Do peptide mimetics of gp41 improve antibody-epitope interactions?

gp41 的肽模拟物是否可以改善抗体-表位相互作用？

基本信息

批准号：
8624504
负责人：
Vincent Joseph Venditto
金额：
$ 5.39万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-03-01 至 2015-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8624504
关键词：
Acquired Immunodeficiency Syndrome Address Affinity Alkanesulfonates Amino Acids Animals Antibodies Antibody Affinity Antibody Formation Antigens Aspartic Acid Autoimmune Process Binding Biological Assay Cells Chemicals Communicable Diseases Cysteic Acid Cysteine Data Development Drug Formulations Enzyme-Linked Immunosorbent Assay Epidemic Epitopes Evaluation Exhibits Extravasation Failure Freund&apos s Adjuvant Future Glutamic Acid HIV HIV Infections HIV vaccine HIV-1 Immune Sera Immune response Immunity Immunization Infection Inflammation Infusion procedures Irrigation Lead Lipid Bilayers Lipids Liposomes Mediating Membrane Modeling Modification Monoclonal Antibodies Mucosal Immune Responses Mucosal Immunity Mus Mutation Oryctolagus cuniculus Patients Peptides Phospholipids Phosphorylation Population Positioning Attribute Post-Translational Protein Processing Primates Proteins Reaction Sampling Serine/Threonine Phosphorylation Serum Site Structure-Activity Relationship Surface Testing Tyrosine Vaccinated Vaccines Viral Load result Virus Walkers Work abstracting analog base clinically relevant cross reactivity design env Gene Products immunogenic immunogenicity immunoreactivity improved in vitro Assay inorganic phosphate mimetics monophosphoryl lipid A neonatal Fc receptor neutralizing antibody neutralizing monoclonal antibodies nitration novel phosphonate polyclonal antibody prevent protein aminoacid sequence receptor binding success vaccine development

项目摘要

Do peptide mimetics of gp41 improve antibody-epitope interactions? Abstract In this F32 application I postulate that the broadly HIV neutralizing monoclonal antibodies, 2F5 and 4E10, against the membrane proximal external region (MPR) of gp41 were induced in a small population of donors by MPR epitopes due to modifications in the envelope proteins. My idea is to generate MPR immunogens incorporating unnatural amino acids that may induce stronger immune responses. I believe that these novel peptides will induce stronger affinity toward monoclonal antibodies, 2F5 and 4E10, and potentially induce higher titer polyclonal antibodies in a prime-boost immunization assay. The idea to use unnatural amino acids is unique. MPR immunogen design has focused on eliciting antibody responses that cross-react with phospholipids because the mAbs 2F5 and 4E10 exhibit unusual cross-reactivity with phospholipids. Incorporation of chemically modified amino acids into known epitopes may induce stronger immune responses. I will synthesize a group of MPR-based peptide sequences containing modified residues, which will be used to determine antibody affinity using a BIAcore assay. Structure activity relationships will be derived from these data and will guide subsequent rounds of chemically modified peptides to find sequences that will bind optimally to the monoclonal antibodies. I will then determine the immunogenicity of each peptide in rabbits using a prime-boost immunization strategy. Lipopeptides based on the peptides used in the BIAcore assay will be presented in a liposomal formulation to further enhance immunogenicity by presenting the epitopes in a lipid bilayer. Rabbits will be immunized subcutaneously and intramuscularly and antisera from these animals will be obtained and evaluated in a validated ELISA assay to determine the antibody titer induced by each peptide immunogen. Furthermore, formulations containing the same lipopeptides will be delivered intranasally to evaluate the mucosal immune response. Formulations will be prepared in the presence and absence of a peptide that binds and is transported by the neonatal Fc receptor (FcRn). ELISA assays will be used to determine the antibody responses in serum from SC/IM immunized animals, while mucosal lavage and serum samples will be evaluated from IN immunized animals. Differences in antisera titers from SC/IM immunized animals will be compared to IN immunized animals. Antisera and mucosal lavage samples with high titers in rabbits will be used in traditional inhibition assays to determine the neutralization activity of primary HIV-1 isolates in PBMCs. Success in generating neutralizing antibodies in rabbits will lead to future work to generate monoclonal antibodies. The chemically modified peptides that induce neutralizing antibodies in rabbits could provide a widely applicable epitope design strategy for the development of vaccines to a variety of infectious diseases including HIV.

gp41 的肽模拟物是否可以改善抗体-表位相互作用？抽象的在这个 F32 应用中，我假设，由于包膜蛋白的修饰，MPR 表位在一小群供体中诱导了针对 gp41 近膜外部区域 (MPR) 的广泛 HIV 中和单克隆抗体 2F5 和 4E10。我的想法是产生包含非天然氨基酸的 MPR 免疫原，可以诱导更强的免疫反应。我相信这些新型肽将诱导对单克隆抗体、2F5 和 4E10 更强的亲和力，并可能在初免-加强免疫测定中诱导更高滴度的多克隆抗体。使用非天然氨基酸的想法是独特的。 MPR 免疫原设计的重点是引发与磷脂发生交叉反应的抗体反应，因为 mAb 2F5 和 4E10 表现出与磷脂不同寻常的交叉反应性。将化学修饰的氨基酸掺入已知表位可能会诱导更强的免疫反应。我将合成一组包含修饰残基的基于 MPR 的肽序列，这些序列将用于使用 BIAcore 测定法确定抗体亲和力。结构活性关系将从这些数据中得出，并将指导后续几轮化学修饰肽找到与单克隆抗体最佳结合的序列。然后，我将使用初免-加强免疫策略确定每种肽在兔子中的免疫原性。基于 BIAcore 测定中使用的肽的脂肽将以脂质体制剂的形式呈现，通过在脂质双层中呈现表位来进一步增强免疫原性。对兔子进行皮下和肌内免疫，从这些动物中获得抗血清，并在经过验证的 ELISA 测定中进行评估，以确定每种肽免疫原诱导的抗体滴度。此外，含有相同脂肽的制剂将被鼻内递送以评估粘膜免疫反应。将在存在或不存在与新生儿 Fc 受体 (FcRn) 结合并由其转运的肽的情况下制备制剂。 ELISA 测定将用于确定 SC/IM 免疫动物血清中的抗体反应，同时将评估 IN 免疫动物的粘膜灌洗液和血清样本。将 SC/IM 免疫动物的抗血清滴度差异与 IN 免疫动物进行比较。兔子体内高滴度的抗血清和粘膜灌洗样品将用于传统的抑制测定，以确定 PBMC 中初级 HIV-1 分离株的中和活性。在兔子中成功产生中和抗体将导致未来产生单克隆抗体的工作。在兔子体内诱导中和抗体的化学修饰肽可以为开发包括艾滋病毒在内的多种传染病的疫苗提供广泛适用的表位设计策略。