Antibody Engineering Program

抗体工程项目

• 批准号: 10262782
• 负责人: Mitchell Ho
• 金额: $ 58.26万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10262782
• 关键词: 2019-nCoV; Adult; Animals; Antibodies; Antibody Therapy; Antibody-drug conjugates; Antigen Receptors; Antigens; Award; Bacteria; Bacteriophages; Binding; Binding Proteins; CD276 gene; Camels; Cell Surface Receptors; Communicable Diseases; Communication; Communities; Complex; Consult; Development; Epitopes; Exhibits; Family; Funding; GPC3 gene; Generations; Glypican; Grant; Hepatology; Human; Hybridomas; Immunization; Immunize; Immunoglobulin Fragments; Immunoglobulin G; Immunoglobulins; Immunotoxins; Ion Channel; Knock-out; Knowledge; Laboratory Research; Libraries; Malignant Neoplasms; Malignant neoplasm of liver; Mammalian Cell; Manuscripts; Methods; Mus; NCI Center for Cancer Research; Names; National Cancer Institute; Nature; Nurses; Oryctolagus cuniculus; Phage Display; Preparation; Property; Proteins; Publishing; Recombinants; Reporting; Research; Research Personnel; Resources; Scientist; Services; Shark; Signal Transduction; Site; Source; Technology; Therapeutic; Therapeutic antibodies; Tumor Cell Line; United States National Institutes of Health; antibody engineering; antibody libraries; antigen binding; base; beta catenin; cancer cell; cancer immunotherapy; cancer therapy; chimeric antigen receptor T cells; drug candidate; human disease; immunogenic; innovation; interest; mesothelin; nanobodies; new technology; next generation sequencing; novel; preclinical development; programs; screening; technology development; tool; web site

While antibody-based therapeutics have emerged as a major component in cancer treatment, the generation of antibodies to important targets such as cell surface receptors and ion channels remains difficult. These proteins contain buried functional sites that are unreachable by classical mouse or human IgG-based antibodies. Single domain antibodies have shown a promising ability to target difficult antigens and hidden epitopes. Dr. Mitchell Ho at the NCI has demonstrated that single domain antibodies are capable of targeting buried functional sites in cancer signaling complexes [Feng et al. PNAS, 2013; Gao et al Nature Communications, 2015; Li et al. PNAS, 2017; Li et al. Hepatology, 2019]. The Ho lab has constructed large shark and camel single-domain ('nanobody') libraries and isolated binders to a wide range of antigens [Feng et al. Antibody Therapeutics, 2019], indicating that the phage-displayed single domain antibody libraries can be a valuable source to isolate therapeutic antibodies. In FY20, the AEP published two articles and collaborated in several other manuscripts in preparation. One is a review article that analyzes the shark library developed by the Ho lab [English et al. Antibody Therapeutics 2020; PMID: 32118195]. The other is a method article that describes the isolation of rabbit single domain antibodies for the cancer target B7-H3 (CD276) [Feng et al. Antibody Therapeutics 2020; PMID: 32166218]. The antigen binding variable domain (VNAR) of the shark immunoglobulin new antigen receptor (IgNAR) evolved approximately 500 million years ago and it is one of the smallest antibody fragments in the animal kingdom with sizes of 12-15 kDa. In the review article, the AEP evaluates the current knowledge of the shark VNAR single domain sequences and ongoing development of shark VNARs as research tools as well as potential therapeutics, in particular highlighting the recent next-generation sequencing analysis of 1.2 million shark VNAR sequences and construction of a large phage displayed shark VNAR library from six adult nurse sharks (Ginglymostoma cirratum) [English et al. Antibody Therapeutics 2020; PMID: 32118195]. The large phage-displayed VNAR single domain library covers all the four known VNAR types (Types I-IV) and many previously unknown types. Ongoing preclinical development will help define the utility of shark VNAR single domains as a potentially new family of drug candidates for treating cancer and other human diseases. Rabbit antibodies can recognize diverse epitopes, including those that are poorly immunogenic in mice and humans. The AEP has established a method to isolate rabbit VH single domain antibodies for potential cancer therapy [Feng et al. Antibody Therapeutics 2020; PMID: 32166218]. We immunized rabbits with recombinant human B7-H3 (CD276) protein, made a phage-displayed rabbit VH single domain library with a diversity of 109, and isolated two binders (A1 and B1) from phage panning. Both rabbit VH single domains exhibited antigen-dependent binding to B7-H3-positive tumor cell lines but not B7-H3 knockout tumor cell lines. Our study shows that protein immunization followed by phage display screening can be used to isolate rabbit single domain antibodies. The two single domain antibodies reported by the AEP may have potential applications in cancer immunotherapy. We are also supporting the development of neutralizing nanobodies against SARS-CoV-2.

