Rapid isolation of high-affinity human antibodies from large synthetic libraries

从大型合成文库中快速分离高亲和力人类抗体

• 批准号: 7803512
• 负责人: MATTHEW P DELISA
• 金额: $ 20万
• 依托单位: VYBION, INC.
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-02-01 至 2011-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7803512
• 关键词: Accounting; Affinity; Antibodies; Antibody Affinity; Antibody Therapy; Antigen Targeting; Antigens; Arginine; Binding; Biological; Biotechnology; Cell surface; Cells; Clinical; Clinical Trials; Collection; Complex; Culture Media; Cytoplasm; Development; Diagnostics Research; Disease; Engineering; Ensure; Environment; Enzymes; Evolution; Future; Genes; Goals; Health; Human; Immobilization; Immunization; Immunoglobulin Fragments; Immunotoxins; Inflammatory; Lactamase; Libraries; Light; Malignant Neoplasms; Marketing; Membrane; Methods; Molecular; Monoclonal Antibodies; Nature; Neurodegenerative Disorders; Pathway interactions; Peptide Hydrolases; Peptides; Phage Display; Pharmaceutical Preparations; Production; Property; Protein Export Pathway; Proteins; Quality Control; Reagent; Recombinant Antibody; Recombinant Proteins; Relative (related person); Reporter; Research; Resistance; Screening procedure; Series; Solid; Specificity; Staging; Structure; Surface; System; Techniques; Technology; Twin Multiple Birth; Virus Diseases; Yeasts; antigen antibody binding; antigen binding; base; combinatorial; commercial application; cost; design; directed evolution; drug development; genetic selection; human disease; innovation; interest; new technology; novel; novel therapeutics; periplasm; polypeptide; pre-clinical; protein expression; protein folding; public health relevance; research study; secretory protein; success; tat Protein; therapeutic protein; therapeutic target; tool

DESCRIPTION (provided by applicant): With over 300 therapeutic proteins currently in various stages of clinical trials, the road to a healthier future will require new methods for producing safer and less expensive recombinant proteins. In particular, next generation therapeutics derived from monoclonal antibodies (e.g. Fab, scAb, scFv, immunotoxins, etc) show great clinical promise in treating a range of human disorders including bacterial and viral infections, cancer, inflammatory diseases and neurodegenerative disorders. Currently, the bacterial Sec protein export pathway is the dominant mechanism for protein display (e.g., phage display, cell surface display) and also for the expression and engineering of small, non-glycosylated antibody fragments. However, due to a number of limitations associated with Sec export, bacterial production of stable, high-affinity antibody fragments in high yields for preclinical and clinical trials can be a serious bottleneck in the antibody drug pipeline. Therefore, the overall objective of the proposal is to develop a versatile antibody expression and engineering platform by capitalizing on the remarkable properties of the recently discovered bacterial twin-arginine translocation (Tat) pathway. Towards this objective, the current proposal encompasses the following specific aims: (1) creation of synthetic libraries of single-chain Fv (scFv) antibody fragments; and (2) application of a proprietary Tat-based genetic selection strategy for efficient isolation of super-stable, ultra- high affinity human scFvs against disease-relevant antigens. The proposed studies are expected to result in a rapid, single-step selection platform for the discovery of scFv sequences that fold and function in intra- or extra-cellular environments. Successful completion of the proposed studies is expected to result in a powerful new technology for generating potent antibody-based biopharmaceuticals, diagnostics and research reagents. PUBLIC HEALTH RELEVANCE: By 2008, engineered antibody fragments are predicted to account for >30% of all revenues in the biotechnology market and will be used to treat a wide array of human diseases including bacterial and viral infections, cancer, inflammatory diseases and neurodegenerative disorders. Since antibody therapies are an increasingly large fraction of the drugs in development, with ever escalating increases in the cost of drug development, any improvements to the production or discovery of efficacious antibody fragments will have a significant impact on human health. Thus, this proposal seeks to develop a novel bacterial selection strategy for rapid, low-cost isolation of potent human antibodies against virtually any target antigen of interest.

描述（由申请人提供）：目前有300多个治疗蛋白处于临床试验的各个阶段，通往更健康的未来的道路将需要新的方法来生产更安全且较便宜的重组蛋白。特别是，源自单克隆抗体（例如Fab，SCAB，SCFV，免疫毒素等）的下一代治疗剂在治疗一系列人类疾病（包括细菌和病毒感染），包括癌症，炎性疾病，炎症性疾病和神经减退性疾病方面显示出巨大的临床前景。当前，细菌SEC蛋白出口途径是蛋白质显示的主要机制（例如噬菌体显示，细胞表面显示），也是小型非糖基化抗体片段的表达和工程。但是，由于与SEC出口相关的许多局限性，用于临床前试验和临床试验的高产量的稳定，高亲和力抗体片段的细菌可能是抗体药物管道中的严重瓶颈。因此，该提案的总体目标是通过利用最近发现的细菌双精氨酸易位（TAT）途径的显着特性来开发一种多功能抗体表达和工程平台。为了实现这一目标，当前的提案包括以下特定目的：（1）创建单链FV（SCFV）抗体片段的合成库； （2）将基于TAT的专有遗传选择策略应用于有效地分离出与疾病相关的抗原的超稳定性超高亲和力人类SCFV。预计拟议的研究将导致一个快速的单步选择平台，以发现在细胞内或细胞外环境中折叠和功能的SCFV序列。预计拟议的研究的成功完成将导致一项强大的新技术，用于产生有效的基于抗体的生物药物，诊断和研究试剂。 公共卫生相关性：到2008年，预计工程抗体碎片将占生物技术市场中所有收入的30％，并将用于治疗多种人类疾病，包括细菌和病毒感染，癌症，炎症性疾病，炎症性疾病和神经退行性疾病。由于抗体疗法越来越大的开发药物，随着药物开发成本的不断提高，对生产或发现有效抗体片段的任何改善都会对人类健康产生重大影响。因此，该提案旨在制定一种新型的细菌选择策略，以快速，低成本的人类抗体几乎对任何目标抗原的抗原进行快速隔离。

项目成果

Engineering antibody fitness and function using membrane-anchored display of correctly folded proteins.

• DOI: 10.1016/j.jmb.2011.12.021
• 发表时间: 2012-02-10
• 期刊: JOURNAL OF MOLECULAR BIOLOGY
• 影响因子: 5.6
• 作者: Karlsson, Amy J.;Lim, Hyung-Kwon;Xu, Hansen;Rocco, Mark A.;Bratkowski, Matthew A.;Ke, Ailong;DeLisa, Matthew P.
• 通讯作者: DeLisa, Matthew P.