Mechanisms Underlying Clearance of the Persistently Infected CNS

清除持续感染的中枢神经系统的潜在机制

基本信息

批准号：
7969711
负责人：
Dorian McGavern
金额：
$ 172.33万
依托单位：
NATIONAL INSTITUTE OF NEUROLOGICAL DISORDERS AND STROKE
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/7969711
关键词：
Address Adoptive Immunotherapy Adoptive Transfer Adult Antigen-Presenting Cells Antiviral Agents B-Lymphocytes Biological Models Birth Brain CD4 Positive T Lymphocytes CD80 gene CD8B1 gene Carrier State Cell Count Cells Central Nervous System Infections Central Nervous System Viral Diseases Clinical Dendritic Cells Exposure to HIV Health Human Image Immune Immune system Immunotherapeutic agent Immunotherapy Infection Interferon Type I Kidney Label Laboratories Life Lymphocyte Lymphocytic choriomeningitis virus Memory Microscopy Modeling Mus Neuraxis Neurons Outcome Peptides Peripheral Population Process Production T memory cell T-Lymphocyte Therapeutic Time Tissues Translating Treatment Protocols Up-Regulation Viral Viral Antigens Virus Virus Diseases cell injury cytotoxic follow-up immunopathology improved in utero in vivo interest migration pathogen prevent purge residence success tool two-photon

