Novel antivirals targeting the RNase H activity of HIV reverse transcriptase

描述(申请人提供)：高效抗逆转录病毒疗法(HAART)药物主要针对四种HIV酶中的三种：逆转录酶(RT)DNA聚合酶、蛋白酶和整合酶。尽管HAART在抑制HIV感染患者的病毒载量方面非常有效，但长期治疗不可避免地会导致耐药病毒株的出现。因此，开发对新的艾滋病毒靶点起作用的制剂是至关重要的。第四种HIV酶，RT-相关核糖核酸酶H(RNH)就是这样的靶标之一。RNH在逆转录过程中降解病毒RNA基因组，对HIV复制至关重要。这是艾滋病毒唯一一种尚未被抗逆转录病毒药物解决的酶活性。这类药物可能会对目前所有的抗药性病毒株起作用。我们的目标是开发有效的RNH抑制剂(RNHI)，在基于细胞的复制分析中具有纳摩尔效力，为此，我们将使用基于我们在结构生物学、计算生物学、药物化学、酶学、生物化学和病毒学方面的广泛专业知识的多学科方法。我们将通过基于结构的设计过程，包括RNH-RNHI复合体的结构确定和计算分析、药物化学以及新合成的抑制剂的生化和病毒学表征的迭代循环，寻求改善现有的先导和实现抗病毒活性的NM效力。为此，我们提出了三个具体目标：具体目标1.构效关系和化学合成。我们将准备一个经过验证的RNHI筛选命中数据库和我们开发的新型支架。我们将基于数据库分析以及从结晶学和分子对接研究中获得的结构信息，对两类不同的RNHI进行完整的SAR。具体目标2.RT-RNHI相互作用的结晶学和计算分析。基于结构的设计是该应用程序的主要关注点。我们将使用我们实验室已经建立的结晶学工具，这些工具通常会在含有抑制剂的复合体中产生高分辨率的RT和/或RNH结构(分辨率高达1.5？)。这些结构信息将用于指导新抑制剂的设计。具体目标3.重组人免疫缺陷病毒的生化和病毒学特征我们将使用生化和病毒学分析来评估选定的有效筛选命中和将在AIM 1中准备的新的RNHI。这些信息将被整合到迭代的SAR介导的新抑制剂的设计中。我们的多学科方法将导致新的艾滋病毒抑制剂，将有效地对野生型和抗药性病毒株。

DESCRIPTION (provided by applicant): Highly active antiretroviral therapy (HAART) drugs primarily target three of the four HIV enzymes: reverse transcriptase (RT) DNA polymerase, protease, and integrase. Although HAART is very effective in suppressing viral load in HIV-infected patients, prolonged treatment inevitably leads to the emergence of drug-resistant viral strains. Hence, it is essential to develop agents that act on novel HIV targets. The fourth HIV enzyme, RT- associated ribonuclease H (RNH) is one such target. RNH degrades the viral RNA genome during reverse transcription and is essential for HIV replication. It is the only enzymatic activity of HIV that has yet to be addressed by antiretroviral drugs. Such drugs will likely be active against all current drug-resistant viral strains. Our goal is to develop potent RNH inhibitors (RNHIs) with nanomolar efficacy in cell-based replication assays and for this we will use a multidisciplinary approach based on our extensive expertise in structural biology, computational biology, medicinal chemistry, enzymology, biochemistry and virology. We will pursue improvement of current leads and achievement of nM potencies of antiviral activities through a structure-based design process involving iterative cycles of structure determination and computational analysis of RNH-RNHI complexes, medicinal chemistry, and biochemical and virological characterization of newly synthesized inhibitors. To this end we propose three specific aims: Specific Aim 1. Structure-Activity Relationships (SAR) and Chemical Synthesis. We will prepare a database of the validated RNHI screening hits and novel scaffolds that we have developed. We will perform complete SAR for two different classes of RNHIs based on the database analysis and the structural information gained from crystallographic and molecular docking studies in aim 2. Specific Aim 2. Crystallographic and computational analysis of RT-RNHI interaction. Structure-based design is a main focus of this application. We will use crystallographic tools that are already established in our lab and that routinely result in high resolution structures of RT and/or RNH in complex with inhibitors (resolutions up to 1.5 ¿). The structural information will be used to guide the design of new inhibitors. Specific Aim 3. Biochemical and virologic profiling of RNHIs. We will use biochemical and virological assays to assess selected validated screening hits and new RNHIs to be prepared in Aim 1. This information will be integrated in the iterative SAR-mediated design of new inhibitors. Our multidisciplinary approach will lead to new inhibitors of HIV that will be effective against both wild- type and drug-resistant viral strains.