HIV neutralizing antibodies induced by chemically modified MPR epitopes

由化学修饰的 MPR 表位诱导的 HIV 中和抗体

基本信息

批准号：
8146237
负责人：
FRANCIS C. SZOKA
金额：
$ 19.31万
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：
2011
资助国家：
美国
起止时间：
2011-08-15 至 2013-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8146237
关键词：
Acquired Immunodeficiency Syndrome Affinity Alkanesulfonates Amino Acids Animals Antibodies Antibody Formation Antigens Biological Assay Cells Charge Chemicals Circular Dichroism Communicable Diseases Complex Cysteine Drug Formulations Enzyme-Linked Immunosorbent Assay Epidemic Epitopes Exhibits Failure Future Generations HIV HIV Infections HIV vaccine Head Human Immune Sera Immune response Immune system Immunization In Vitro Infection Inflammation Infusion procedures Lead Learning Length Lipids Liposomes Mammals Measures Mediating Membrane Modeling Modification Monoclonal Antibodies Mus Nitrates Oryctolagus cuniculus Partition Coefficient Patients Peptide antibodies Peptides Peripheral Blood Mononuclear Cell Phospholipids Polyethylene Glycols Positioning Attribute Post-Translational Protein Processing Primates Property Protein Region Research Personnel Serine Serine/Threonine Phosphorylation Serum Side Site Source Structure Testing Threonine Tyrosine Vaccinated Vaccination Vaccines Vesicle Viral Viral Load result Virus Diseases Walkers base carboxylate cholesteryl hemisuccinate clinically relevant cross reactivity design high risk immunogenic immunogenicity immunoreactivity in vitro Assay in vivo neutralizing antibody neutralizing monoclonal antibodies nitration novel physical property prevent prophylactic protein aminoacid sequence research study scaffold success triple helix vaccine development

项目摘要

DESCRIPTION (provided by applicant): In this R21 application we postulate that the broadly HIV neutralizing monoclonal antibodies 2F5 and 4E10, against the membrane proximal external region (MPR) of GP41 were induced in the donors by MPR epitopes that had been post-translational modified. Our high-risk idea is to generate MPR immunogens incorporating chemically-modified amino acid residues that reflect the post-translational modifications that can occur during viral infections. These modifications are immediately adjacent to the MPR epitopes recognized by 2F5 and 4E10. We believe these novel immunogens will enhance the potency of existing epitopes and/or 'unmask' epitopes previously hidden from the immune system that induce neutralizing antisera. The idea to use such peptides containing the modified residues is unique. MPR immunogen design has focused on eliciting antibody responses that cross-react with phospholipids because the broadly neutralizing mAbs 2F5 and 4E10 exhibit unusual cross-reactivity with lipids. This led many investigators to attribute their dual affinity for both gp41 and phospholipids in the viral membrane as the source of their potency. However, the "lipid reactivity" of these antibodies is primarily directed at negatively charged lipid head groups, which are chemically similar to phosphorylated or nitrated amino acids. Thus, amino acid modifications, not lipids, may be the true cause of the unusual dual recognition properties of 2F5 and 4E10. We will synthesize MPR peptide sequences containing nitrated, phosphorylated, or non-hydrolyzable charge equivalent, phosphonated or sulfonated residues, attach them to lipid anchors, characterize their secondary structure and measure their partition coefficient in lipid vesicles. The lipopeptides will be formulated in an immunogenic liposome and used to immunize mice and rabbits. Antisera will be tested for antibody titers using a validated ELISA and neutralization will be measured in a clinically relevant PBMC HIV infection assay. To learn if presentation of the modified epitopes in a viral-mimic triple helix bundle increases immunogenicity and/or results in the generation of neutralizing antibodies, the modified peptides will be incorporated into a scaffold that presents three epitopes on the same lipid anchor and evaluated as described for the lipopeptides. To determine if the immunogenicity of the modified epitopes requires a lipid anchor, we will replace the lipid on the modified epitopes with polyethylene glycol and repeat the in vitro characterization and in vivo immunization studies. Results from these experiments will guide subsequent rounds of epitope design to optimize antisera titers and HIV neutralization activity. Success in generating neutralizing antibodies from chemically-modified peptide epitopes would enable structural studies on the epitope-antibody complex and lead to protection studies in primate HIV challenge models. This unusual approach could provide a widely applicable epitope design strategy for development of vaccines to prevent infectious disease such as HIV. PUBLIC HEALTH RELEVANCE: We will synthesize MPR immunogens that incorporate chemically-modified amino acids which reflect the post- translational modifications that can occur during HIV infections. We will use the immunogens in a prophylactic HIV vaccine to induce broadly HIV neutralizing antibodies in mammals which could be a precursor to a vaccine to prevent AIDS.

描述（由申请人提供）：在此 R21 申请中，我们假设针对 GP41 近膜外部区域 (MPR) 的广泛 HIV 中和单克隆抗体 2F5 和 4E10 是由经过翻译后修饰的 MPR 表位在供体中诱导的。我们的高风险想法是产生包含化学修饰氨基酸残基的 MPR 免疫原，这些氨基酸残基反映了病毒感染期间可能发生的翻译后修饰。这些修饰紧邻 2F5 和 4E10 识别的 MPR 表位。我们相信这些新型免疫原将增强现有表位的效力和/或“揭露”先前隐藏在免疫系统中的表位，从而诱导中和抗血清。使用此类含有修饰残基的肽的想法是独特的。 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，因为广泛中和的 mAb 2F5 和 4E10 表现出与脂质不寻常的交叉反应性。这使得许多研究人员将其对病毒膜中 gp41 和磷脂的双重亲和力归因于其效力的来源。然而，这些抗体的“脂质反应性”主要针对带负电荷的脂质头部基团，其化学性质类似于磷酸化或硝化氨基酸。因此，氨基酸修饰，而不是脂质，可能是 2F5 和 4E10 不寻常的双重识别特性的真正原因。我们将合成含有硝化、磷酸化或不可水解的电荷当量、膦化或磺化残基的 MPR 肽序列，将它们附着到脂质锚定上，表征它们的二级结构并测量它们在脂质囊泡中的分配系数。脂肽将被配制在免疫原性脂质体中并用于免疫小鼠和兔子。将使用经过验证的 ELISA 测试抗血清的抗体滴度，并在临床相关的 PBMC HIV 感染测定中测量中和作用。为了了解模拟病毒三螺旋束中修饰的表位的呈现是否会增加免疫原性和/或导致中和抗体的产生，修饰的肽将被纳入在同一脂质锚上呈现三个表位的支架中，并进行评估：描述了脂肽。为了确定修饰表位的免疫原性是否需要脂质锚，我们将用聚乙二醇替换修饰表位上的脂质，并重复体外表征和体内免疫研究。这些实验的结果将指导后续几轮表位设计，以优化抗血清滴度和 HIV 中和活性。成功地从化学修饰的肽表位产生中和抗体将使表位-抗体复合物的结构研究成为可能，并导致灵长类HIV攻击模型的保护研究。这种不寻常的方法可以为开发预防艾滋病毒等传染病的疫苗提供广泛适用的表位设计策略。公共健康相关性：我们将合成含有化学修饰氨基酸的 MPR 免疫原，这些氨基酸反映了 HIV 感染期间可能发生的翻译后修饰。我们将使用预防性艾滋病毒疫苗中的免疫原在哺乳动物中诱导广泛的艾滋病毒中和抗体，这可能是预防艾滋病疫苗的前身。