Role of Viral Reservoirs in the Pathogenesis of HIV Disease

病毒库在 HIV 疾病发病机制中的作用

• 批准号: 8745373
• 负责人: Anthony S. Fauci
• 金额: $ 99.79万
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 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/8745373
• 关键词: Antigen Receptors; Autologous; Back; CD19 Antigens; CD28 gene; CD3 Antigens; CD4 Positive T Lymphocytes; CD8B1 gene; Cancer Patient; Cell Count; Cell Line; Cells; Chronic Lymphocytic Leukemia; Coculture Techniques; Coupled; Cytotoxic T-Lymphocytes; Data; Detection; Disease; Disease Progression; Disease remission; Effector Cell; Enzyme-Linked Immunosorbent Assay; Escape Mutant; Frequencies; Goals; HIV; HIV Infections; Immune; Immunity; Immunologics; Individual; Infusion procedures; Interferon Type II; Longevity; Lymphoid Tissue; Macrophage Inflammatory Protein-1; Monitor; Pathogenesis; Patients; Plasma; Research; Residual state; Rest; Role; Site; Specificity; T-Lymphocyte; TNF gene; Therapeutic; Time; Viral; Viral Antigens; Viremia; Virus; antiretroviral therapy; base; chemokine; cytokine; cytotoxic; in vivo; interest; killings; patient population; peripheral blood; receptor; therapeutic development

Over a decade ago, we demonstrated that the latent viral reservoir in the resting CD4+ T cell compartment persists in virtually all HIV-infected individuals receiving clinically effective ART. In addition, we demonstrated that HIV continually replicates at low levels in chronically infected individuals who are consistently aviremic during prolonged periods of receiving ART. Consequently, the persistent viral reservoir has become a major impediment to the eradication of HIV in infected individuals receiving ART. In recent years, major research efforts have been dedicated to a better understanding of the pathogenesis of persistent HIV infection and to the development of therapeutic strategies aimed at eradicating virus in infected individuals receiving ART. During the past year, we have focused our research on: 1) examining the effect of ART on HIV reservoirs in elite controllers and 2) developing immune-based therapeutic strategies that target persistently infected CD4+ T cells. First, we investigated the effect of ART on the persistent HIV reservoirs in elite controllers. Although ART suppresses plasma viremia to below the limit of detection in the vast majority of HIV-infected individuals, the virus cannot be eradicated by therapy alone due in part to the persistence of viral reservoirs in the peripheral blood, lymphoid tissues, and other sequestered sites. Consequently, there is a growing interest in developing therapeutic strategies to eliminate persistent HIV reservoirs and/or to enhance host immunity against the virus in order to control viral replication in the absence of ART. In this regard, it has been shown that a small proportion of HIV-infected individuals spontaneously control plasma viremia in the absence of ART (controllers). Moreover, a subset of such individuals (elite controllers) are capable of maintaining relatively normal CD4+ T cell counts and undetectable levels of plasma viremia for years to decades without the need for ART. Although previous studies have suggested that ongoing viral replication may occur in elite controllers, precise frequencies of infected CD4+ T cells carrying replication-competent virus and/or the impact of ART on immunologic and virologic parameters have not been fully delineated in this patient population. Therefore, we conducted a study in order to determine the effect of ART on the dynamics of persistent HIV reservoirs and various immunologic parameters in elite controllers. We demonstrated that a short course (9 months) of ART dramatically decreased the infectious HIV burden in elite controllers, providing compelling evidence that onging/residual viral replication occurs in such aviremic individuals. The infectious HIV burden in the CD4+ T cell compartment of the elite controllers rebounded back to their original baseline levels upon cessation of ART, an indication that a virologic set-point exists even at this very low level of viral replication, and that host immunity can efficiently control plasma viremia without completely eliminating the infected CD4+ T cells. We also demonstrated that the level of HIV-specific CD8+ T cells in elite controllers gradually declined upon initiation of ART, further supporting the concept that low levels of viral replication occur in these infected individuals. Our data provide compelling evidence that ongoing HIV replication occurs in elite controllers. Given that these individuals effectively control viral replication and lack disease progression in the absence of ART, our study has implications for the feasibility of attaining sustained virologic remission in non-controllers without complete suppression or elimination of viral replication in the face of discontinuation of ART. Second, we investigated the feasibility of eliminating HIV-infected CD4+ T cells using autologous CD8+ T cells that express a chimeric antigen receptor (CAR). Recent studies have demonstrated that complete remission can be achieved in patients with chronic lymphoid leukemia upon infusion of autologous CD8+ T cells expressing CAR with specificity for the B cell antigen CD19 coupled with CD3-zeta, and costimulatory molecules. Inspired by these studies, we investigated whether autologous CD8+ T cells expressing CAR specific for HIV Env could eliminate persistently infected CD4+ T cells from HIV-infected individuals receiving ART. Three CAR constructs were evaluated. All constructs included CD4 for recognition of HIV Env on infected targets, CD3-zeta for cytotoxic activity, and one of the following constimulatory molecules: mock, 4-1BB, or CD28. Autologous CD8+ T cells expressing the above CAR proteins were cocultured with pre-stimulated CD4+ T cells from HIV-infected individuals receiving ART and the level of viral replication was monitored by p24 ELISA over time. Our preliminary data suggest that the highest level of HIV suppression can be achieved when CD8+ T cells expressing the CAR that contains CD28 are cocultured with infected target cells. Our data also show that CD8+ T cells expressing the above CAR construct produce higher levels of intracellular cytokines and chemokines (such as IFN-g, TNF-a, MIP-1) compared to those expressing CAR with mock or 4-1BB upon incubation with a cell line expressing HIV Env. Taken together, our preliminary data suggest that the elimination of persistently infected CD4+ T cells from HIV-infected individuals ex vivo can be achieved with autologous CD8+ T cells that express a CAR that recognizes HIV Env. Because CD8+ cytotoxic T cells do not require viral antigen specificity (besides recognition of HIV Env) for killing, such effector cells could potentially eliminate infected target cells without inducing viral escape mutants. In addition, considering the longevity of CAR-transduced CD8+ T cells observed in cancer patients (years to decades), these cells could restrict viral replication in vivo for extended periods of time.

