Learning from attenuated CMV how to broaden HIV-specific T cell responses

从减毒 CMV 中学习如何扩大 HIV 特异性 T 细胞反应

• 批准号: 8732086
• 负责人: Sylvie Le Gall
• 金额: $ 53.14万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2018-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8732086
• 关键词: Affect; Antigen-Presenting Cells; Antigens; Area; Attenuated; Binding; Biochemical; Biological Assay; Bypass; CD4 Positive T Lymphocytes; CD8B1 gene; Cell membrane; Cells; Cellular Tropism; Computer Analysis; Cross Presentation; Cytomegalovirus; Cytomegalovirus Infections; Cytosol; Dendritic Cells; Development; Disease; Endoplasmic Reticulum; Epitopes; Event; Fibroblasts; Frequencies; Gagging; HIV; HIV Antigens; HIV Infections; HIV vaccine; Health; Human; Immune response; Immunologic Deficiency Syndromes; In Vitro; Infection; Kinetics; Knowledge; Lead; Learning; Length; Life; Lysosomes; MHC binding peptide; Macaca; Macaca mulatta; Mass Spectrum Analysis; Measures; Memory; Methodology; Mus; Outcome; Pattern; Peptide Hydrolases; Peptide/MHC Complex; Peptides; Play; Process; Production; Proteins; Role; Route; SIV; Shapes; Source; System; T cell response; T-Lymphocyte; Testing; Time; Vaccinated; Vaccination; Vaccine Design; Vaccines; Variant; Viral Proteins; Virion; Virus; Walkers; antigen processing; base; cell type; design; in vivo; lymph nodes; multidisciplinary; nanoparticle; pathogen; protein degradation; public health relevance; response; trafficking; vector

DESCRIPTION (provided by applicant): Vaccination of Rhesus Macaques with an attenuated CMV virus carrying SIV proteins (RhCMV-SIV) led to an unprecedented durable control and clearance of SIV in 50% of vaccinated Macaques, a positive outcome predicted by the magnitude of CD8 T cell responses during vaccination and effector T cell responses in lymph nodes after infection. RhCMV-SIV vaccination broke natural immunodominance and led to broad CD4 T cells and CD8 responses covering many areas of SIV antigens. Surprisingly most SIV-specific CD8 T cells responses elicited by this vaccine were restricted by MHC-II. Whereas the results are among the most promising to date in the HIV vaccine field the use of CMV vectors for HIV vaccination raise health concerns. The first event required to prime T cell responses is the presentation of MHC-I- and MHC-II-bound peptides by antigen presenting cells (APC) to T cells. These peptides come from the multistep intracellular degradation of proteins by the antigen processing machinery. Considering the capacity of WT CMV to alter MHC-peptide presentation we propose that attenuated CMV expressing HIV proteins (HCMV-HIV) creates unique conditions for processing and presentation of HIV epitopes leading to broad unconventional T cell responses, and that these conditions can be replicated without the use of CMV by transiently manipulating the antigen processing machinery during the delivery of the immunogen in APC. To unveil these mechanisms of protein degradation and epitope presentation in the presence of attenuated HCMV-HIV we have developed throughput assays to measure antigen processing activities in primary cells, mass spectrometry-based assays to follow the degradation of proteins or virions in the two cellular compartments where the virus may enter: cytosol and endo-lysosomes, and methodologies to identify intracellular epitope precursors and MHC-bound peptides in HIV-infected cells. We showed that variations in the levels of peptidase activities among cell subsets play a critical role in shaping the lengths and kinetics of peptides produced for epitope presentation. Using a combination of biochemical, computational, in vitro and in vivo immunological approaches we propose to 1) Determine the effect of attenuated HCMV-HIV on HIV protein degradation in target cell subsets, 2) Assess epitope presentation and priming of T cell responses in the context of attenuated HCMV-HIV infection, and 3) Design and test a CMV-free antigen delivery system leading to broad HIV-specific T cell responses. This project builds on a multidisciplinary collaborative approach between the PI, Dr Heckerman for computational analysis of HIV degradation products, Dr Picker for HCMV expertise in CMV, Dr Moris for in vitro priming assays of HIV-specific T cell responses, Dr Walker for HIV-specific T cells, Dr Tager for humanized mice and Dr Irvine for the development of nanoparticles for antigen delivery.

描述（由申请人提供）：用携带 SIV 蛋白的减毒 CMV 病毒（RhCMV-SIV）对恒河猴进行疫苗接种，在 50% 的接种猕猴中实现了史无前例的持久控制和清除 SIV，根据疫苗接种期间 CD8 T 细胞反应的强度和感染后淋巴结中效应 T 细胞反应的程度预测，这是一个积极的结果。 RhCMV-SIV 疫苗接种打破了自然免疫优势，导致广泛的 CD4 T 细胞和 CD8 反应，覆盖了 SIV 抗原的许多区域。令人惊讶的是，该疫苗引发的大多数 SIV 特异性 CD8 T 细胞反应均受到 MHC-II 的限制。尽管这些结果是迄今为止 HIV 疫苗领域最有希望的结果之一，但使用 CMV 载体进行 HIV 疫苗接种引起了健康问题。启动 T 细胞反应所需的第一个事件是抗原呈递细胞 (APC) 将 MHC-I 和 MHC-II 结合肽呈递给 T 细胞。这些肽来自抗原加工机器对蛋白质的多步细胞内降解。考虑到 WT CMV 改变 MHC 肽呈递的能力，我们建议表达 HIV 蛋白的减毒 CMV（HCMV-HIV）为 HIV 表位的加工和呈递创造了独特的条件，从而导致广泛的非常规 T 细胞反应，并且这些条件可以在不使用 CMV 的情况下通过在免疫原递送过程中瞬时操纵抗原加工机制来复制。 装甲运兵车。为了揭示减毒 HCMV-HIV 存在下蛋白质降解和表位呈递的这些机制，我们开发了通量测定法来测量原代细胞中的抗原加工活性，基于质谱的测定法来跟踪病毒可能进入的两个细胞区室中蛋白质或病毒体的降解：胞质溶胶和内溶酶体，以及识别细胞内表位的方法 HIV 感染细胞中的前体和 MHC 结合肽。我们表明，细胞亚群之间肽酶活性水平的变化在塑造用于表位呈递的肽的长度和动力学方面发挥着关键作用。通过结合生化、计算、体外和体内免疫学方法，我们建议 1) 确定减毒 HCMV-HIV 对靶细胞亚群中 HIV 蛋白降解的影响，2) 评估减毒 HCMV-HIV 感染背景下的表位呈递和 T 细胞反应的启动，以及 3) 设计和测试无 CMV 的抗原递送系统，从而产生广泛的 HIV 特异性 T 细胞反应。该项目建立在 PI 之间的多学科合作方法之上，Heckerman 博士负责 HIV 降解产物的计算分析，Picker 博士负责 CMV 方面的 HCMV 专业知识，Moris 博士负责 HIV 特异性 T 细胞反应的体外启动测定，Walker 博士负责 HIV 特异性 T 细胞，Tager 博士负责人源化小鼠，Irvine 博士负责开发用于抗原递送的纳米颗粒。