Discovery of Bunyaviral Endonuclease Inhibitors for Antiviral Therapy

用于抗病毒治疗的布尼亚病毒核酸内切酶抑制剂的发现

• 批准号: 10481430
• 负责人: Gaya K. Amarasinghe
• 金额: $ 29.65万
• 依托单位: MICROBIOTIX, INC
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-12 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10481430
• 关键词: Acids; Address; Affinity; African; Americas; Animals; Antiviral Agents; Antiviral Therapy; Binding; Biochemical; Biological Assay; Bunyaviridae; Case Fatality Rates; Category A pathogen; Cell Culture Techniques; Cell Line; Cells; Chemicals; Collaborations; Country; Culicidae; Development; Disease Outbreaks; Drug Kinetics; Drug Targeting; Epidemic; Europe; Exhibits; FDA approved; Family; Fluorescence Resonance Energy Transfer; Genus Phlebovirus; Goals; Human; In Vitro; Infection; Influenza; Insect Vectors; Isotope Labeling; Knowledge; Life Cycle Stages; Livestock; Mammalian Cell; Measurement; Measures; Medical; Modeling; Mutation; N-terminal; National Institute of Allergy and Infectious Disease; Orthobunyavirus; Pathogenicity; Patients; Pharmaceutical Preparations; Phase; Property; Proteins; Publishing; RNA Polymerase Inhibitor; RNA Viruses; RNA-Directed RNA Polymerase; Rift Valley fever virus; Risk; Running; Sampling; Series; Serum; Specificity; Structure; Structure-Activity Relationship; System; Therapeutic; Therapeutic Agents; Toxic effect; Vaccines; Validation; Vesicular stomatitis Indiana virus; Viral; Viral Genome; Viral Hemorrhagic Fevers; Virus; Virus Diseases; Virus Replication; base; counterscreen; cytotoxicity; drug discovery; economic impact; endonuclease; epizootic; experience; global health; high throughput screening; human disease; in vivo; influenzavirus; inhibitor; innovation; lead series; mosquito-borne; novel; nucleoside analog; pathogenic virus; prophylactic; small molecule; small molecule inhibitor; small molecule libraries; therapeutic vaccine

Abstract Segmented negative-sense, single-stranded RNA viruses (sNSVs), which include bunyaviruses, are causative agents of human diseases. Rift Valley Fever Virus (RVFV), a bunyavirus, causes hemorrhagic fever in humans with a case fatality rate of patients developing hemorrhagic fever reaching approximately 50% and has been classified by the NIAID as a Category A Priority Pathogen. RVFV is mosquito-borne, but is also capable of using a wide range of insect vectors with potential to spread to Europe and the Americas. RVFV has the potential to cause significant global health and economic impact. Unfortunately, there are no FDA-approved drugs or vaccines for the treatment of the RVFV infection. Therefore, there is an urgent medical need for more potent therapeutics tailored for RVFV. The overall goal of this project is to identify and develop small molecule prophylactics and/or therapeutics for RVFV infections and preferably also for infections of other highly related bunyaviruses. The strategy is to address the unmet medical need by identifying small molecule inhibitors targeting the enzymatic activity of the essential RVFV endonuclease, which exhibits significant structural similarity to other sNSV endonucleases. The approach is to leverage the team’s experience with bunyaviral endonucleases and a homogeneous FRET-based biochemical assay to identify small molecules that inhibit the enzymatic activity of bunyaviral endonucleases. In Preliminary Studies, we developed a FRET-endonuclease activity (FRET-EA) assay for RVFV endonuclease and applied the assay in a low/medium-throughput format with Z’-factors ≥0.9. The FRET assay confirmed the inhibition of an FDA approved antiviral to treat Influenza virus (IAV), Baloxavir acid (BXA), against IAV endonuclease. In Phase I, for Aim 1, the FRET-EA assay will be optimized for high-throughput screening. Biochemical and cellular secondary assays, including FRET, thermal shift, and cell-based infectious assays, will be optimized to further evaluate confirmed hits. In Aim 2, the FRET- EA HTS will be applied to diverse chemical libraries of ≥250,000 small molecules for the identification and confirmation of small molecules that inhibit the enzymatic activity of RVFV endonuclease. In Aim 3, hits identified in Aim 2 will be validated in secondary assays for binding to endonuclease, anti-sNSV spectrum, and potency against infectious RVFV. They will also be prioritized based on their drug-likeness and their ADME properties. In Phase II, we will perform structural based mechanism studies and further chemically optimize priority inhibitors for potency, selectivity, in vitro and in vivo pharmacokinetics properties and evaluate them in animal infection models.

抽象的 分段负义单链 RNA 病毒 (sNSV)（包括布尼亚病毒）是致病原因 人类疾病的媒介。裂谷热病毒 (RVFV) 是一种布尼亚病毒，可引起人类出血热 出血热患者病死率高达50%左右 被 NIAID 列为 A 类优先病原体。 RVFV 是由蚊子传播的，但也能够利用 多种昆虫媒介有可能传播到欧洲和美洲。 RVFV 有潜力 造成重大的全球健康和经济影响。不幸的是，没有 FDA 批准的药物或 用于治疗 RVFV 感染的疫苗。因此，医学界迫切需要更有效的药物 针对 RVFV 量身定制的治疗方法。该项目的总体目标是识别和开发小分子 用于RVFV感染并且优选地还用于其他高度相关的感染的预防剂和/或治疗剂 布尼亚病毒。该策略是通过识别小分子抑制剂来解决未满足的医疗需求 靶向必需 RVFV 核酸内切酶的酶活性，该酶表现出显着的结构 与其他 sNSV 核酸内切酶相似。该方法是利用团队在布尼亚病毒方面的经验 核酸内切酶和基于均相 FRET 的生化测定来鉴定抑制 布尼亚病毒核酸内切酶的酶活性。在初步研究中，我们开发了一种 FRET 核酸内切酶 RVFV 核酸内切酶活性 (FRET-EA) 测定，并以低/中通量形式应用该测定 Z’因子≥0.9。 FRET 测定证实了 FDA 批准的抗病毒药物治疗流感的抑制作用 病毒（IAV），巴洛沙韦酸（BXA），抗 IAV 核酸内切酶。在第一阶段，对于目标 1，FRET-EA 检测将是 针对高通量筛选进行了优化。生化和细胞二次检测，包括 FRET、热分析 转变和基于细胞的感染测定将被优化，以进一步评估已确认的命中。在目标 2 中，FRET- EA HTS 将应用于 ≥250,000 个小分子的多种化学库，用于识别和 确认抑制 RVFV 核酸内切酶酶活性的小分子。在目标 3 中，已确定命中 目标 2 中的目标将在核酸内切酶结合、抗 sNSV 谱和效力的二次测定中得到验证 对抗传染性RVFV。还将根据它们的药物相似性和 ADME 特性对它们进行优先排序。 在第二阶段，我们将进行基于结构的机制研究，并进一步对优先抑制剂进行化学优化 效力、选择性、体外和体内药代动力学特性并在动物感染中进行评估 模型。