A MERS-CoV Receptor Decoy

MERS-CoV 受体诱饵

• 批准号: 8981265
• 负责人: KEITH WYCOFF
• 金额: $ 80.18万
• 依托单位: PLANET BIOTECHNOLOGY, INC.
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-06-01 至 2017-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8981265
• 关键词: Active Sites; Affinity; Amino Acids; Antibodies; Binding; Biotechnology; Camels; Case Fatality Rates; Cell Surface Proteins; Cells; Cessation of life; Contracts; Coronavirus; Coronavirus Infections; Coronavirus spike protein; Country; Development; Dipeptidyl Peptidases; Dose; Drug Kinetics; Enzyme-Linked Immunosorbent Assay; Evaluation; Extracellular Domain; Fc Immunoglobulins; Glucose; Half-Life; Health; Human; Hydrolase; IgA1; IgA2; IgG1; Immunoglobulin Isotypes; Immunoglobulins; Infection; Laboratories; Lead; Link; Middle East; Middle East Respiratory Syndrome Coronavirus; Modeling; Mus; Mutate; Mutation; Peptide Hydrolases; Phase; Physiological Processes; Planets; Plants; Polysaccharides; Process; Production; Protein Binding; Protein S; Proteins; Rattus; Recombinant Proteins; Route; Safety; Serum; Site; Surface; Syndrome; System; Testing; Toxicology; Variant; Virus; base; coronavirus receptor; design; efficacy testing; epidemiology study; improved; in vivo; inhibitor/antagonist; intraperitoneal; novel therapeutics; pandemic disease; phase 1 study; phase 2 study; protective efficacy; public health relevance; receptor; respiratory; scale up; transmission process

 DESCRIPTION (provided by applicant): Middle East respiratory syndrome coronavirus (MERS-CoV) is a newly emerging human health threat with a ~35% case fatality rate. MERS-CoV uses dipeptidyl peptidase 4 (DPP4), a cell surface protein, to enter and infect cells. During our Phase I study, using a transient plant expression system, we produced fusions of human DPP4 and the human immunoglobulin Fc sequences of IgG1, IgA1 and IgA2, to produce "receptor decoys" to block cellular infection with MERS-CoV. We demonstrated that DPP4-Fc binds to the S1 domain of MERS-CoV S protein, and that DPP4-Fc is a more potent inhibitor of MERS-CoV cellular infection than soluble DPP4. We showed that a DPP4-Fc fusion based on IgA1 might possibly be more effective than one based on IgG1. In addition, we demonstrated that binding and virus neutralization could be improved by more than 10-fold by modifying a single amino acid where human DPP4 and MERS-CoV spike protein contact. DPP4-Fc is also expected to have superior pharmacokinetics, as Fc will confer a long circulating half-life and the ability to be delivered to airway mucosal surfaces, the site of MERS-CoV infection. In a phase II study, we will produce new DPP4-Fc constructs to improve DPP4's affinity for MERS-CoV spike protein, and eliminate DPP4's peptidase activity by mutating the active site. We will optimize new constructs to improve expression in plants and to produce DPP4-Fc with human-like N-linked glycans. New DPP4-Fc fusions will be ranked for MERS-CoV spike protein-binding by ELISA and tested for the ability to neutralize MERS-CoV infection in susceptible cells. The best performing DPP4-Fc variants will be tested for protective efficacy in a mouse MERS model, evaluating both intraperitoneal and intranasal routes of administration. We will scale up production and purification of our lead molecule to purify 7 grams of DPP4-Fc, which will be used for a pilot 2-week repeat-dose safety/toxicology study in rats. We will evaluate safety and quantify DPP4-Fc in serum to obtain pharmacokinetic parameters. We will also screen for anti-DPP4-Fc antibodies in these studies.

 描述（由申请人提供）：中东呼吸综合征冠状病毒（MERS-CoV）是一种新出现的人类健康威胁，病死率约为35%。MERS-CoV使用二肽基肽酶4（DPP 4），一种细胞表面蛋白，进入并感染细胞。在我们的I期研究期间，使用瞬时植物表达系统，我们产生了人DPP 4和IgG 1、IgA 1和IgA 2的人免疫球蛋白Fc序列的融合物，以产生“受体诱饵”来阻断MERS-CoV的细胞感染。我们证明了DPP 4-Fc与MERS-CoV S蛋白的S1结构域结合，并且DPP 4-Fc是比可溶性DPP 4更有效的MERS-CoV细胞感染抑制剂。我们表明，基于IgA 1的DPP 4-Fc融合体可能比基于IgG 1的DPP 4-Fc融合体更有效。此外，我们证明，通过修饰人DPP 4和MERS-CoV刺突蛋白接触的单个氨基酸，可以将结合和病毒中和提高10倍以上。还预期DPP 4-Fc具有上级药代动力学，因为Fc将赋予长的循环半衰期和递送至气道粘膜表面（MERS-CoV感染部位）的能力。在II期研究中，我们将生产新的DPP 4-Fc构建体，以提高DPP 4对MERS-CoV刺突蛋白的亲和力，并通过突变活性位点消除DPP 4的肽酶活性。我们将优化新的构建体，以改善在植物中的表达，并产生具有人样N-连接聚糖的DPP 4-Fc。将通过ELISA对新的DPP 4-Fc融合体的MERS-CoV刺突蛋白结合进行排名，并测试其在易感细胞中中和MERS-CoV感染的能力。将在小鼠MERS模型中测试表现最佳的DPP 4-Fc变体的保护效力，评价腹膜内和鼻内给药途径。我们将扩大先导分子的生产和纯化，以纯化7克DPP 4-Fc，用于大鼠中为期2周的重复给药安全性/毒理学试验。我们将评估安全性并定量血清中的DPP 4-Fc以获得药代动力学参数。我们还将在这些研究中筛选抗DPP 4-Fc抗体。