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: 10666579
• 负责人: Thomas Hope
• 金额: $ 23.77万
• 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/10666579
• 关键词: Ablation; Anatomy; Animals; Antibodies; Aquila; Area; Autopsy; Binding; Biological Models; CD4 Positive T Lymphocytes; Cells; Cellular Morphology; Collaborations; DNA; Dendritic Cells; Detection; Down-Regulation; Environment; Evaluation; Grant; HIV; Image; Immobilization; Infection; Inflammation; Interruption; Intervention; Investigation; Kinetics; Knowledge; Label; Location; Lymphoid Tissue; Macaca mulatta; Macrophage; 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; Reproducibility; Resolution; Role; SIV; Sampling; Scanning; 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; Visualization; acute infection; antiretroviral therapy; innovation; insight; 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.

项目摘要/摘要 如果没有完全了解艾滋病毒水库的形成，其持久性和潜在反弹，它将 很难开发疗法以获得艾滋病毒的治疗方法。艾滋病毒领域需要克服的主要局限性 为了促进我们对病毒反弹的理解，无法系统地研究储层 组织。在P01的支持下，以早期储层的病毒反弹为重点，我们最近优化了 我们命名为PET-CT指导尸检的新工作流程。此工作流程基于使用64CU标记的使用 抗Siv Env Fab2（64CU-7D3FAB2）指导采样和/或尸检过程以识别和收集碎片 含有64CU信号的焦点的组织揭示了SIV复制位点。病毒感染细胞的存在 这些“热”组织现在通过4个标记（GAG和ENV表达以及CD4和MHC-I下调）来验证。 出乎意料的是，我们仅在ATI后的头7-10天内检测到SIVMAC239髓样细胞的感染。没有SIV 尽管CD4 T细胞在这些焦点中很丰富，但观察到感染的T细胞。最后，我们的64CU-7D3FAB2问题 引导我们到达组织，我们可以通过髓样细胞形态可重复地可视化生成病毒粒子的细胞 透射电子显微镜。这种多刻度成像，利用PET/CT引导的尸检，使我们能够 达到难以捉摸的组织晶状体。在该计划1的该项目1中，我们建议利用我们的进步 与组织的PET-CT指导采样以及非人类的其他创新抽样技术 灵长类动物（NHP）模型，以剖析持续的SIV储层的动力学。具体来说，我们计划了3 不同类型的NHP研究，借助临时干预，将为我们提供前所未有的分辨率 进入水库的动力及其反弹。我们计划采取旨在“挠痒”和的干预措施 在购物车期间干扰储层，这将允许PET-CT信号增加并收集组织样品 这将通过包括多尺度成像在内的不同技术进行分析，并分发给其他两个项目。 我们将通过感染前的DSS治疗中的DSS治疗来“覆盖”储层，以覆盖储层。 然后在购物车计划时再次对组织的研究器进行深入研究，并在几次之后 几个月的抑制车。最后，我们将破坏或刺激髓样细胞，我们的研究揭示了这些细胞 作为ATI后病毒反弹的主要贡献者，并研究了这些干预措施对储层的影响 反弹病毒血症的动力和时机。总而言之，我们的项目将在NHP研究中使用不同的策略 阐明SIV储层的性质和来源，并特别关注粘膜组织和髓样细胞。