虽然基于抗体的疗法已经成为癌症治疗的主要组成部分，但针对细胞表面受体和离子通道等重要靶点的抗体的产生仍然困难。这些蛋白质含有经典的鼠或人免疫球蛋白抗体所不能达到的埋藏的功能位点。单域抗体已显示出针对困难抗原和隐藏表位的前景看好的能力。NCI的Mitchell Ho博士已经证明了单域抗体能够靶向癌症信号复合体中埋藏的功能部位[冯等人]。PNAS，2013；高等人，2015；Li等人。PNAS，2017；Li等人。肝病，2019年]。Ho实验室已经构建了大型鲨鱼和骆驼单域(‘纳米体’)文库，并分离了多种抗原的结合剂[冯等人]。抗体治疗学，2019]，表明噬菌体展示的单域抗体库可以作为分离治疗性抗体的有价值的来源。在2020财年，AEP发表了两篇文章，并合作编写了其他几篇手稿。一篇是一篇评论文章，分析了Ho实验室开发的Shark库[English等人。抗体治疗，2020年；PMID：32118195]。另一篇是描述分离抗癌靶标B7-H3(CD276)的兔单域抗体的方法文章[冯等人。抗体治疗，2020年；PMID：32166218]。鲨鱼免疫球蛋白新抗原受体(IgNAR)的抗原结合可变区(VNAR)大约在5亿年前进化，是动物界中最小的抗体片段之一，大小为12-15 kDa。在这篇综述文章中，AEP评估了鲨鱼VNAR单域序列的当前知识、鲨鱼VNAR作为研究工具的持续发展以及潜在的治疗方法，特别是重点介绍了最近对120万条鲨鱼VNAR序列的下一代测序分析，以及从六只成年护士鲨(Ginglymostoma Cirratum)[English等人]构建的大型噬菌体展示鲨鱼VNAR文库。抗体治疗，2020年；PMID：32118195]。噬菌体展示的大型VNAR单域文库涵盖了所有四种已知的VNAR类型(类型I-IV)和许多以前未知的类型。正在进行的临床前开发将有助于确定鲨鱼VNAR单结构域的用途，作为治疗癌症和其他人类疾病的潜在新候选药物家族。兔抗体可以识别不同的表位，包括那些在小鼠和人类中免疫原性较差的表位。AEP已经建立了一种分离兔VH单域抗体的方法，用于潜在的癌症治疗[冯等人。抗体治疗，2020年；PMID：32166218]。我们用重组人B7-H3(CD276)蛋白免疫兔，构建了109个多样性的噬菌体展示兔VH单域文库，并从噬菌体淘洗中分离出两个结合子A1和B1。这两个兔VH单区均与B7-H3阳性肿瘤细胞呈抗原依赖性结合，但不能与B7-H3基因敲除肿瘤细胞结合。我们的研究表明，蛋白免疫和噬菌体展示筛选可用于兔单域抗体的分离。AEP报告的两个单域抗体可能在癌症免疫治疗中有潜在的应用。我们还支持开发针对SARS-CoV-2的中和纳米抗体。