项目摘要

Persistent viruses, such as human immunodeficiency virus, cause major health problems worldwide and are extraordinarily difficult to clear following the establishment of persistence. Given the challenges associated with clearing persistent infections, it is important to develop and mechanistically understand therapeutic strategies that successfully achieve viral eradication without inducing permanent damage to the host. Studies using the lymphocytic choriomeningitis virus (LCMV) model system have demonstrated convincingly that a systemic persistent viral infection can be completely purged from a murine host by using a therapeutic approach referred to as adoptive immunotherapy. Remarkably, total body control of multiple persistent viral infections in both the mouse and humans can be achieved using adoptive immunotherapy. When mice are persistently infected at birth or in utero with LCMV (referred to as carrier mice), the virus establishes systemic persistence. Adult LCMV carrier mice are tolerant to the virus at the T cell level and thus are unable to eradicate the pathogen, which provides an excellent model to study immunotherapeutic regimens. Immunocytotherapy relies on the adoptive transfer of virus-specific memory CD8 and CD4 T cells from LCMV-immune donor mice into recipient carrier mice. Following the therapeutic administration of memory cells, LCMV is purged from most peripheral tissues of carrier mice in 14 days, whereas more than 100 days are required to clear virus from the central nervous system (CNS) and kidneys. Furthermore, successful viral clearance can be achieved with antiviral memory but not effector T cells. Thus, in addition to its proven therapeutic relevance, this model also provides a paradigm to understand factors that regulate memory T cells following secondary exposure to pathogens in vivo. Our laboratory is interested in mechanistically understanding how the immune system can be harnessed to resolve persistent viral infections, particularly those that take residence in the CNS. LCMV carrier mice provide an ideal model to study because every tissue, including the CNS, contains an abundance of viral antigen. Thus, following administration of therapeutic memory T cells, the mechanisms underlying clearance of a persistent viral infection can be studied in all tissues simultaneously. Clearance of the persistently infected CNS is of particular interest in LCMV carrier mice because neurons are the primary cell population infected within the brain parenchyma, and these cells are thought to express minimal to no MHC I. Nevertheless, adoptively transferred memory T cells achieve clearance of these cells with very little evidence of cellular injury. Our recent studies demonstrated that the success of adoptive immunotherapy in LCMV carrier mice relies on assistance from the recipients immune system. For example, we observed that migration of anti-viral cytotoxic lymphocytes (CTL) into the CNS coincided temporally with the arrival of recipient antigen-presenting cells (e.g. dendritic cells). We became particularly interested in dendritic cells (DCs) because they are not normally found in the uninflamed brain parenchyma; however, following adoptive immunotherapy, they not only migrated into the brain, but also presented viral peptides to therapeutic CTL. Using depletion studies, we revealed that recipient DCs were in fact required for the success of adoptive immunotherapy both in the periphery as well as the CNS. More recently, we exploited the involvement of DCs in this process by using a costimulation blocker approved for clinical use (i.e., CTLA-4-Fc) to modulate the activities of therapeutic memory T cells. We observed that within days of adoptive immunotherapy memory T cells induced the recruitment of DCs into the CNS as well as peripheral tissues. As expected, this migration was associated with an upregulation of the costimulatory molecules, CD80 and CD86, on the DCs. Previously, it was thought that memory T cells, unlike their nave counterparts, did not require costimulation to be reactivated. However, more recent studies, including ours, called this dogma into question by noting that memory T cells do in fact require costimulation for optimal proliferation and effector functions. Importantly, we demonstrated that costimulation blockade with CTLA-4-Fc could be used to control the proliferation of therapeutic memory T cells without impeding their ability to ultimately purge persistent virus. This finding has the potential to translate into an important therapeutic tool because memory T cells can cause severe (sometimes fatal) immunopathology. Lessening T cell numbers without impacting viral clearance should reduce unwanted immunopathology. We are presently in the process of following up on these exciting observations. The overall aim of this project is to define how therapeutic memory T cells coordinate with host innate immune cells to achieve clearance of the persistently infected CNS, particularly neurons. We demonstrated this past year that the host innate immune system is chronically stimulated by type I interferon production during a LCMV carrier state, and previous studies have demonstrated that type I interferons are required for the success of adoptive immunotherapy. We, therefore, propose that the dialogue between the innate and adaptive immune systems is critical for the eradication of a persistent viral infection. Using real time imaging approaches, we are now attempting to reveal the precise mechanism(s) by which the innate and adaptive immune systems purge neurons of virus. Despite the low level of MHC I expression, it is conceivable that CTL directly engage neurons. Alternatively, it is possible that CTL indirectly achieve clearance of neurons through their interactions with DCs. These hypotheses can be easily addressed by simultaneously visualizing in real time infected neurons, DCs, and CTL each labeled with distinct fluorescent tags. This will be accomplished by imaging the living brains of immunotherapy recipients using two photon microscopy. We are also interested in defining the mechanism(s) used by neurons to uniquely regulate the immune system and prevent unwanted immunopathology. We postulate that CTL can directly interact with CNS neurons but are uniquely regulated upon doing so. Harnessing regulatory mechanisms therapeutically might improve the outcome of CNS infections that have severe consequences in humans.

