human; Immunoglobulin Constant Region; Immunoglobulin Variable Region; Infection; Knowledge; Light; Measures; Monoclonal Antibodies; Mus; Patients; Proteins; Reagent; Research Personnel; Serum; Shock; Staphylococcal Enterotoxin B; Staphylococcus aureus; T-Lymphocyte; Techniques; Technology; Testing; Therapeutic Uses; Time; Toxin; Treatment Protocols; Work; base; biodefense; cost; design; improved; in vivo; novel therapeutics; research study; Affinity; Animal Model; Arts; B-Lymphocytes; Binding; Biological Assay; C-terminal; Collaborations; Dose; Engineering; Enzyme-Linked Immunosorbent Assay; Epitopes; Funding; Goals; Human; Immunoglobulin Constant Region; Immunoglobulin Variable Region; Infection; Knowledge; Light; Measures; Monoclonal Antibodies; Mus; Patients; Proteins; Reagent; Research Personnel; Serum; Shock; Staphylococcal Enterotoxin B; Staphylococcus aureus; T-Lymphocyte; Techniques; Technology; Testing; Therapeutic Uses; Time; Toxin; Treatment Protocols; Work; base; biodefense; cost; design; improved; in vivo; novel therapeutics; research study

This application will bring together the expertise of different investigators and incooperate state of the art platform technologies with the goal to optimize monoclonal antibodies (mAb) to Staphylococcal enterotoxin B (SEE) for human use. Beyond the immediate benefit of yielding a novel therapeutic reagent, this work will investigate several hypotheses that pertain to toxin-neutralization and clearance. The underlying working hypothesis of our proposal is that high affinity anti-SEB mAbs are best suited for therapeutic use because low amounts of mAb will be sufficient to successfully compete with already bound SEB. We will investigate if human mAbs differ and are superior in their ability to neutralize toxin, In addition we propose that targeted engineering of the C-terminal with effects on FcR binding will change toxin clearance in vivo. In the past funding period 11 mAbs to SEB have been subcloned, characterized and tested in murine animal model for SEB induced shock. A highly sensitive capture ELISA was developed that will allow us in proposed experiments to measure SEB toxin in serum of intoxicated mice. In addition the prevalance of SEB producing S. aureus strains has been established and identified important sequence variabilities in different S. aureus strains which will be taken into consideration. Three lgG1 candidate mAbs have been selected for targeted Ab engineering. They recognize epitopes in the C-terminal part of the protein, which is also the predominant epitope recognized by human B-cells. Protection was documented in animal models and human T-cell stimulation assays. In addition heavy and light chain variable regions of the mAbs were cloned. In Aim one we propose to optimize these mAbs by affinity maturation in collaboration with Dr. Scharff. In Aim 2 we propose to generate new human SEB apecific mAbs one involving a genetically modified mouse that expresses the human variable region and the other one we will use Drs. Ahmed/Wilson's technique to directly clone the variable region from patients with primary S. aureus infection. Finally in aim 3 we will engineer the FC binding portion of the mAbs in collaboration with Dr. Ravetch and test the intriguing hypothesis that combination of mAbs enhances neutralization and clearance.

该申请将汇集不同的研究人员的专业知识，并融合了艺术状态 平台技术的目标是将单克隆抗体（MAB）优化为葡萄球菌肠毒素B （请参阅）使用人类。除了产生一种新颖的治疗试剂的直接利益之外，这项工作还将 研究几种与毒素中和化学有关的假设。基础工作 我们提出的假设是高亲和力抗seb mab最适合治疗，因为 低量的mAB将足以成功与已经绑定的SEB竞争。我们将调查是否 人物mAB的不同，并且在中和毒素的能力上是优越的，此外，我们提出了针对性的 C末端对FCR结合的影响的工程将在体内改变毒素清除率。在过去 在鼠类动物模型中，已将11个单克重的筹集时间纳入了SEB的表征 SEB引起的冲击。开发了高度敏感的捕获ELISA 测量陶醉小鼠血清中SEB毒素的实验。此外，SEB产生的流行 金黄色葡萄球菌菌株已经建立并确定了不同金黄色葡萄球菌的重要序列变化 将考虑的菌株。已选择了三个LGG1候选mAb以进行针对性 AB工程。他们识别蛋白质的C末端部分的表位，这也是主要的 人类B细胞认可的表位。在动物模型和人类T细胞中记录了保护 刺激测定。另外，将单元素的重链变量区域克隆。在目标中 我们建议通过与Scharff博士合作，通过亲和力成熟来优化这些mAB。在目标2中我们 建议生成新的人类SEB Apecific mabs，其中涉及一种转基因的小鼠 表示人类可变区域，而我们将使用DRS的另一个区域。艾哈迈德/威尔逊的技术 直接从原发性金黄色葡萄球菌感染的患者中直接克隆可变区域。终于在目标3中 工程师与Mabs的FC装订部分与Ravetch博士合作，并测试有趣的 MAB组合可以增强中和和清除率的假设。