HTS for Synergistic Activators of Latent HIV-1 Infection

描述(申请人提供)：HIV-1潜伏期，允许病毒在抗逆转录病毒治疗存在的情况下持续存在，可能是开发HIV-1感染根治疗法所要克服的主要障碍。虽然很明显需要根除潜伏的HIV-1病毒宿主，但如何实现这一点尚不清楚。在分子生物学水平上，目前的HIV-1研究集中在对潜伏HIV-1感染的分子控制必须不同于对可诱导的细胞基因的控制，并且其特点是在病毒LTR处存在限制性的染色质环境。但是，当潜伏的病毒整合到主动表达的宿主基因中时，这种限制性的染色质环境是如何形成的尚不清楚。当一个复杂的限制性染色质结构被认为屏蔽了潜伏的病毒LTR时，如何在潜伏的病毒LTR上发现暂停的RNAP II？这种对限制性染色质环境的关注，作为HIV-1潜伏感染的分子控制机制，转化为药物发现和临床应用，其中HDAC抑制剂被视为HIV-1重新激活策略的圣杯。然而，仔细阅读文献后，无论是在许多潜伏感染模型中，还是在体外实验或临床研究中，作为HIV-1再活化剂的HDAC抑制剂都没有显示出令人信服的疗效。可能的例外是SAHA，临床批准为HDAC抑制剂涡流调节剂。SAHA在包括我们在内的大多数实验系统中都显示出一些HIV-1重新激活的能力，但SAHA最初是作为一种高效的细胞分化剂开发的，使用HMBA，另一种细胞分化剂，重新激活潜伏的HIV-1作为结构模板。为此，我们在此报告，先前对HIV-1重新激活药物组合的药物筛选显示，FDA批准的一组药物或化合物具有已报告的细胞分化能力，包括放线菌素、阿克拉霉素、阿糖胞苷和阿昔双林，它们使潜伏感染通过低水平激活而重新激活。我们假设，细胞分化药物可以启动潜伏的HIV-1感染，然后通过协同激活剂重新激活，而协同激活剂本身只起到最小的激活作用。因此，该应用的两个目标是：(I)识别协同激活剂，与启动药物一起触发系统范围的重新激活；(Ii)描述不同的药物如何改变细胞转录因子谱和信号转导途径，以启动潜伏的HIV-1感染的重新激活。该应用程序的目标是识别多种药物组合，这些药物组合针对潜在的HIV-1感染，在几个分子控制水平上触发重新激活。这将通过药物筛选或通过合理选择化合物/药物来实现，随着我们对已确定的药物如何改变对潜伏的HIV-1感染事件的细胞控制有越来越详细的了解。

DESCRIPTION (provided by applicant): HIV-1 latency, which allows the virus to persist in the presence of antiretroviral therapy, is likely the major hurdle to overcome towards the development of a curative therapy for HIV-1 infection. While it is clear that the latent HIV-1 reservoir needs to be eradicated, it is unclear how this can be achieved. At the molecular biology level, current HIV-1 research focuses on the idea that molecular control of latent HIV-1 infection must be different from the control of inducible cellular genes and is characterized by the presence of a restrictive chromatin environment at the viral LTR. But how such a restrictive chromatin environment can form when the latent virus is integrated into actively expressed host-genes is unclear. How can paused RNAP II be found at the latent viral LTR, when a complex restrictive chromatin structure supposedly shields the LTR? This focus on a restrictive chromatin environment as the molecular control mechanism for latent HIV-1 infection translates into drug discovery and clinical application, where HDAC inhibitors are viewed as the holy grail of HIV-1 reactivation strategies. However, upon closer review of the literature, HDAC inhibitors, which are clinically pursued as HIV-1 reactivating agents, do not show convincing efficacy, neither in many models of latent infection, nor in ex vivo experiments or clinical studies. The possible exception is SAHA, clinically approved as the HDAC inhibitor vorinostat. SAHA exhibits some HIV-1 reactivating capacity in most experimental systems, including ours, but SAHA was initially developed as a highly potent cell-differentiating agent, using HMBA, another cell differentiating agent that reactivates latent HIV-1 as a structural template. To this end, we here report that a previous drug screen for HIV-1 reactivating drug combinations revealed that a panel of FDA-approved drugs or compounds with reported cell-differentiating capacity including dactinomycin, aclacinomycin, cytarabine and aphidicolin prime latent infection for reactivation by low-level activation. We hypothesize that cell-differentiating drugs can act to prime latent HIV-1 infection for reactivation by synergistic activators, which by themselves only exert a minimal activating effect. Thus the two objectives of the application are (i) to identify synergistic activators that trigger system-wide reactivation in combination with the priming drugs and (ii) to describe how differentiating drugs alter the cellular transcription factor profiles and signal transduction pathways to prime latent HIV-1 infection for reactivation. The goal of the application is to identiy multi-drug combinations that target latent HIV-1 infection at several levels of molecular control t trigger reactivation. This will be achieved through drug screening or by rational selection of compounds/drugs that will be enabled as we gain increasingly detailed insights into how the identified drugs alter the cellular control over the latent HIV-1 infection events.