Genetic analysis of unspliced HIV RNA produced during HDAC inhibitor therapy

HDAC 抑制剂治疗期间产生的未剪接 HIV RNA 的遗传分析

• 批准号: 8730254
• 负责人: Sarah Elizabeth Palmer
• 金额: $ 15.66万
• 依托单位: UNIVERSITY OF SYDNEY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-03-01 至 2016-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8730254
• 关键词: Address; Australia; Biological Assay; Blood; CD4 Positive T Lymphocytes; Cells; Clinical Trials; Clonal Expansion; Collaborations; Conduct Clinical Trials; Data; Denmark; Dose; Enrollment; Gene Expression; Genetic; Genetic Transcription; Genome; Gut associated lymphoid tissue; HIV; HIV-1; Histone Deacetylase Inhibitor; Histones; Human; Individual; Infection; Interruption; Investigation; Lytic Phase; Memory; Myeloid Cells; Oral; Patients; Phylogenetic Analysis; Plasma; Production; RNA; RNA Sequences; Regimen; Research; Rest; Sampling; Sorting - Cell Movement; Source; Structure; T memory cell; T-Lymphocyte Subsets; Time; Universities; University Hospitals; Up-Regulation; Viral; Viral Genome; Viremia; Virus; Vorinostat; antiretroviral therapy; cell type; effective therapy; genetic analysis; inhibitor/antagonist; memory CD4 T lymphocyte; peripheral blood; public health relevance; research study

PROJECT SUMMARY Current antiretroviral therapy effectively suppresses but does not eradicate HIV-1 infection. During combination antiretroviral therapy (cART), reduction of HIV-1 RNA levels to less than 50 copies/ml is frequently achieved; however, persistent low-level viremia has been detected in plasma using ultrasensitive assays. The source and dynamics of this persistent viremia is currently under investigation. One well-defined reservoir of HIV-1 is memory CD4+ T cells, where HIV-1 latency is established when an activated CD4+ T cell becomes infected by HIV-1 but transitions to a terminally differentiated memory T cell instead of undergoing lytic infection. Upregulating cellular transcription to induce HIV gene expression has been proposed as a strategy for reducing the pool of latently- infected resting memory CD4+ T cells and possibly myeloid cells carrying an integrated form of the viral genome. This includes inhibiting cellular histone deacetylases (HDACs) because these HDACs promote latency by regulating genome structure and transcriptional activity. A recent treatment of eight HIV-infected patients on suppressive cART with a single dose of the histone deacetylase inhibitor (HDACi), vorinostat, resulted in a significant increase in cell associated unspliced (CA-US) HIV RNA in resting memory CD4+ T cells. During a clinical trial in Australia, a similar significant increase in CA- US HIV RNA was observed in CD4+ T-cells in blood from HIV-infected patients treated with 14 days of the HDACi vorinostat. Preliminary data from a multidose study of the HDACi panobinostat in Denmark also demonstrates an increase in CA-US HIV RNA. Given these findings, we hypothesize that: 1) the sequence of CA-US HIV RNA following HDACi treatment will be clonal which is consistent with transcription from a subset of infected resting memory T cells responsive to HDAC inhibitors in the blood and gut; 2) specific cell types are producing CA-US HIV RNA; 3) this CA-US HIV RNA will be genetically similar to pre-therapy (pre-cART and pre-HDACi) plasma-derived HIV-1 sequences, indicating this reservoir was established prior to cART initiation; 4) if virus rebounds during an analytical treatment interruption post-panobinostat therapy, it will be genetically similar to CA-US HIV RNA reactivated during HDACi therapy. These hypotheses will be addressed in the experiments of the following Specific Aims: (1) to determine the genetic makeup and diversity of CA-US HIV RNA from CD4+ T cells of patients prior to and following multi- dose treatment with vorinostat or panobinostat; (2) to determine the genetic makeup and diversity of unspliced HIV-1 RNA in myeloid cells and specific memory CD4+ T cell subsets; and (3) to compare the genetic makeup of CA-US HIV RNA to plasma-derived HIV-1 sequences from pre-therapy samples and samples taken after an analytical treatment interruption following panobinostat therapy. If this exploratory study reveals clonal expansion of CA-US HIV RNA in myeloid cells and specific resting memory T cells from HIV-1 infected patients on HDAC inhibitors, this would provide direct evidence that these compounds are disrupting proviral latency within these cells and that HIV latency can be therapeutically targeted in humans.

项目摘要 当前的抗逆转录病毒治疗可有效抑制，但不能消除HIV-1感染。在组合期间 经常实现抗逆转录病毒疗法（CART），HI​​V-1 RNA水平降低至少于50份/mL； 但是，使用超敏化测定法在血浆中检测到持续的低级病毒血症。来源和 目前正在研究这种持续性病毒血症的动力学。 HIV-1的一个定义明确的水库是内存 CD4+ T细胞，其中当激活的CD4+ T细胞被HIV-1感染时，建立HIV-1潜伏期 过渡到末端分化的记忆T细胞，而不是经历裂解感染。上调细胞 已经提出了诱导HIV基因表达的转录作为减少潜伏的库的一种策略 感染的静息记忆CD4+ T细胞以及可能具有综合形式的病毒基因组形式的髓样细胞。 这包括抑制细胞组蛋白脱乙酰基酶（HDAC），因为这些HDAC促进了通过 调节基因组结构和转录活性。 最近在抑制车上对八名HIV感染患者的治疗 脱乙酰基酶抑制剂（HDACI），Vorinostat，导致与细胞相关的无斑（CA-US）显着增加（CA-US） HIV RNA在静止的记忆CD4+ T细胞中。在澳大利亚的一项临床试验中，CA-的显着增长类似 在接受14天治疗的HIV感染患者的血液中，在CD4+ T细胞中观察到US HIV RNA hdaci vorinostat。来自丹麦HDACI Panobinostat的多蛋白研究的初步数据也 证明CA-US HIV RNA的增加。 鉴于这些发现，我们假设：1）HDACI处理后CA-US HIV RNA的序列将 是克隆纳尔，这与从感染的静息记忆T细胞的子集中转录一致 血液和肠道中的HDAC抑制剂； 2）特定细胞类型正在产生CA-US HIV RNA； 3）此CA-US HIV RNA在遗传上类似于疗法（前卡特和HDACI前）等离子体衍生的HIV-1序列， 表明该储层是在货车启动之前建立的； 4）如果病毒在分析治疗过程中反弹 中断后苯并症治疗后，它将与HDACI期间重新激活的CA-US HIV RNA在遗传上相似 治疗。这些假设将在以下特定目的的实验中解决：（1）确定 在多个患者的CD4+ T细胞中，CA-US HIV RNA的遗传构成和多样性在多个患者之前和之后 用伏立诺替塔或panobinostat治疗剂量； （2）确定未填充的基因组成和多样性 髓样细胞中的HIV-1 RNA和特定的记忆CD4+ T细胞子集； （3）比较基因组成 从疗法样品和样品中采集的CA-US HIV RNA到血浆衍生的HIV-1序列 panobinostat疗法后的分析治疗中断。 如果这项探索性研究揭示了髓样细胞中CA-US HIV RNA的克隆膨胀和特定的静止记忆 来自HIV-1感染HDAC抑制剂患者的T细胞，这将提供直接证据表明这些化合物 正在破坏这些细胞内的病毒潜伏期，并且在人类中可以治疗艾滋病毒潜伏期。