Small molecule inhibitory leads against influenza RdRp

描述(申请人提供)：该项目的总体目标是开发一种安全有效的新的流感病毒感染治疗剂，通过阻断流感聚合酶发挥作用，流感聚合酶是一种独特的流感蛋白，对病毒复制和感染至关重要。该项目建立在通过创新的流感聚合酶分析筛选靶向文库而发现新的活性分子的基础上。最初发现的活性分子被进一步发展成新的类药物分子，这种分子对流感聚合酶的抑制能力大约是后者的100倍。该研究计划提出，通过额外的文库合成和定向药物化学循环，进一步将这些化合物开发成优化的药物先导。然后，这些先导化合物可以转移到葛兰素史克(GSK)手中，葛兰素史克是“生物防御伙伴关系”的产品开发合作伙伴。葛兰素史克将负责这种新的一流流感药物的开发、临床试验和分销。与这项工作相关的一些主要创新是：1)我们最初观察到HIV整合酶(临床验证的靶点)的两个金属结合部位与流感核酸内切酶之间的机械相似性，这导致了我们的新假设，即这两个机械相似部位的药效团应该在靶向文库设计中共享有用的共同特征。2)高通量化学的新的特定设计和相关开发，允许合成针对双金属结合位点药效团的5992个新化合物的文库。3)我们内部开发的IRdRp微型基因组(复制子)分析的扩展，允许对&gt；10,000种化合物进行定向筛选。4)以荧光标记的已知核酸内切酶抑制剂和重组PAN核酸内切酶亚单位为关键试剂，设计并建立了一种全新的体外荧光偏振(FP)结合分析方法。5)我们发现了新的类药物化合物，它们在IRdRp复制子实验中表现出良好的细胞抑制作用，在FP内切酶实验中也表现出结合。6)我们证明，上述FP分析中的结合与已发表的对一组已知的天然流感核酸内切酶抑制剂的酶抑制相关。7)开发了一种新的重组PAN结构，首次给出了缓蚀剂与PAN的共晶结构。本项目的具体目标是：目标1.引导优化新型核酸内切酶、聚合酶和双抑制物，利用迭代文库合成和指导药物化学选择两个或两个以上优化的线索进行开发。目的2.评价铅化合物的体内外保护和治疗作用。目的3.通过非GLP毒性和药代动力学研究对最有希望的口服活性最优的候选药物进行优化，以满足所建立的标准。

DESCRIPTION (provided by applicant): The overall goal of this project is to develop a safe and effective new therapeutic agent for influenza virus infections that works by blocking influenza polymerase, which is a unique influenza protein that is critical for virus replication an infection. This project builds on the discovery of new active molecules found by the screening of targeted libraries with innovative influenza polymerase assays. The active molecules that were initially discovered were further developed into new drug-like molecules that are about 100 times more potent in the inhibition of influenza polymerase. The research plan proposes to further develop these compounds into optimized drug leads through additional cycles of library synthesis and directed medicinal chemistry. These lead compounds can then be transferred to GlaxoSmithKline (GSK), who is the product development partner as part of a "Partnership for Biodefense". GSK will be responsible for development, clinical trials and distribution of the new first-in-class influenza drug. Some of the major innovations that are associated with this work are; 1) Our original observation regarding the mechanistic similarity between the two-metal binding site of HIV integrase (which is a clinically validated target) and influenza endonuclease, this led to our novel hypothesis that the pharmacophore of these two mechanistically similar sites should share useful common features for targeted library design. 2) The novel specific design and associated development of the high throughput chemistry that allowed for the synthesis of a library of 5,992 new compounds that target the two-metal binding site pharmacophore. 3) Our extension of the internally developed IRdRp mini-genome (replicon) assay that allows for the targeted screening of >10,000 compounds. 4) The design and development of a completely novel in vitro fluorescent polarization (FP) binding assay using a fluorescently labeled known inhibitor of endonuclease and a recombinant PAn endonuclease subunit as the key reagents. 5) Our discovery of novel drug-like compounds that show good cell-based inhibition in the IRdRp replicon assay and also binding in the FP endonuclease assay. 6) Our demonstration that binding in the above FP assay correlates with the published enzyme inhibition for a set of known inhibitors of native influenza endonuclease. 7) The development of a new recombinant PAn construct that has given the first co-crystal structure of an inhibitor with PAn. The Specific Aims of this project are: Aim 1. Lead optimization of novel endonuclease, polymerase and dual inhibitors, using iterative library synthesis and directed medicinal chemistry to select two or more optimized leads for development. Aim 2. Evaluation of the protective and therapeutic efficacy of lead compounds in vitro and in vivo. Aim 3. Conduct lead optimization through non-GLP toxicity and pharmacokinetic studies for the most promising orally-active optimized lead candidates that meet the established criteria.