Trispecific monoclonal antibody for botulinum neurotoxin intoxication therapy

用于肉毒杆菌神经毒素中毒治疗的三特异性单克隆抗体

• 批准号: 8469825
• 负责人: James D. Marks
• 金额: $ 20.1万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-06-01 至 2014-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8469825
• 关键词: Affinity; Allergic Reaction; Amino Acid Sequence; Antibodies; Antibody Therapy; Antitoxins; Award; Binding; Binding Sites; Biological Assay; Biotechnology; Bioterrorism; Blood Circulation; Bontoxilysin; Botulinum Toxin Type A; Botulism; Clinical; Clinical Trials; Contracts; Cyclic GMP; Drug Kinetics; Epitopes; Equus caballus; Event; FDA approved; Funding; Goals; Grant; Half-Life; Human; Immunoglobulin Variable Region; Incidence; Individual; Intensive Care; Intoxication; Monoclonal Antibodies; Mus; Pharmaceutical Preparations; Phase I Clinical Trials; Positioning Attribute; Prevention; Production; Property; Proteins; Quality Control; Recombinant Antibody; Relative (related person); Serotyping; Serum; Testing; Therapeutic Effect; Toxic effect; Toxin; Work; base; biothreat; botulinum toxin type B; botulinum toxin type E; cost; cost effective; high risk; human disease; in vivo; neutralizing monoclonal antibodies; safety testing; stoichiometry; Affinity; Allergic Reaction; Amino Acid Sequence; Antibodies; Antibody Therapy; Antitoxins; Award; Binding; Binding Sites; Biological Assay; Biotechnology; Bioterrorism; Blood Circulation; Bontoxilysin; Botulinum Toxin Type A; Botulism; Clinical; Clinical Trials; Contracts; Cyclic GMP; Drug Kinetics; Epitopes; Equus caballus; Event; FDA approved; Funding; Goals; Grant; Half-Life; Human; Immunoglobulin Variable Region; Incidence; Individual; Intensive Care; Intoxication; Monoclonal Antibodies; Mus; Pharmaceutical Preparations; Phase I Clinical Trials; Positioning Attribute; Prevention; Production; Property; Proteins; Quality Control; Recombinant Antibody; Relative (related person); Serotyping; Serum; Testing; Therapeutic Effect; Toxic effect; Toxin; Work; base; biothreat; botulinum toxin type B; botulinum toxin type E; cost; cost effective; high risk; human disease; in vivo; neutralizing monoclonal antibodies; safety testing; stoichiometry

DESCRIPTION (provided by applicant): The goal of this project is to generate highly potent trispecific antibodies for the prevention and treatment of botulism due to botulinum neurotoxins (BoNT) serotypes A, B, E, and F. BoNTs are one of the six highest-risk threat agents for bioterrorism, due to their extreme potency and lethality, ease of production, and need for prolonged intensive care. Equine antitoxin has a high incidence of allergic reactions, a short serum half-life leading to reintoxication, is challenging to administer, and cannot be given prophylactically. Given the extraordinary toxicity of BoNTs, antitoxin must be of very high potency. Previous studies have found that no single mAb neutralizes BoNT with the requisite potency, however, combining mAbs binding non-overlapping epitopes leads to highly potent BoNT neutralization due to rapid clearance of BoNT from the circulation. We have generated combinations of three mAbs that bind all subtypes of BoNT/A, B, or E and result in highly potent BoNT neutralization in vivo. Based on the potency, HHS has awarded contracts to a biotechnology company (XOMA (US) LLC) to develop these mAb combinations for clinical use. The BoNT/A three mAb combination has entered phase 1 clinical trials. The BoNT/B and BoNT/E three mAb combinations are in cGMP manufacturing and will enter clinical trials by 2013-2014. A challenge of this approach is the large number of mAbs that need to be manufactured and quality controlled (3 per serotype). This adds to the cost and complexity of the product, especially if the mAbs are all going to be combined to create a multi-serotype drug. Our proposed alternative is to create a single trispecific mAb for each serotype incorporating the binding sites of each of the three mAbs. For this work, we hypothesize that such trispecific mAb can be generated which will have the same binding and BoNT neutralization properties as the three mAb combination. In the R21 portion we will construct trispecific antibodies (TsAbs) binding three non-overlapping BoNT/A epitopes. Four different TsAbs will be constructed that differ with respect to where each of the three variable domains are positioned relative to the antibody Fc. Each TsAb will be characterized and optimized with respect to affinity for each of the three epitopes, stoichiometry of binding to BoNT/A, stability, and ease of expression and purification. Multiple TsAb constructs will be evaluated for in vivo pharmacokinetics (PK) and BoNT/A clearance and neutralization potency in the mouse neutralization assay. In vivo studies will be conducted at the USDA by our long-term collaborator, Dr. Luisa Cheng. These studies will identify those constructs with optimal properties and will result in a single mAb that potentl neutralizes BoNT/A. In the R33 portion of this grant, the optimal construct identified for BoNT/A will be applied to three other BoNT serotypes, B, E and F. The result of this project will be antibodies that would treat over 99% of human botulism cases.

DESCRIPTION (provided by applicant): The goal of this project is to generate highly potent trispecific antibodies for the prevention and treatment of botulism due to botulinum neurotoxins (BoNT) serotypes A, B, E, and F. BoNTs are one of the six highest-risk threat agents for bioterrorism, due to their extreme potency and lethality, ease of production, and need for prolonged intensive care.马抗毒素的过敏反应发生率很高，这是一种短血清半衰期，导致重新毒性，这是挑战性的，不能预防性地给出。鉴于BONT的特殊毒性，抗毒素必须具有很高的效力。先前的研究发现，没有单个mAb以必要的效力中和骨，但是，由于循环中的循环迅速清除了BONT，因此将MAB结合非重叠表位的MAB会导致高度有效的BONT中和。我们已经生成了三个mab的组合，它们结合了BONT/A，B或E的所有亚型，并导致体内高效的BONT中和。根据该效力，HHS已将合同授予生物技术公司（XOMA（US）LLC），以开发这些MAB组合供临床使用。 BONT/A三MAB组合已进入1期临床试验。 BONT/B和BONT/E三个mAb组合在CGMP制造中，并将在2013 - 2014年之前进行临床试验。这种方法的挑战是需要制造和控制质量的大量mAB（每个血清型3）。这增加了产品的成本和复杂性，尤其是如果将单克隆变量全部组合在一起以创建多种型药物时。我们提出的替代方法是为每个血清型创建一个单个特异性mAb，并结合了三个mAb的每个MAB的结合位点。对于这项工作，我们假设可以生成这种特异性mAb，该单元具有与三个mAb组合相同的结合和骨骼中和特性。在R21部分中，我们将构建三个非重叠BONT/A表位的三个特异性抗体（TSAB）。将构建四个不同的TSAB，这些TSAB相对于抗体FC的位置而言，这些TSAB与位置相对于位置中的每个TSAB。每个TSAB都将针对三个表位中的每个表位的亲和力进行表征和优化，结合到BONT/A的化学计量，稳定性以及表达和纯化的易感性。将评估多个TSAB构建体的体内药代动力学（PK）和小鼠中和测定中的清除和中和效力。我们的长期合作者Luisa Cheng博士将在USDA进行体内研究。这些研究将识别具有最佳特性的构造，并导致单个mAb的电位中和bont/a。在该赠款的R33部分中，确定的BONT/A的最佳结构将应用于其他三种BONT血清型B，E和F。该项目的结果将是将超过99％的人类肉毒杆菌病例治疗的抗体。