持续性病毒，例如人类免疫缺陷病毒，在全球范围内引起重大健康问题，并且在建立持久性后难以清除。鉴于与清除持续感染相关的挑战，重要的是要开发和机械理解的治疗策略，这些策略成功地消除了病毒性，而不会诱导宿主永久损害。使用淋巴细胞绒毛膜炎病毒（LCMV）模型系统的研究令人信服地证明，可以通过使用称为收养免疫疗法的治疗方法从鼠宿主中完全清除系统性的持续病毒感染。值得注意的是，可以使用过继的免疫疗法来实现小鼠和人类多种持续病毒感染的总体控制。当小鼠在出生时或用LCMV（称为载体小鼠）的子宫内持续感染时，该病毒会建立系统性持久性。成年LCMV载体小鼠在T细胞水平上耐受病毒的耐受性，因此无法消除病原体，该病原体为研究免疫治疗方案提供了出色的模型。免疫细胞疗法依赖于从LCMV-免疫供体小鼠中的病毒特异性记忆CD8和CD4 T细胞转移到受体载体小鼠中。记忆细胞的治疗性给药后，在14天内从载体小鼠的大多数外围组织清除LCMV，而从中枢神经系统（CNS）和肾脏清除病毒需要100天以上。此外，可以通过抗病毒记忆来实现成功的病毒清除率，而不能实现效应T细胞。因此，除了其可靠的治疗性相关性外，该模型还提供了一个范式，以了解在体内接触病原体后调节记忆T细胞的因素。我们的实验室有兴趣机械地了解如何利用免疫系统来解决持续的病毒感染，尤其是那些居住在中枢神经系统中的病毒感染。 LCMV载体小鼠提供了研究的理想模型，因为包括CNS在内的每个组织都有大量的病毒抗原。因此，在治疗记忆T细胞进行治疗后，可以同时研究所有组织中持续病毒感染的机制。在LCMV载体小鼠中，持续感染的CNS的清除尤其引起了人们的关注，因为神经元是脑实质内感染的原发性细胞群体，并且这些细胞被认为表现出最小的MHCI。不过，不过，不采用的记忆T细胞可清除这些细胞，几乎没有细胞损伤的证据。我们最近的研究表明，LCMV载体小鼠中收养免疫疗法的成功取决于受体免疫系统的帮助。例如，我们观察到，抗病毒细胞毒性淋巴细胞（CTL）迁移到中枢神经系统中，随着受体抗原抗原呈递细胞的到来（例如，树突状细胞）的到来。我们对树突状细胞（DC）特别感兴趣，因为它们通常在未发明的脑实质中发现。然而，在采继的免疫疗法之后，它们不仅迁移到大脑中，而且还向治疗性CTL呈现了病毒肽。使用耗竭研究，我们透露，实际上，受体DC在周围和中枢神经系统中都取养了免疫疗法所必需。最近，我们通过使用批准用于临床使用的共刺激阻滞剂（即CTLA-4-FC）来调节治疗记忆T细胞的活性，从而利用了DC在此过程中的参与。我们观察到，在收养免疫疗法记忆中，T细胞在几天内诱导DC募集到中枢神经系统和周围组织。如预期的那样，这种迁移与DC上的共刺激分子CD80和CD86的上调有关。以前，人们认为记忆T细胞与中殿的同类细胞不同，不需要重新激活共刺激。但是，包括我们的包括我们的教条在内的最新研究称记忆T细胞实际上需要进行最佳的增殖和效应子功能。重要的是，我们证明了与CTLA-4-FC的共刺激阻滞可用于控制治疗记忆T细胞的增殖，而不会阻碍其最终清除持久病毒的能力。这一发现有可能转化为重要的治疗工具，因为记忆T细胞会引起严重（有时是致命的）免疫病理学。减少T细胞数量而不会影响病毒清除率，应减少不良的免疫病理学。目前，我们正在跟进这些令人兴奋的观察结果。该项目的总体目的是定义治疗记忆T细胞如何与宿主先天免疫细胞坐标以清除持续感染的CNS，尤其是神经元。我们证明了过去的一年，在LCMV载体状态下，宿主先天免疫系统长期受到I型干扰素产生的刺激，先前的研究表明，I型干扰素是为了成功地采用免疫疗法而需要的。因此，我们建议先天和适应性免疫系统之间的对话对于消除持续的病毒感染至关重要。使用实时成像方法，我们现在正在尝试揭示先天和适应性免疫系统清除病毒神经元的精确机制。尽管MHC I表达的水平较低，但可以想象CTL直接吸引神经元。另外，CTL可能通过与DC的相互作用间接实现神经元的清除率。这些假设可以通过实时感染的神经元，DC和CTL同时可视化每个假设，每个假设都标有不同的荧光标签。这将通过使用两个光子显微镜成像免疫疗法受体的活体大脑来实现。我们还有兴趣定义神经元用来唯一调节免疫系统并防止不必要的免疫病理学的机制。我们假设CTL可以直接与CNS神经元相互作用，但在这样做时受到了唯一的调节。在治疗上利用监管机制可能会改善中枢神经系统感染的结果，这些感染对人类产生严重后果。