Project 1: Dissecting Persistent Virus Reservoirs in Tissues

项目 1：剖析组织中的持久病毒库

• 批准号: 10460076
• 负责人: Thomas Hope
• 金额: $ 23.02万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-15 至 2027-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10460076
• 关键词: Anatomy; Animals; Antibodies; Aquila; Area; Autopsy; Biological Models; CD4 Positive T Lymphocytes; Cells; Cellular Morphology; Collaborations; DNA; Dendritic Cells; Detection; Down-Regulation; Environment; Evaluation; Grant; HIV; Image; Infection; Inflammation; Interruption; Intervention; Investigation; Kinetics; Knowledge; Label; Location; Lymphoid Tissue; Macaca mulatta; Methods; Modeling; Monitor; Mucous Membrane; Myeloid Cells; Names; Nature; Nucleic Acids; PET/CT scan; Phase; Physiology; Play; Population; Population Dynamics; Positron-Emission Tomography; Process; Production; Proteins; RNA; Radioactive; Resolution; Role; SIV; Sampling; Scanning; Seeds; Signal Transduction; Site; Source; T-Lymphocyte; Techniques; Technology; Time; Tissue Banks; Tissue Sample; Tissues; Transmission Electron Microscopy; Trauma; Viral Antigens; Viral Proteins; Viral reservoir; Viremia; Virion; Virus; acute infection; antiretroviral therapy; base; innovation; insight; macrophage; mast cell; nonhuman primate; pathogen; prevent; programs; recruit; simian immunodeficiency virus gp120; therapy development; tissue repair; viral rebound

PROJECT SUMMARY/ABSTRACT Without a complete knowledge of the formation of the HIV reservoir, its persistence, and potential rebound, it will be difficult to develop therapies to achieve a cure for HIV. A major limitation that the HIV field needs to overcome in order to advance our understanding of viral rebound is the inability to systemically study the reservoir in tissues. With the support of a P01 focused on viral rebound of the early reservoir, we have recently optimized a new workflow that we named PET-CT guided necropsy. This workflow is based on the use of a 64Cu-labeled anti-SIV env FAB2 (64Cu-7D3FAB2) to guide the sampling and/or necropsy process to identify and collect pieces of tissues containing foci of 64Cu signal revealing sites of SIV replication. The presence of virus infected cells in these “hot” tissues is now validated by 4 markers (Gag and Env expression and CD4 and MHC-I downregulation). Unexpectedly, we only detect SIVmac239 infection of myeloid cells during the first 7-10 days after ATI. No SIV infected T cells are observed although CD4 T cells are abundant in these foci. Finally, our 64Cu-7D3FAB2 probe led us to tissues where we can reproducibly visualize virion producing cells, with a myeloid cell morphology, by transmission electron microscopy. This multiscaling imaging, leveraging PET/CT guided necropsy, allows us to peak into the elusive tissue reservoir. In this Project 1 of the Program, we propose to leverage our advancements with PET-CT-guided sampling of tissues as well as additional innovative sampling techniques in non-human primate (NHP) models to dissect the dynamics of the persistent SIV reservoir. Specifically, we have planned 3 different type of NHP studies that, with the help of ad-hoc interventions, will give us an unprecedented resolution into the dynamics of the reservoir and its rebound. We have planned interventions that aim at “tickling” and disturbing the reservoir during cART, which will allow PET-CT-signal increase and collection of tissue samples that will be analyzed with different techniques including multiscale imaging and distributed to the other 2 projects. We will “mulch” the reservoir by favoring its seeding into mucosal tissues with DSS treatment before infection and then again an in-depth investigation of the tissue reservoir at the time of cART initiation and after several months of suppressive cART. Finally, we will deplete or stimulate myeloid cells, which our studies have revealed as main contributor to viral rebound after ATI and study the impact of these interventions on the reservoir dynamics and timing of rebound viremia. In summary, our project will use different strategies in NHP studies to clarify the nature and source of the SIV reservoir with particular focus at the mucosal tissue and myeloid cells.

项目总结/摘要 如果不完全了解艾滋病病毒库的形成、持久性和潜在的反弹， 很难开发出治愈艾滋病毒的疗法。艾滋病毒领域需要克服的一个主要限制 为了提高我们对病毒反弹的理解，我们无法系统地研究病毒反弹的宿主， 组织中在专注于早期水库病毒反弹的P01的支持下，我们最近优化了 我们命名为PET-CT引导尸检的新工作流程。该工作流程基于使用64 Cu标记的 抗SIV env FAB 2（64 Cu-7 D3 FAB 2），用于指导采样和/或尸检过程，以识别和收集碎片 的组织含有病灶的64铜信号揭示网站的SIV复制。病毒感染细胞的存在， 这些“热”组织现在通过4种标记物（Gag和Env表达以及CD 4和MHC-I下调）验证。 出乎意料的是，我们仅在ATI后的前7-10天期间检测到骨髓细胞的SIVmac 239感染。无SIV 尽管CD 4 T细胞在这些病灶中是丰富的，但是观察到感染的T细胞。最后，我们的64 Cu-7 D3 FAB 2探针 引导我们找到了组织，在那里我们可以再现地看到产生病毒体的细胞，具有髓样细胞形态， 透射电子显微镜这种多尺度成像利用PET/CT引导尸检，使我们能够 进入难以捉摸的组织库在该计划的项目1中，我们建议利用我们的进步 通过PET-CT引导的组织采样以及其他创新的非人类采样技术， 灵长类动物（NHP）模型来剖析持续性SIV库的动态。具体来说，我们计划3 不同类型的NHP研究，在特别干预的帮助下，将为我们提供前所未有的解决方案 水库的动态及其反弹。我们已经计划了旨在“挠痒痒”的干预措施， 在cART期间干扰储液器，这将允许PET-CT信号增加和组织样本收集 将使用不同的技术进行分析，包括多尺度成像，并分发给其他2个项目。 我们将通过在感染前用DSS治疗促进其接种到粘膜组织中来“覆盖”水库 然后再次在cART开始时和几次治疗后对组织储库进行深入研究， 几个月的抑制性CART最后，我们将耗尽或刺激骨髓细胞，我们的研究已经揭示了这一点 作为ATI后病毒反弹的主要贡献者，并研究这些干预措施对储库的影响 病毒血症反弹的动力学和时间。总之，我们的项目将在NHP研究中使用不同的策略， 阐明SIV储库的性质和来源，特别关注粘膜组织和骨髓细胞。