Defining HIV Env protein expression in latently infected cells

定义潜伏感染细胞中的 HIV 包膜蛋白表达

• 批准号: 10762524
• 负责人: Alberto Bosque
• 金额: $ 24.23万
• 依托单位: GEORGE WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-04 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10762524
• 关键词: Address; Affect; Antibodies; B-Lymphocytes; Binding; Biological Assay; Cell Separation; Cell model; Cell surface; Cells; Cellular biology; Characteristics; Chronic; Clinical Trials; Data; Detection; Development; Effector Cell; Epitopes; Flow Cytometry; Future; Gender; Genetic; Genetic Variation; HIV; HIV Core Protein p24; HIV-1; Immune; Immunotherapy; In Vitro; Infection; Interruption; Investigation; Knowledge; Lead; Learning; Length; Measures; Mediating; Methodology; Methods; Modeling; Molecular Conformation; Molecular Virology; Outcome; Persons; Phagocytosis; Protein Conformation; Proteins; Publishing; RNA; RNA Splicing; Role; Surface; Testing; Therapeutic; Therapeutic Uses; Transcript; Translating; Translations; Viral; Viral Proteins; Virus; Virus Replication; antibody-dependent cell cytotoxicity; antiretroviral therapy; cohort; env Gene Products; flexibility; gag Gene Products; high reward; high risk; improved; lonely individuals; monomer; neutralizing antibody; novel; prevent; protein expression; reactivation from latency; treatment strategy; virus genetics

Project Summary/Abstract Advances in B cell biology and molecular virology have enabled the discovery, characterization, and commercial development of broadly neutralizing antibodies (bNAbs). They are a promising immunotherapy that can be incorporated in many strategies for the treatment and/or cure of HIV-1. Antibodies have many effector functions beyond neutralization and their most important mechanism of action for treatment/cure may be their ability to opsonize infected cells, tagging them for antibody-dependent cellular cytotoxicity or phagocytosis (ADCC/ADCP) by immune effector cells. It is not clear that bNAbs are mediating clearance of infected cells in clinical trials when they are passively infused into chronically infected people living with HIV (PLWH). An obvious reason for this lack of efficacy of bNAbs may be a lack of Env protein expression while on suppressive ART. Our lack of knowledge about the level of Env expression during HIV latency and latency reversal prevents effective use of bNAbs as therapeutics. To address this gap, the ability to detect low level Env protein expression in cells infected with different subtypes of HIV-1 is needed. The Bosque lab has published an ultrasensitive method to detect p24 Gag protein down to the fg/ml level and has preliminary data for a newly developed Env assay. Here we propose to use these assays to detect low levels of Env and Gag protein in a well-described model of latency and compare the data to Env surface expression as measured by flow cytometry (Aim 1). Within these assays, we have the flexibility to test Env detection using bNAbs targeting different epitopes to probe for Env conformation (i.e., trimer, monomer, etc) and extend these analyses to diverse HIV isolates from different subtypes to measure differences in Env expression and latency attributable to genetically diverse virus (Aim 2). Knowing these crucial factors about Env and Gag expression in a carefully controlled model of latency will allow us to further investigate HIV protein translation in future studies using cohorts of PLWH. These studies will improve the ability of therapeutics using Env-targeting strategies to target latently infected cells.

项目总结/摘要 B细胞生物学和分子病毒学的进展使得能够发现、表征和商业化。 广泛中和抗体（bNAb）的发展。它们是一种很有前途的免疫疗法， 在治疗和/或治愈HIV-1的许多策略中均采用了这种方法。抗体具有许多效应子功能 除了中和之外，它们最重要的治疗/治愈作用机制可能是它们的能力， 调理感染的细胞，标记它们的抗体依赖性细胞毒性或吞噬作用（ADCC/ADCP） 免疫效应细胞。目前尚不清楚bNAb是否在临床试验中介导感染细胞的清除， 它们被被动地输注到患有HIV的慢性感染者（PLWH）体内。一个明显的原因是 bNAb缺乏功效可能是在抑制性ART时缺乏Env蛋白表达。 关于HIV潜伏期和潜伏期逆转期间Env表达水平的知识妨碍了有效使用 bNAb作为治疗剂。为了解决这一差距，检测感染细胞中低水平Env蛋白表达的能力， 不同亚型的HIV-1感染。博斯克实验室发表了一种超灵敏的方法来检测p24 Gag蛋白降至fg/ml水平，并具有新开发的Env测定的初步数据。在这里我们建议 使用这些检测方法在一个描述良好的潜伏期模型中检测低水平的Env和Gag蛋白，并比较 通过流式细胞术测量Env表面表达的数据（Aim 1）。在这些测定中，我们有 使用靶向不同表位的bNAb检测Env构象的灵活性（即，三聚体， 单体等），并将这些分析扩展到来自不同亚型的不同HIV分离株，以测量差异 在Env中的表达和潜伏期归因于遗传多样性病毒（目的2）。了解这些关键因素 关于Env和Gag在一个精心控制的潜伏期模型中的表达将使我们能够进一步研究HIV 在未来的研究中使用PLWH的队列蛋白质翻译。这些研究将提高治疗的能力 使用Env靶向策略靶向潜伏感染的细胞。