Harnessing Stem-Like CD8 T Cells for Immunotherapies to Eradicate HIV Reservoirs

项目摘要 尽管联合抗逆转录病毒疗法(CART)有效地抑制了人类免疫缺陷病毒 (艾滋病毒)复制，它不能治愈艾滋病毒感染，需要昂贵的终身治疗。面临的一个重大挑战 治愈艾滋病毒感染是长期潜伏的艾滋病毒蓄水池，它逃避免疫识别，并对 在购物车中断后不久病毒反弹。大量的研究工作集中在消除 潜伏感染细胞通过所谓的“休克和杀死”策略，通过潜伏期重新激活潜伏的艾滋病毒- 反转剂(LRA)，允许通过细胞溶解或免疫介导的清除来“杀死”。特别是根除 抗HIV T细胞作为一种“活的”药物，维持患者的抗病毒免疫，是HIV储存库的一部分 要么完全解决感染，要么在不接受CART治疗的情况下保持长期控制，这是一个很有希望的策略。 然而，通过基于T细胞的免疫疗法清除HIV储存库仍然具有挑战性，因为1)T- 细胞耗尽，2)需要终生抗HIV免疫以取代CART，3)B细胞滤泡等避难所 这排除了大多数HIV特异的CD8 T细胞，以及4)LRA的影响不清楚，其中大多数针对表观遗传学 体内抗病毒CD8 T细胞的功能和分化途径。我们和其他人最近 猴慢性淋巴细胞性脉络膜脑膜炎病毒(LCMV)中干细胞样CD8 T细胞亚群的特征 免疫缺陷病毒(SIV)和艾滋病毒感染，以及在小鼠和人类肿瘤中。与.相比 终末耗尽的CD8T细胞，干细胞样的CD8T细胞耗尽较少，介导长期免疫，以及 在动物和人类的免疫疗法治疗后，反应更有力。慢性LCMV、SIV和 HIV感染时，这些细胞表达CXCR5，迁移到B细胞滤泡并杀死感染的T滤泡辅助细胞，a HIV和SIV的主要潜在宿主。此外，这些细胞的频率与病毒血症呈负相关。 SIV或HIV。最近，我们发现了干细胞样CD8的单细胞转录和表观遗传学特征 T细胞不同于急性或慢性LCMV感染后产生的其他CD8亚群。此外，我们 发现了一个涉及转录因子TOX的转录程序，该转录因子对干细胞样CD8T细胞是必不可少的 抗病毒CD8 T细胞在慢性病毒感染过程中的分化和长期存留在这里，我会 确定最佳T细胞所需的干细胞样CD8T细胞的转录和表观遗传程序 针对艾滋病毒的免疫疗法。我将使用慢性LCMV和SIV感染的动物模型以及样本 并使用尖端技术，包括T细胞转录本的单细胞图谱 和表观基因组，CRISPR/Cas9筛选，以及嵌合抗原受体(CAR)T细胞治疗，以确定 表观遗传修饰LRA的“休克”如何影响干细胞样CD8T细胞的程序和抗病毒免疫 是否可以采用干细胞样CD8T细胞的转录和表观遗传程序来增强 以T细胞为基础的免疫疗法“杀死”艾滋病病毒携带者。本研究的结果将为以后的研究奠定基础 用于实现艾滋病毒感染的长期缓解而不需要CART的新型免疫疗法。

Project Abstract Although combination antiretroviral therapy (cART) effectively suppresses human immunodeficiency virus (HIV) replication, it does not cure HIV infection and requires costly lifelong treatment. A major challenge for curing HIV infection is the long-lived latent HIV reservoir, which evades immune recognition and is responsible for viral rebound shortly after interruption of cART. Substantial research efforts have focused on eliminating latently infected cells through so-called “shock and kill” strategy, which reactivates latent HIV using latency- reversing agents (LRAs) to allow for the “kill” by cytolysis or immune-mediated clearance. Particularly, eradication of theHIV reservoir by anti-HIV T cells, which maintain antiviral immunity in patients as a “living” drug, presents a promising strategy to either fully resolve the infection or maintain long-term control without cART treatment. However, clearance of the HIV reservoir by T-cell based immunotherapy remains challenging because of 1) T- cell exhaustion, 2) the need of lifelong anti-HIV immunity to replace cART, 3) sanctuary sites like B cell follicles that exclude most HIV-specific CD8 T cells, and 4) unclear impact of LRAs, most of which target epigenetic pathways, on the function and differentiation of antiviral CD8 T cells in vivo. We and others have recently characterized a stem-like CD8 T cell subset in chronic lymphocytic choriomeningitis virus (LCMV), simian immunodeficiency virus (SIV), and HIV infections, as well as in mouse and human tumors. Compared to terminally exhausted CD8 T cells, stem-like CD8 T cells are less exhausted, mediate long-term immunity, and respond more potently after treatment of immunotherapies in animals and human. In chronic LCMV, SIV, and HIV infections, these cells express CXCR5, migrate to B cell follicles and kill infected T follicular helper cells, a major latent reservoir of HIV and SIV. In addition, frequency of these cells inversely correlates with viremia of SIV or HIV. Most recently, we showed that the single-cell transcriptomic and epigenetic profiles of stem-like CD8 T cells are distinct from other CD8 subsets generated after acute or chronic LCMV infection. In addition, we identified a transcriptional program involving transcription factor TOX that is essential for stem-like CD8 T cell differentiation and the long-term persistence of antiviral CD8 T cells during chronic viral infection. Here, I will determine the transcriptional and epigenetic programs of stem-like CD8 T cells required for optimal T-cell based immunotherapy against HIV. I will use animal models of chronic LCMV and SIV infections as well as samples from HIV patients, and employ cutting-edge technologies, including single-cell profiling of T-cell transcriptomes and epigenomes, CRISPR/Cas9 screening, and chimeric antigen receptor (CAR) T-cell therapy, to determine how “shock” by epigenetic modifying LRAs affects the program and antiviral immunity of stem-like CD8 T cells and whether transcriptional and epigenetic programs of stem-like CD8 T cells can be adopted to enhance the “kill” of the HIV reservoir by T-cell based immunotherapies. The results from this study will build the foundation for novel immunotherapies that achieve long-term remission from HIV infection without a need for cART.