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.

项目概要 目前的抗逆转录病毒疗法可以有效抑制但不能根除 HIV-1 感染。组合时 抗逆转录病毒治疗 (cART)，通常可将 HIV-1 RNA 水平降低至 50 拷贝/毫升以下； 然而，使用超灵敏检测在血浆中检测到持续的低水平病毒血症。来源和 目前正在调查这种持续病毒血症的动态。 HIV-1 的一个明确定义的储存库是记忆 CD4+ T 细胞，当激活的 CD4+ T 细胞被 HIV-1 感染时，HIV-1 潜伏期就建立起来，但 转变为终末分化的记忆 T 细胞，而不是经历裂解感染。上调细胞 转录诱导 HIV 基因表达已被提议作为减少潜在病毒库的策略。 受感染的静息记忆 CD4+ T 细胞和可能携带病毒基因组整合形式的骨髓细胞。 这包括抑制细胞组蛋白脱乙酰酶 (HDAC)，因为这些 HDAC 通过以下方式促进潜伏期： 调节基因组结构和转录活性。 最近使用单剂量组蛋白对 8 名 HIV 感染者进行了抑制性 cART 治疗 脱乙酰酶抑制剂 (HDACi) 伏立诺他导致细胞相关未剪接 (CA-US) 显着增加 静息记忆 CD4+ T 细胞中的 HIV RNA。在澳大利亚的一项临床试验中，CA- 也出现了类似的显着增加 在接受 14 天的 HIV 感染患者血液 CD4+ T 细胞中观察到 HIV RNA HDACi 伏立诺他。丹麦 HDACi 帕比司他多剂量研究的初步数据也 表明 CA-US HIV RNA 增加。 鉴于这些发现，我们假设：1) HDACi 治疗后 CA-US HIV RNA 的序列将 是克隆的，这与受感染的静息记忆 T 细胞子集的转录一致 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) 加拿大-美国艾滋病毒 RNA 在遗传上与治疗前（cART 前和 HDACi 前）血浆来源的 HIV-1 序列相似， 表明该储库是在 cART 启动之前建立的； 4) 如果病毒在分析治疗期间反弹 帕比司他治疗中断后，它将与 HDACi 期间重新激活的 CA-US HIV RNA 相似 治疗。这些假设将在以下具体目标的实验中得到解决：（1）确定 多组治疗前后患者 CD4+ T 细胞的 CA-US HIV RNA 的基因组成和多样性 用伏立诺他或帕比司他进行剂量治疗； (2) 确定未剪接的基因组成和多样性 骨髓细胞和特定记忆 CD4+ T 细胞亚群中的 HIV-1 RNA； (3) 比较基因组成 将 CA-US HIV RNA 转化为治疗前样本和治疗后样本中血浆衍生的 HIV-1 序列 帕比司他治疗后分析治疗中断。 如果这项探索性研究揭示了骨髓细胞中 CA-US HIV RNA 的克隆扩增和特定的静息记忆 来自使用 HDAC 抑制剂的 HIV-1 感染患者的 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前病毒潜伏期，并且艾滋病毒潜伏期可以在人类中进行治疗。