Evaluating the role of the novel apoptosis inhibitor TRAILshort, in maintaining HIV persistence.

评估新型细胞凋亡抑制剂 TRAILshort 在维持 HIV 持久性方面的作用。

• 批准号: 8990167
• 负责人: ANDREW D BADLEY
• 金额: $ 50.57万
• 依托单位: MAYO CLINIC ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-06-15 至 2020-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8990167
• 关键词: Acute; Antiviral Agents; Apoptosis; Apoptosis Inhibitor; Apoptotic; Autologous; B-Lymphocytes; Binding; Biological Assay; CD4 Positive T Lymphocytes; CD8B1 gene; Caspase; Caspase-1; Cell Count; Cell Death; Cell Death Signaling Process; Cell Line; Cell Survival; Cell model; Cell physiology; Cells; Cessation of life; Coculture Techniques; Complex; DNA; DNA-dependent protein kinase; Data; Death Receptor 5; Dendritic Cells; Disease Progression; Dominant-Negative Mutation; Failure; Goals; Granzyme; HIV; HIV Infections; Herpesvirus 1; Homologous Gene; Human; Human Herpesvirus 4; Human Herpesvirus 8; Human Papillomavirus; IL6 gene; Immune; Immune response; Immune system; Immunologic Surveillance; Immunologics; Impairment; In Vitro; Integrase; Interferon-alpha; Interleukin-6; Intervention; Length; Ligand Binding; Ligands; Malignant - descriptor; Malignant Neoplasms; Measures; Mediating; Names; Natural Killer Cells; Nested PCR; Outer Mitochondrial Membrane; Pathogenesis; Pathway interactions; Patients; Peripheral Blood Mononuclear Cell; Plasma; Play; Production; Proteins; Proviruses; RNA Splicing; Regulation; Research; Research Proposals; Resistance; Role; Signal Transduction; Stimulus; Subfamily lentivirinae; System; T-Lymphocyte; TNF gene; TNF-related apoptosis-inducing ligand; Testing; Transcript; Tumor Necrosis Factor Ligand Superfamily Member 6; Variant; Viral; Virion; Virus; Virus Diseases; Work; apoptosis inducing factor; base; cancer cell; cell killing; clinically relevant; cohort; cytotoxic; cytotoxicity; death receptor-4; design; genetic approach; human TNFRSF10A protein; immune activation; in vivo; inhibitor/antagonist; innovation; insight; interest; killings; knock-down; macrophage; novel; novel strategies; prevent; public health relevance; purge; reactivation from latency; receptor; response; sensor; small hairpin RNA; tool

 DESCRIPTION (provided by applicant): The interplay between a pathogenic viral infection and the human host reflects a bi-directional interaction of host sensors (e.g. TLR, RIG, etc.) which detect virus infection, host effector mechanisms which eliminate infected cells (e.g. caspase activation, expression of death inducing ligands including Fas Ligand and TRAIL), and viral adaptations which attempt to subvert the host response (e.g. virally encoded apoptosis inhibitors e.g. EBV encoded BALF1, KSHV encoded vFLIP etc.). Experimental elimination of virally encoded apoptosis inhibitors enhances host cell clearance of virally infected cells, and reduces viral persistence - as examples: inhibiting BALF1 prevents EBV persistence, inhibiting LAT inhibits HSV1 persistence, and disrupting E6 or E7 inhibits HPV persistence. HIV infection can be sensed in human cells for example by IFI-16 sensing of accumulated reverse transcripts leading to caspase 1 dependent pyroptosis, or DNA-PK sensing of integrase mediated host DNA "nicking" leading to p53 dependent apoptosis. However these sensing mechanisms do not eradicate all HIV infected cells, and HIV is as a result, persistent. HIV is not known to encode a protein which antagonizes the hosts ability to eradicate infected cells, yet accumulating evidence suggests that HIV infected cells are more resistant to cell death that corresponding uninfected cells, through an as yet undefined mechanism TNF related apoptosis inducing factor (TRAIL) is a molecule whose principal function is as an effector of immune surveillance, and it has been implicated in the pathogenesis of malignancies, as well as viral infections including HIV. Concerning the role of TRAIL in HIV, considerable data indicate that TRAIL is dysregulated during HIV infection in vivo, and there is ample evidence that treatment of cells from ART suppressed HIV- infected patients with exogenous TRAIL, reduces the number of latently infected cells, as measured by undetectable levels of replication competent virus in quantitative co-culture assays. The current research proposal concerns a novel TRAIL splice variant, which we have discovered and named TRAILshort, which is produced during HIV infection in vitro and in vivo. TRAILshort binds to TRAIL receptor 2 (TRAILR2), yet does not transmit a cell death signal, whereas TRAIL binding to TRAILR2 rapidly results in apoptotic death. Moreover, expression of TRAILshort, prevents TRAIL from engaging TRAILR2, demonstrating that TRAILshort acts as a dominant negative inhibitor of TRAIL mediated killing. Because NK cells and CD8+ CTL kill target cells by TRAIL (in addition to other mechanisms such as Granzyme B), it follows that TRAILshort expression reduces both NK cell and CD8 T cell cytotoxicity. Our underlying hypothesis is that TRAILshort production during HIV infection prevents TRAIL dependent HIV clearance mechanisms which allows HIV infected cells to persist. This hypothesis is supported by novel preliminary data included in this application, where shRNA mediated knockdown of TRAILshort does not alter uninfected T cell survival, yet selectively enhances killing of HIV infected cells, thereby reducing HIV replication by 3.5 logs, and decreasing the number of cells containing HIV DNA. The long-term goal of our work is to fully understand and manipulate TRAILshort production in order that normal immune mechanisms can contribute to clearance of virally infected cells, thereby providing an additional tool with which to contribute towards a cure for HIV. This will be fully explored by: (i) studying the regulation of TRAILshort, in order to identify means of inhibiting its production. (ii) Optimize lentivirus knock down of TRAILshort in primary CD4 T cells from HIV infected patients, and test whether HIV reactivation with TRAILshort knockdown, plus or minus co-culture with autologous CD8 T cells, or NK cells, reduces HIV reservoir size ex vivo (iii) Evaluate TRAILshort expression in HIV elite controllers, partial controllers and non-controllers, and determine the association of TRAILshort expression with HIV reservoir size.

