PROTEOLYTIC ANTIBODIES FOR BIODEFENSE

用于生物防御的蛋白水解抗体

• 批准号: 7538168
• 负责人: ROBERT F BALINT
• 金额: $ 30万
• 依托单位: CYTODESIGN, INC.
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-06-01 至 2009-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7538168
• 关键词: Accounting; Address; Animals; Antibodies; Antibody Formation; Antigen Targeting; Antigens; Binding; Bioterrorism; Bontoxilysin; Botulism; Catalytic Domain; Categories; Cells; Centers for Disease Control and Prevention (U.S.); Class; Cleaved cell; Clinic; Complementarity Determining Regions; Disease; Dose; Engineering; Epitopes; Escherichia coli; Fab Immunoglobulins; Facility Construction Funding Category; Factor VIII; Fd fragments; Germ Lines; Human; Hydrolysis; Immune; Libraries; Light; Link; Monoclonal Antibodies; Pharmaceutical Preparations; Pharmacologic Substance; Phase; Phenotype; Production; Public Health; Rate; Reporter; Resistance; Risk; Safety; Serine Protease; Serotyping; Structure; System; Therapeutic; Therapeutic Uses; Time; Toxin; Triad Acrylic Resin; Work; antibody engineering; base; biodefense; cost; cross reactivity; desire; pre-clinical; prevent; research clinical testing; sensor; small molecule; therapeutic target; tool

DESCRIPTION (provided by applicant): Antibodies have shown great promise in the clinic for neutralizing a wide variety of target antigens. However, significant limitations have prevented monoclonal antibodies (mAbs) from realizing their full therapeutic potential. Antibody target neutralization and effector functions work best when multiple epitopes are targeted. Thus, mAbs have sub-optimal potency for many therapeutic applications. Oligoclonal combinations of mAbs targeting multiple epitopes have shown more promise, however, large doses are generally required, effector functions are compromised by the need for non-human hosts, and production costs are prohibitive for many applications. These limitations could be overcome by replacing conventional antibody effector function with proteolytic activities against target antigens. Proteolytic antibodies have been observed naturally, and certain disease states provoke the formation of antibodies with physiologically significant proteolytic activities, suggesting the potential efficacy of proteolytic antibodies in the clinic. The ability to harness the catalytic power of proteolytic antibodies for therapeutic uses could provide more potent pharmaceuticals than conventional antibodies that could be effective at lower doses. A third of the human germline kappa light chain variable domain (V() repertoire contain canonical serine protease-like catalytic triads in their complementarity- determining regions (CDR) and many have detectable peptidolytic activity. The tools of antibody engineering allow the construction of large libraries based on these human V( structures, which may contain effective proteolytic activities against a broad spectrum of therapeutic targets. What is lacking is a robust system for direct selection of desired activities from such libraries. To address this need, a proprietary cell-based system has been developed for selecting target-specific proteolytic activities with potentially unprecedented catalytic efficiencies from large libraries of proteolytic V( domains. The selection system comprises a proteolytic activity sensor in which an auto-inhibited reporter is linked to the target in such a way that cleavage of the target leads to activation of the reporter. When the sensor is expressed in E. coli cells along with a proteolytic V( library, cells expressing target-cleaving V( domains can be identified by the selectable phenotype conferred on the cells by the activated reporter. The system will be used to select and optimize human antibody light chains and Fab fragments containing V( domains with high proteolytic activity against Botulinum neurotoxin (BoNT). BoNTs are the most potent known toxins, and have been classified by the CDC as one of the six highest-risk agents for bioterrorism. Currently there are no effective small-molecule drugs for botulism, and due to the extreme lethality of BoNTs, it has not been possible to produce mAbs of adequate potency. In Phase I, human antibody Fab fragments that inactivate the catalytic subunit of BoNT will selected and optimized. In Phase II, the most active Fabs from Phase I will be subjected to pre-clinical evaluation in animals for both pre- and post- exposure safety and efficacy. PUBLIC HEALTH RELEVANCE: Monoclonal antibodies have shown great promise in the clinic, but significant limitations have prevented them from realizing their full therapeutic potential, including sub-optimal potencies and high costs. These limitations could be overcome by replacing conventional antibody effector function with proteolytic activities against target antigens. In this project, a system which has been developed for engineering proteolytic activity into antibodies will be applied to developing proteolytic antibodies against Botulinum neurotoxin, the most potent known toxin, and one of the greatest bioterrorism threats, for which there are no adequate treatments at present.

