High-throughput assays and small-molecule discovery of antiviral candidates targeting influenza hemagglutinin

针对流感血凝素的抗病毒候选药物的高通量测定和小分子发现

• 批准号: 10612773
• 负责人: IAN A WILSON
• 金额: $ 67.85万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-05-01 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10612773
• 关键词: Affinity; Animal Model; Animals; Antibodies; Antiviral Agents; Appearance; Binding; Binding Sites; Biochemical; Biological Assay; Biophysics; Biotechnology; Birds; Cells; Collection; Communities; Complementarity Determining Regions; Complex; Cyclic Peptides; Development; Disease; Drug Targeting; Economic Burden; Epidemic; Exhibits; Fluorescence Polarization; Fluorescence Resonance Energy Transfer; Foundations; Future; Goals; Half-Life; Head; Health; Hemagglutinin; Housing; Human; Hydrogen Bonding; Hydrophobicity; Infection; Influenza; Influenza A Virus, H1N1 Subtype; Influenza A Virus, H2N2 Subtype; Influenza A Virus, H3N2 Subtype; Influenza A Virus, H5N1 Subtype; Influenza A Virus, H7N7 Subtype; Influenza A Virus, H7N9 Subtype; Influenza A Virus, H9N2 Subtype; Influenza A virus; Influenza B Virus; Influenza Hemagglutinin; Journals; Label; Laboratories; Lead; Libraries; Membrane; Membrane Glycoproteins; Mission; Mutation; Nature; Neuraminidase; Peptides; Pharmaceutical Chemistry; Pharmaceutical Preparations; Point Mutation; Predisposition; Property; Proteins; Publishing; Recurrence; Reporter; Resistance; Roentgen Rays; Science; Seasons; Serotyping; Signal Transduction; Site; Specificity; Structure; Surface; Surface Antigens; United States National Institutes of Health; Viral; Viral Physiology; Virus; X-Ray Crystallography; Zoonoses; active method; anti-influenza; combat; combinatorial; cost effective; design; drug discovery; feature detection; fitness; flu; fluorophore; high throughput screening; improved; influenza infection; influenzavirus; inhibitor; innovation; interest; iterative design; mutant; nanomolar; neutralizing antibody; new pandemic; novel; novel therapeutics; pandemic disease; pandemic influenza; pandemic potential; pathogenic virus; prevent; rational design; receptor binding; scaffold; seasonal influenza; small molecule; small molecule libraries; stem; stoichiometry; therapeutic candidate; tool

PROJECT SUMMARY / ABSTRACT Influenza A viruses exhibit extreme diversity as exemplified by the multiple serotypes of the hemagglutinin (HA, H1-H18) and neuraminidase (NA, N1-N11) surface antigens. To date, only 3 of 198 possible combinations of HA and NA in avian and other animal reservoirs have been associated with human pandemics (H1N1, H2N2, H3N2). Recent appearances of H5N1, H6N1, H7N7, H7N9, H9N2, and H10N8 in humans are constant reminders of the potential for devastating new pandemics. Influenza B viruses with its two lineages further increase the health and economic burdens of seasonal influenza. No effective antiviral drugs are currently available for preventing entry of influenza A or B viruses into host cells (scientific premise). However, relatively recent discoveries of broadly neutralizing antibodies to human influenza viruses and concomitant structural studies have identified sites-of-vulnerability on the HA in pandemic, seasonal, and emerging influenza viruses. These HA surface sites include the receptor binding site and membrane-proximal stem housing the fusion machinery, both of which are essential for cellular infection. Common features for recognition of these sites can now be exploited in design of small molecules to ultimately develop broadly applicable influenza antivirals. Here, we will employ this structural information into the optimization and execution of high-throughput assays to identify new small-molecule scaffolds that target the highly conserved and vulnerable stem-binding site. High- throughput screening will be performed in parallel on representative HAs from influenza A group 1 against 600K structurally diverse molecules (SA1). We will also subject group 2 and influenza B HAs to a 300K compound screen (SA2). Validated hit compounds will be prioritized based on affinity and breadth across HAs and top candidates will be rigorously optimized into lead molecules by x-ray structure-based design cycled with medicinal chemistry. Biophysical binding, cellular infectivity and resistance assays (e.g., combinatorial viral libraries of HA mutants) will aid in iterative design, selection, and characterization of potential novel therapeutic candidates with favorable drug-like properties. All of these methods are actively employed in the Wolan and Wilson laboratories. As proof-of-concept for this approach, we identified a molecule with modest affinity to the stem of group 1 HAs with an HT assay of our own design. Its co-crystal structure with HA provided critical information towards design and synthesis of a focused compound library, which we used to produce a stereoselective molecule with nanomolar affinity and antiviral activity. Our overall goal is to identify and improve molecules with broad potency against the stem of groups 1 and 2 as well as flu B HAs. To our knowledge, we are the first to design an assay against group 2 and flu B HAs amenable to HTS (innovation). We anticipate that several classes of stem-targeted compound scaffolds will be identified with nanomolar affinity to HAs with cellular antiviral activity and suitable PK-ADME properties. Future efforts will include animal models of influenza infections to further validate our antivirals with the ultimate goal of combatting future influenza pandemics and seasonal epidemics.

