Epitope-Focused HIV-1 Envelope Vaccine for HIV-1/AIDS

描述（由申请人提供）：本项目的总体目标是探索和推进HIV-1包膜（Env）蛋白的结构限制形式，用于开发HIV-1疫苗。病毒包膜蛋白gp 120的配方，可以编程人类免疫系统识别和阻断HIV-1包膜蛋白一直是艾滋病疫苗开发的主要目标。然而，已经发现gp 120能够逃避免疫监视，这主要是由于其构象的灵活性。鉴定诱导增加的结构稳定性的方法，特别是通过保守表位聚焦，被视为提高包膜疫苗引发中和免疫应答的能力的重要目标。最近，我们开发了一组称为“变构双位点拮抗剂”的分子，其可以以这样的方式与病毒gp 120强烈结合，从而将gp 120捕获在三维结构中，该三维结构在功能上受到抑制，但具有可接近的CD 4 Phe 43结合口袋。我们假设变构双位点拮抗剂可以诱导HIV-1包膜蛋白中免疫原性景观的改变，并且这种改变的景观可能会导致针对保守受体结合位点（包括CD 4）的中和免疫应答增强。我们将在本PIA项目的R21阶段测试这一假设，主要目标有两个。（R21-Aim 1）使用重组蛋白工程和肽合成，我们的目标是产生HIV-1 Env免疫原，其中HNG-156类的变构双位点拮抗剂与HIV-1 gp 120的重组三聚体和单体形式非共价或共价结合。（R21-Aim 2）使用小动物模型，我们的目的是检验这样的假设，即变构gp 120抑制剂与重组三聚体和单体形式的HIV-1 gp 120的复合物和共价融合物的疫苗接种将引起增强的抗HIV gp 120特异性抗体和T细胞表位宽度、幅度和B细胞的中和能力。R21项目的关键里程碑将是证明HIV-1 Env gp 120的变构双位点拮抗剂促进的增强的血清中和活性。 这一里程碑的实现将导致R33的开发阶段，包括鉴定具有优化的HIV-1 Env与变构抑制剂非共价和共价结合的组合物的高级免疫原（R33-Aim 1）;使用非人灵长类动物模型进行粘膜激发的免疫原检测（R33- Aim2）;和产生由Env变构抑制剂免疫原引发的潜在中和单克隆抗体，其将表征Env结合表位和中和宽度（R33-Aim 3）。 总的来说，该项目将通过构象约束使用表位聚焦，以了解变构配体如何以免疫原性生产方式控制HIV-1 Env蛋白的构象，确定HIV-1 Env中的中和位点以获得保护性免疫，开发用于非人灵长类动物疗效/毒性评估的Env构建体，并将免疫原制剂推向灵长类动物，并最终进行人体试验。 公共卫生相关性：该项目旨在测试新发现的HIV-1病毒外壳蛋白形式，作为设计艾滋病疫苗的工具，可用于为人类提供免疫保护。

DESCRIPTION (provided by applicant): The overall goal of this project is to explore and advance structurally constrained forms of HIV-1 envelope (Env) protein for development of an HIV-1 vaccine. Formulations of viral envelope protein gp120 that could program the human immune system to recognize and block the HIV-1 envelope protein have been a major goal of AIDS vaccine development. However, gp120 has been found to be able to evade immune surveillance, due significantly to its conformational flexibility. Identifying approaches to induce increased structural stability, in particular by conserved epitope focusing, is seen as an important goal to improve the ability of envelope vaccines to elicit neutralizing immune responses. Recently, we have developed a set of molecules, called "allosteric dual site antagonists", that can bind strongly to the viral gp120 in such a way as to entrap gp120 in a three dimensional structure that is functionally suppressed but with an accessible CD4 Phe 43 binding pocket. We hypothesize that allosteric dual site antagonists can induce an altered immunogenic landscape in HIV-1 envelope protein, and that such an altered landscape can potentially lead to an enhanced neutralizing immune response against conserved receptor binding sites, including that for CD4. We will test this hypothesis in the R21 phase of this PIA project with 2 major aims. (R21-Aim1) Using recombinant protein engineering and peptide synthesis, we aim to produce HIV-1 Env immunogens in which allosteric dual site antagonists of the HNG-156 class are bound either noncovalently or covalently with recombinant trimeric and monomeric forms of HIV-1 gp120. (R21-Aim2) Using small animal models, we aim to test the hypothesis that vaccination of complexes and covalent fusions of allosteric gp120 inhibitors with recombinant trimeric and monomeric forms of HIV-1 gp120 will elicit enhanced anti-HIVgp120 specific antibody and T cell epitope breadth, magnitude and the neutralization capacity of B cells. The key milestone of the R21 project will be to demonstrate enhanced serum neutralization activities promoted by allosteric dual site antagonists of HIV-1 Env gp120. Achievement of this milestone will lead to an R33 development phase that includes identification of advanced immunogens with optimized compositions of HIV-1 Env bound noncovalently and covalently to allosteric inhibitors (R33-Aim1); testing of immunogens using a non-human primate model with mucosal challenge (R33- Aim2); and production of potential neutralizing monoclonal antibodies, elicited by Env -allosteric inhibitor immunogens, that will be characterized for Env binding epitopes and breadth of neutralization (R33-Aim3). Overall, this project will use epitope focusing through conformational constraint to learn how allosteric ligands can exert control over conformations of HIV-1 Env protein in an immunogenically productive way, identify neutralization sites in HIV-1 Env for protective immunity, develop Env constructs for non-human primate efficacy/toxicity evaluation, and advance immunogen formulations to primate and, ultimately, human trials. PUBLIC HEALTH RELEVANCE: This project seeks to test newly identified forms of the HIV-1 virus coat protein as tools for designing AIDS vaccines that could be used to provide immunoprotection in humans.