十多年前，我们证明了静息CD 4 + T细胞区室中的潜伏病毒库在几乎所有接受临床有效ART的HIV感染个体中持续存在。此外，我们证明了HIV在长期接受ART期间持续存在的慢性感染个体中持续低水平复制。因此，持续的病毒储库已经成为在接受ART的受感染个体中根除HIV的主要障碍。近年来，主要的研究工作致力于更好地了解持续性HIV感染的发病机制，并开发治疗策略，在过去的一年中，我们的研究重点是：1）检查ART对精英控制者中HIV储库的影响，2）开发针对持续感染的CD 4 + T细胞的基于免疫的治疗策略。 首先，我们调查了ART对精英控制者中持续存在的HIV库的影响。尽管ART在绝大多数HIV感染者中将血浆病毒血症抑制至检测限以下，但病毒不能通过单独治疗根除，部分原因是外周血、淋巴组织和其他隔离部位中病毒储库的持续存在。因此，有越来越多的兴趣在开发治疗策略，以消除持久的HIV水库和/或增强宿主对病毒的免疫力，以控制病毒复制在ART的情况下。在这方面，它已被证明，一小部分的HIV感染的个人自发控制血浆病毒血症在ART（控制器）的情况下。此外，这些人的一个子集（精英控制者）能够维持相对正常的CD 4 + T细胞计数和不可检测的血浆病毒血症水平数年至数十年而不需要ART。尽管先前的研究表明，精英控制者中可能发生持续的病毒复制，携带可复制病毒的受感染CD 4 + T细胞的精确频率和/或或ART对免疫学和病毒学参数的影响尚未在该患者人群中完全描述。因此，我们进行了一项研究，以确定ART对持续的HIV水库和精英控制器中的各种免疫参数的动态的影响。我们证明，短期（9个月）的ART显着降低了精英控制者的感染性HIV负担，提供了令人信服的证据表明，在这种病毒血症个体中发生了持续/残留病毒复制。在停止ART后，精英控制者的CD 4 + T细胞区室中的感染性HIV负荷反弹回到其原始基线水平，这表明即使在这种非常低的病毒复制水平下也存在病毒学设定点，并且宿主免疫可以有效地控制血浆病毒血症而不完全消除感染的CD 4 + T细胞。我们还证明，精英控制者的HIV特异性CD 8 + T细胞水平在开始ART后逐渐下降，进一步支持了这些感染个体中发生低水平病毒复制的概念。我们的数据提供了令人信服的证据表明，正在进行的艾滋病毒复制发生在精英控制者。鉴于这些人有效地控制病毒复制和缺乏疾病进展的情况下，ART，我们的研究有可能实现持续的病毒学缓解的非控制者没有完全抑制或消除病毒复制的ART停止。 其次，我们研究了使用表达嵌合抗原受体（CAR）的自体CD 8 + T细胞消除HIV感染的CD 4 + T细胞的可行性。最近的研究表明，在输注表达CAR的自体CD 8 + T细胞后，慢性淋巴细胞白血病患者可以实现完全缓解，所述CAR对与CD 3-zeta偶联的B细胞抗原CD 19和共刺激分子具有特异性。受这些研究的启发，我们研究了表达HIV Env特异性CAR的自体CD 8 + T细胞是否可以消除接受ART的HIV感染个体中持续感染的CD 4 + T细胞。评估了三种CAR构建体。所有构建体均包括用于识别感染靶标上的HIV Env的CD 4、用于细胞毒性活性的CD 3-zeta以及以下共刺激分子之一：模拟物、4-1BB或CD 28。将表达上述CAR蛋白的自体CD 8 + T细胞与来自接受ART的HIV感染个体的预刺激的CD 4 + T细胞共培养，并通过p24 ELISA随时间监测病毒复制水平。我们的初步数据表明，当表达含有CD 28的CAR的CD 8 + T细胞与感染的靶细胞共培养时，可以实现最高水平的HIV抑制。我们的数据还显示，表达上述CAR构建体的CD 8 + T细胞在与表达HIV Env的细胞系孵育后，与表达CAR与模拟物或4-1BB的那些相比，产生更高水平的细胞内细胞因子和趋化因子（如IFN-g、TNF-α、MIP-1）。综上所述，我们的初步数据表明，用表达识别HIV Env的CAR的自体CD 8 + T细胞可以实现从HIV感染个体中体外消除持续感染的CD 4 + T细胞。因为CD 8+细胞毒性T细胞不需要病毒抗原特异性（除了识别HIV Env）来杀伤，所以这种效应细胞可以潜在地消除感染的靶细胞而不诱导病毒逃逸突变体。此外，考虑到在癌症患者中观察到的CAR转导的CD 8 + T细胞的寿命（数年至数十年），这些细胞可以在体内长时间限制病毒复制。