 描述（由适用提供）：致病病毒感染与人类宿主之间的相互作用反映了宿主传感器的双向相互作用（例如TLR，钻机等），该相互作用检测到病毒感染，宿主效应机制，消除了感染细胞的宿主效应机制病毒编码的凋亡抑制剂，例如EBV编码的BALF1，KSHV编码的VFLIP等。实验消除虚拟编码的凋亡抑制剂可以增强虚拟细胞的宿主细胞清除，并降低病毒持久性 - 作为示例：抑制BALF1阻止EBV持久性，抑制LAT抑制HSV1抑制HSV1持久性，并破坏E7或E7抑制HPV持久性。例如，可以通过IFI-16的IFI-16敏感性来感知HIV感染，导致caspase 1依赖性凋亡的累积逆转录，或对综合介导的宿主DNA“刻痕”的DNA-PK敏感性，从而导致p53依赖性凋亡。但是，这些感应机制并不能放射所有受HIV感染的细胞，因此HIV持续存在。尚不知道HIV编码一种使宿主拮抗放射性感染细胞的能力的蛋白质，但积累的证据表明，HIV感染的细胞对与未感染的细胞相对应的细胞死亡具有更大的抵抗力，该细胞通过TNF相关的凋亡因子（TRAIL）是一种分子，并且具有效应的效果，并且具有效应的效果，并且具有效应的效果，并且具有效应的效果。恶性肿瘤以及包括艾滋病毒在内的病毒感染。同意TRAIL在HIV中的作用，这表明在体内HIV感染期间TRAIL失调，并且有充分的证据表明，从ART抑制的ART抑制细胞的外源性trail的细胞的治疗可以减少潜在受感染细胞的数量，这是通过在定量coulture sass中的不可检测的复制病毒水平来测量的。当前的研究建议涉及一种新型的Trail Splice变体，我们发现并命名为Trailshort，该变体是在艾滋病毒感染期间在体外和体内产生的。 Trailshort与TRAIL接收器2（TRAILR2）结合，但不会传输细胞死亡信号，而Trail与TrailR2结合迅速导致凋亡死亡。此外，Trailshort的表达可防止Trail与TrailR2吸引，这表明Trailshort是Trail介导的杀戮的主要负面抑制剂。由于NK细胞和CD8+ CTL通过TRAIL杀死靶细胞（除其他机制（例如颗粒酶B）之外，因此Trailshort表达降低了NK细胞和CD8 T细胞细胞毒性。我们的基本假设是，在HIV感染期间的Trailshort产生可防止依赖性的HIV清除机制，从而允许HIV感染的细胞持续存在。该假设得到了本应用程序中包含的新初步数据的支持，在该数据中，SHRNA介导的Trailshort的敲低不会改变未感染的T细胞存活，而是选择性地增强了HIV感染细胞的杀死，从而减少了3.5对日志的HIV复制，并减少了含有HIV DNA的细胞的数量。我们工作的长期目标是充分理解和操纵步道生产，以便正常的免疫组机制可以有助于清除几乎受感染的细胞，从而提供了一种额外的工具，可以为治愈HIV治愈。 （i）研究步伐的调节，以确定抑制其产量的方法，对此进行了充分的探讨。 (ii) Optimize lentivirus knockdown of TRAILshort in primary CD4 T cells from HIV infected patients, and test whether HIV reactivation with TRAILshort knockdown, plus or minus co-culture with autologous CD8 T cells, or NK cells, reduces HIV reservoir size ex vivo (iii) Evaluate TRAILshort expression in HIV elite controllers, partial controllers and non-controllers, and determine the association of 带有艾滋病毒储层大小的径流表达。