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

• 批准号: 8320078
• 负责人: FRANCIS C. SZOKA
• 金额: $ 23.18万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 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/8320078
• 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

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.

描述(申请人提供)：在这份R21申请中，我们假设针对GP41膜近端外区(MPR)的广泛的HIV中和单抗2F5和4E10是由翻译后修饰的MPR表位在供者体内诱导的。我们的高风险想法是产生MPR免疫原，其中包含化学修饰的氨基酸残基，反映病毒感染期间可能发生的翻译后修饰。这些修饰直接与2F5和4E10识别的MPR表位相邻。我们相信，这些新的免疫原将增强现有表位和/或先前对免疫系统隐藏的“揭开”表位的效力，这些表位诱导中和抗血清。 使用这种含有修饰残基的肽的想法是独一无二的。MPR免疫原设计的重点是激发与磷脂交叉反应的抗体反应，因为广泛中和的mAb2F5和4E10与脂类表现出不同寻常的交叉反应。这导致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它们的效力来源。然而，这些抗体的“脂质反应性”主要针对带负电荷的脂头基团，这些基团在化学上类似于磷酸化或硝化氨基酸。因此，氨基酸修饰，而不是脂类，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、磷酸化或非水解性电荷当量、磷酸化或磺化残基的MPR多肽序列，将它们连接到脂质锚上，表征它们的二级结构，并测量它们在脂泡中的分配系数。这些脂肽将被制成免疫原脂质体，并用于免疫小鼠和兔子。抗血清将使用有效的ELISA法进行抗体滴度测试，中和将在临床相关的PBMC HIV感染测试中进行测量。为了了解以病毒模拟的三螺旋束呈现修饰表位是否提高免疫原性和/或导致中和抗体的产生，将修饰多肽结合到一个支架中，该支架在同一脂锚上呈现三个表位，并按照脂肽的描述进行评估。为了确定修饰表位的免疫原性是否需要脂质锚，我们将用聚乙二醇取代修饰表位上的脂类，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，以优化抗血清效价和HIV中和活性。成功地从化学修饰的多肽表位产生中和抗体将使对表位-抗体复合体的结构研究成为可能，并导致在灵长类艾滋病毒挑战模型中进行保护研究。这种不同寻常的方法可以为疫苗的开发提供一种广泛适用的表位设计策略，以预防艾滋病毒等传染病。