Project 1. Optimization and in vivo evaluation of HIV-1 Env trimer sortase A-conjugated nanoparticles

项目1. HIV-1 Env三聚体分选酶A结合纳米粒子的优化及体内评价

• 批准号: 10541863
• 负责人: KEVIN O SAUNDERS
• 金额: $ 83.42万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-12-17 至 2026-11-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10541863
• 关键词: Adopted; Affinity; Antibodies; Antibody Affinity; Antibody Binding Sites; Antibody Formation; Antibody Response; Antigens; Autologous; B-Cell Antigen Receptor; B-Lymphocytes; Binding Sites; Cell Lineage; Cloning; Codon Nucleotides; Cyclic GMP; Development; Effectiveness; Engineering; Enzymes; Failure; Ferritin; Genes; HIV Antigens; HIV-1; Human; Immunization; Immunize; Knock-in Mouse; Link; Macaca; Molecular Conformation; Monoclonal Antibodies; Mutation; Phenotype; Positioning Attribute; Probability; Process; Production; Protein Subunits; Proteins; Reaction; Recombinants; Regimen; Research; Running; Serum; Somatic Mutation; Surface; Technology; Vaccine Design; Vaccines; Virus; cross reactivity; design; expectation; humanized mouse; immunogenicity; in vivo evaluation; innovation; manufacture; member; mouse model; nanoparticle; neutralizing antibody; next generation sequencing; non-Native; nonhuman primate; novel; phase I trial; research clinical testing; self assembly; sortase; transpeptidation; vaccination strategy

ABSTRACT-PROJECT 1 Previous vaccines have been unable to elicit HIV-1 broadly neutralizing antibodies (bnAbs) in humans. Two of the reasons for this failure are that bnAb precursor B cells are often rare and that the B cell receptors on those rare B cells require extensive somatic mutation to evolve into high-affinity bnAbs. A subset of these somatic mutations—termed improbable mutations—have a low probability of being made by the somatic mutation machinery due to codon bias and positioning of somatic mutation recognition motifs. Our central hypothesis for eliciting bnAbs is that immunogens must have a higher affinity for Abs with the required improbable mutations than for Abs lacking these key mutations. Moreover, we hypothesize that mutation-guided vaccine design will need to engage low-affinity rare bnAb precursors and, then, through immunization with sequential immunogens, select for Abs acquiring the somatic changes requisite for a broad neutralization phenotype. We and others have observed that multimerizing HIV-1 envelope (Env) immunogens on nanoparticles (NPs) can increase Ab titers and affinity maturation of HIV-1 bnAb lineages. However, Env is unstable and can adopt non-native conformations on the surface of NPs, leading to undesired Ab responses. Additionally, NPs bearing HIV-1 envelope can be of low protein yield. To overcome these pitfalls, we have developed an innovative, rapid platform that uses the enzyme sortase A to covalently link well-folded, cleaved Env trimers to self- assembling ferritin protein NPs. These sortase A-conjugated NPs (scNPs) maintain near-native Env trimer conformation, allowing avid selection of improbable mutations in evolving bnAb lineages. In humanized mice, we used a scNP displaying an engineered Env trimer called CH505.M5 G458Y to select for a key improbable mutation required for maturation of the 8ANC131/CH235 class of CD4 binding site (CD4bs) bnAbs. The same scNP elicited CD4bs monoclonal neutralizing Abs and serum neutralizing Abs in nonhuman primates (NHPs). To further guide these Abs to develop neutralization breadth, we have generated a boosting Env trimer scNP immunogen called CH505.TF scNP and have a third boosting Env under development. In CH505.M5 G458Y scNP-primed NHPs, CH505 TF scNP immunization boosts neutralizing Ab titers against multiple CH505 viruses. This IPCAVD will generate a CH505 TF scNP boosting immunogen and a second sequential Env trimer scNP boosting immunogen for a Phase I trial. In Specific Aim 1, Project 1 will determine an optimal NP boosting regimen to broaden the neutralization capacity of current CD4bs B cell lineages in humanized mice. In Specific Aim 2, we will compare conjugation efficiency and immunogenicity of scNPs assembled from research-grade or development-run ferritin and sortase A components from Project 2. Lastly, Specific Aim 3 will demonstrate the induction of CD4bs lineages in NHPs using the prime-boost strategy and cGMP scNPs proposed for the Phase I trial. The impact of this Project is that it will define and demonstrate the effectiveness of a novel, optimal NP vaccine regimen to expand and affinity mature 8ANC131/CH235 class CD4bs bnAbs.

抽象项目1 以前的疫苗无法引起人类中和抗体的广泛中和抗体（BNAB）。两个 发生这种失败的原因是BNAB前体B细胞通常很少 罕见的B细胞需要广泛的体细胞突变才能演变为高亲和力的BNAB。这些躯体的子集 突变 - 不可能的突变 - 通过体细胞突变产生的可能性很低 由于密码子偏置和体细胞突变识别基序的定位引起的机械。我们的中心假设 为了引起bnabs，免疫原子必须具有较高的ABS的亲和力，而所需的不可能 突变比缺乏这些关键突变的ABS。此外，我们假设突变引导的疫苗 设计将需要参与低亲和力的稀有BNAB前体，然后通过顺序进行免疫 免疫原子，选择ABS获得广泛神经化表型所需的躯体变化。我们 其他人也观察到纳米颗粒（NP）上的多颗HIV-1包膜（ENV）免疫原子可以 增加AB滴度和HIV-1 BNAB谱系的亲和力成熟。但是，Env是不稳定的，可以采用 NP表面上的非本性构象，导致不希望的AB反应。另外，NP轴承 HIV-1包膜可以低蛋白质产量。为了克服这些陷阱，我们开发了一种创新的， 快速平台使用酶排序酶A将折叠，切割的Env Trimers的共价连接到自我 组装铁蛋白蛋白NP。这些排序酶A结合的NP（SCNP）保持近乎邻国的ENV触发 构象，可以在不断发展的BNAB谱系中狂热地选择不可能的突变。在人源化的小鼠中， 我们使用的SCNP显示了一个名为CH505.M5 G458Y的工程env trimer来选择一个不可能的钥匙 8anc131/CH235 CD4结合位点（CD4BS）BNABS成熟所需的突变。相同 SCNP在非人类素数（NHP）中引起了CD4BS单克隆中和ABS的中和ABS。 为了进一步指导这些腹肌以发展神经化广度，我们产生了一个增强的env触发scnp 免疫原称为CH505.TF SCNP，并在开发中具有第三个增强ENV。在CH505.M5 G458Y中 SCNP培养的NHP，CH505 TF SCNP免疫增强了AB滴度的中和滴度 病毒。该IPCAVD将产生CH505 TF SCNP，增强免疫原和第二顺序ENV Trimer SCNP在I期试验中增强免疫原。在特定目标1中，项目1将确定最佳NP 增强方案以扩大人性化小鼠中当前CD4BS B细胞谱系的神经元化能力。 在特定的目标2中，我们将比较从 研究级或开发运行的铁蛋白和分类项目2中的成分。最后，特定目标3 将使用Prime-Boost策略和CGMP SCNP来证明NHP中CD4BS谱系的诱导 提出了第一阶段试验的建议。该项目的影响是它将定义和证明有效性 一种新型的最佳NP疫苗方案，以扩展和亲和力成熟8anc131/CH235 CD4BS BNABS。