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

基本信息

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
项目类别：
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

项目摘要

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.

项目概要/摘要甲型流感病毒表现出极大的多样性，血凝素的多种血清型就是例证（HA、H1-H18）和神经氨酸酶（NA、N1-N11）表面抗原。迄今为止，198 种可能的组合中只有 3 种禽类和其他动物宿主中的 HA 和 NA 与人类流行病（H1N1、H2N2、 H3N2）。最近在人类中出现的 H5N1、H6N1、H7N7、H7N9、H9N2 和 H10N8 不断提醒人们造成毁灭性新流行病的可能性。具有两个谱系的乙型流感病毒进一步增加了季节性流感的健康和经济负担。目前尚无有效的抗病毒药物防止甲型或乙型流感病毒进入宿主细胞（科学前提）。然而，相对最近的人类流感病毒广泛中和抗体的发现及伴随的结构研究已经确定了 HA 在大流行性、季节性和新出现的流感病毒中的脆弱点。这些 HA 表面位点包括受体结合位点和容纳融合机制的近膜茎，两者对于细胞感染都是必需的。现在可以使用用于识别这些站点的通用特征用于小分子设计，最终开发出广泛适用的流感抗病毒药物。在这里，我们将利用这些结构信息来优化和执行高通量测定识别针对高度保守且脆弱的茎结合位点的新小分子支架。高的- 将针对 600K 对甲型流感 1 组的代表性 HA 并行进行通量筛选结构多样的分子（SA1）。我们还将对 2 组和乙型流感 HA 进行 300K 化合物的处理屏幕（SA2）。已验证的命中化合物将根据 HA 和顶级之间的亲和力和广度进行优先排序候选药物将通过基于 X 射线结构的设计与药物循环严格优化为先导分子化学。生物物理结合、细胞感染性和耐药性测定（例如 HA 的组合病毒文库）突变体）将有助于潜在新型治疗候选药物的迭代设计、选择和表征良好的药物样特性。所有这些方法都在 Wolan 和 Wilson 实验室中得到积极应用。作为该方法的概念验证，我们鉴定了一种对 1 组 HA 的茎具有适度亲和力的分子使用我们自己设计的 HT 检测。其与 HA 的共晶结构为设计提供了关键信息以及重点化合物库的合成，我们用它来生产立体选择性分子纳摩尔亲和力和抗病毒活性。我们的总体目标是识别和改进具有广泛效力的分子对抗 1 组和 2 组以及 B 型流感 HA 的茎。据我们所知，我们是第一个设计检测方法的人针对 2 组和 B 型流感 HA，适合 HTS（创新）。我们预计几类针对干细胞的复合支架将被鉴定为具有细胞抗病毒活性和合适的 HA 的纳摩尔亲和力 PK-ADME 特性。未来的努力将包括流感感染的动物模型，以进一步验证我们的研究抗病毒药物的最终目标是对抗未来的流感大流行和季节性流行病。