描述（由申请人提供）：抗体在临床上显示出中和多种靶抗原的巨大前景。然而，显着的局限性阻碍了单克隆抗体 (mAb) 充分发挥其治疗潜力。当靶向多个表位时，抗体靶标中和和效应子功能效果最佳。因此，单克隆抗体对于许多治疗应用来说效力不佳。针对多个表位的单克隆抗体寡克隆组合已显示出更大的前景，然而，通常需要大剂量，效应器功能因对非人类宿主的需要而受到损害，并且生产成本对于许多应用来说是令人望而却步的。这些限制可以通过用针对靶抗原的蛋白水解活性取代传统的抗体效应器功能来克服。蛋白水解抗体已在自然中被观察到，某些疾病状态会引发具有生理学显着蛋白水解活性的抗体的形成，这表明蛋白水解抗体在临床中的潜在功效。利用蛋白水解抗体的催化能力进行治疗的能力可以提供比传统抗体更有效的药物，并且可以在较低剂量下发挥作用。三分之一的人种系 kappa 轻链可变结构域 (V() 库在其互补决定区 (CDR) 中包含典型的丝氨酸蛋白酶样催化三联体，并且许多具有可检测的肽分解活性。抗体工程工具允许构建基于这些人 V( 结构的大型文库，其中可能包含针对广谱治疗靶点的有效蛋白水解活性。什么是 缺乏一个强大的系统来从此类库中直接选择所需的活动。为了满足这一需求，开发了一种专有的基于细胞的系统，用于从大型蛋白水解 V( 结构域库中选择具有潜在前所未有的催化效率的靶标特异性蛋白水解活性。选择系统包含一个蛋白水解活性传感器，其中自动抑制报告基因与靶标的连接方式如下： 靶标的裂解导致报告基因的激活。 When the sensor is expressed in E. coli cells along with a proteolytic V( library, cells expressing target-cleaving V( domains can be identified by the selectable phenotype conferred on the cells by the activated reporter. The system will be used to select and optimize human antibody light chains and Fab fragments containing V( domains with high proteolytic activity against Botulinum 神经毒素（BoNT）。 BoNT 是已知最有效的毒素，并已被 CDC 列为六种最高风险的生物恐怖主义制剂之一。目前还没有有效的小分子药物来治疗肉毒杆菌中毒，并且由于肉毒杆菌毒素的极端致死性，还不可能生产出足够效力的单克隆抗体。在第一阶段，人类抗体 Fab 片段使催化失活 BoNT的亚基将被选择和优化。在第二阶段，第一阶段最活跃的 Fab 将在动物身上进行临床前评估，以评估暴露前和暴露后的安全性和有效性。 PUBLIC HEALTH RELEVANCE: Monoclonal antibodies have shown great promise in the clinic, but significant limitations have prevented them from realizing their full therapeutic potential, including sub-optimal potencies and high costs.这些限制可以通过用针对靶抗原的蛋白水解活性取代传统的抗体效应器功能来克服。在该项目中，已开发的用于将蛋白水解活性转化为抗体的系统将用于开发针对肉毒杆菌神经毒素的蛋白水解抗体，肉毒杆菌神经毒素是已知最有效的毒素，也是最大的生物恐怖主义威胁之一，目前尚无适当的治疗方法。