项目摘要 /摘要 流感病毒表现出极端多样性，例如血清谷蛋白的多种血清型 （HA，H1-H18）和神经氨酸酶（Na，N1-N11）表面抗原。迄今为止，只有198个可能的组合中有3个 在鸟类和其他动物水库中的HA和NA与人类大流行有关（H1N1，H2N2， H3N2）。 H5N1，H6N1，H7N7，H7N9，H9N2和H10N8在人类中的最新出现是持续的提醒 有可能破坏新大流行的潜力。流感B病毒及其两个谱系进一步增加了 季节性流感的健康和经济负担。目前没有有效的抗病毒药物可用于 防止流感或B病毒进入宿主细胞（科学前提）。但是，相对较新 对人流感病毒和伴随结构研究的广泛中和抗体的发现 已经确定了大流行，季节性和新兴流感病毒的HA的可变性。这些 HA表面位点包括受体结合位点和膜型茎，它们的融合机械， 两者对于细胞感染都是必不可少的。识别这些网站的常见功能现在可以 在设计小分子的设计中被利用，最终开发出广泛适用的流感抗病毒药。 在这里，我们将使用这些结构信息进行高通量测定的优化和执行 确定针对高度保守且脆弱的茎结合位点的新的小分子支架。高的- 吞吐量筛选将与代表A组1组1集对600K进行 结构多样的分子（SA1）。我们还将对第2组和流感B进行300K的化合物 屏幕（SA2）。经过验证的命中化合物将根据亲和力和宽度的HES和TOP进行优先考虑 候选者将通过用药用循环的基于X射线结构的设计将候选者严格优化为铅分子 化学。生物物理结合，细胞感染性和耐药性测定（例如，HA的组合病毒库 突变体）将有助于迭代设计，选择和表征潜在的新型治疗候选者 有利的药物样特性。所有这些方法都积极地用于Wolan和Wilson实验室。 作为这种方法的概念验证，我们确定了一个与第1组茎相关的分子 使用我们自己的设计的HT分析。其与HA的共结晶结构为设计提供了关键信息 和聚焦化合物库的合成，我们用来产生一个立体选择分子 纳摩尔亲和力和抗病毒活性。我们的总体目标是用广泛的效力识别和改善分子 针对第1组和第2组的茎以及流感B的茎。据我们所知，我们是第一个设计测定的人 针对第2组和流感B具有HTS（创新）。我们预计有几类的词干靶向 复合支架将以纳摩尔亲和力与细胞抗病毒活性和合适 PK-ADME属性。未来的努力将包括流感感染的动物模型，以进一步验证我们 抗病毒药的最终目标是打击未来的流感大流行病和季节性流行病。