Project 1: Perturbation of Pre-Existing Immunity to Chronic Viral Infection Through Immunotherapy

慢性病毒感染仍然是全球健康的主要威胁，病原体如B肝炎病毒（HBV） 和丙型肝炎病毒（HCV），每年造成数百万人死亡。治疗药物的最新发展 HCV的抗病毒治疗已经是一个重大突破。然而，能够产生至少 慢性HBV的功能性治疗是迫切需要的。治疗HBV的一个主要障碍是功能上的 以衰竭为特征的免疫反应受损。PD-1：PD-L1/2抑制性受体 信号通路调节细胞免疫的许多关键方面，包括慢性病毒感染中的T细胞耗竭 和癌症阻断这一途径在晚期癌症的治疗中产生了显著的效果， 引发了免疫革命然而，关于PD-1在人体内的作用知之甚少 阻断对慢性感染的反应，其特征是疲惫和这些机制， 答案是否定的。在我们的U19 CCHI联盟的重要工作的基础上， 慢性HCV中的免疫耗竭并评估其在慢性抗原刺激终止后的逆转， 我们现在将全面研究PD-1作为抗原的关键介体如何直接和特异性阻断， 介导的免疫衰竭，影响抗病毒免疫应答的分子调节层。的 该项目的总体假设是，阻断人类PD-1将改变PD-1的大小、质量、调节和表达。 组合物的预先存在的抗病毒CD 8 T细胞反应，导致更有效的病毒控制，并将 调节细胞免疫的其他方面，包括非典型T细胞和巨噬细胞反应。我们将测试 这一假设通过以下目的。具体来说，在目标1中，我们将测试PD-1治疗如何改变治疗前的 针对血液中持续存在的病毒的现有病毒特异性CD 8 T细胞应答。我们将利用大型 从白细胞分离术中获得的PBMC捐献物用于表型变化的全面深入分析， 功能、克隆组成、转录状态和表观遗传调控，以及CMV-、EBV-、HBV- 和流感特异性CD 8 T细胞，以及非常规MAIT T细胞。在目标2中，我们将测试PD-1如何 治疗改变慢性B型肝炎患者肝脏中HBV特异性CD 8 T细胞应答。我们将 通过肝脏细针直接从感染部位获得的平行数据补充了我们在aim 1中的分析 吸气。最后，在目标3中，我们将定义通过人中的PD-1阻断恢复巨噬细胞 慢性HBV感染。这些数据将把我们对PD-1阻断的影响的分析扩展到其他方面。 通过研究巨噬细胞作为关键调节剂的反应， 肝内先天免疫总的来说，我们希望这些数据能大大提高我们对 PD-I介导的免疫恢复机制不仅可用于病原体的设计， 这不仅有助于特异性免疫疗法，而且还能够进一步改善一般免疫疗法的功效。

Chronic viral infections remain major threats to global health, with pathogens such as hepatitis B virus (HBV) and hepatitis C virus (HCV), responsible for millions of deaths annually. The recent development of curative antiviral therapy for HCV has been a major breakthrough. However, therapies capable of producing at least functional cure for chronic HBV are urgently needed. A major impediment to cure of HBV is a functionally impaired immune response characterized by markers of exhaustion. The PD-1:PD-L1/2 inhibitory receptor pathway regulates many key aspects of cellular immunity, including T cell exhaustion in chronic viral infection and cancer. Blockade of this pathway has produced dramatic effects in the treatment of advanced cancer and unleashed an immunotherapeutic revolution. However, little is known about the human in vivo effect of PD-1 blockade on the response to chronic infections marked by exhaustion and the mechanisms by which these responses are invigorated. Building on our U19 CCHI Consortium’s important work defining the mechanisms of immune exhaustion in chronic HCV and assessing its reversal upon termination of chronic antigen stimulation, we will now comprehensively investigate how direct and specific blockade of PD-1, as a key mediator of antigen- mediated immune exhaustion, affects the layers of molecular regulation of the antiviral immune response. The overall hypothesis of this project is that blocking PD-1 in humans will alter the magnitude, quality, regulation and composition of pre-existing antiviral CD8 T cell responses, leading to more effective viral control, and will modulate other aspects of cellular immunity, including atypical T cell and macrophage responses. We will test this hypothesis through the following aims. Specifically, in Aim 1 we will test how PD-1 therapy alters pre- existing virus-specific CD8 T cell responses targeting persisting viruses in the blood. We will utilize large PBMC donations from leukapheresis for a comprehensive and in-depth analysis of changes in the phenotype, function, clonal composition, transcriptional state, and epigenetic regulation of HBV-, but also CMV-, EBV-, HBV- and influenza-specific CD8 T cells, as well as of unconventional MAIT T cells. In Aim 2 we will test how PD-1 therapy alters HBV-specific CD8 T cell responses in the liver of patients with chronic hepatitis B. We will complement our analyses from aim 1 with parallel data directly from the site of infection through liver fine needle aspirates. Finally, in Aim 3 we will define the recovery of macrophages through PD-1 blockade in persons with chronic HBV infection. These data will extend our analysis on the effects of PD-1 blockade to other components of the antiviral immune response, by studying the response of macrophages as critical modulators of intrahepatic innate immunity. Collectively, we expect these data to dramatically enhance our understanding of the mechanism of PD-1 mediated immune recovery that can be utilized not only for the design of pathogen- specific immunotherapy, but also to enable further improvements in the efficacy of immunotherapy in general.