Antiviral Program for Pandemics (APP) & NCATS: Accelerating Antiviral Development

流行病抗病毒计划 (APP)

• 批准号: 10916053
• 负责人: Matthew Hall
• 金额: $ 3435.8万
• 依托单位: NATIONAL CENTER FOR ADVANCING TRANSLATIONAL SCIENCES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10916053
• 关键词: 2019-nCoV; 3-Dimensional; Acceleration; Address; Air; Alphavirus; Alveolar; Antiviral Agents; Arenavirus; BCAR3 gene; Binding; Biochemical; Biological Assay; Biology; Brain; Bunyavirales; COVID-19 therapeutics; COVID-19 treatment; Cells; Chemicals; Chemistry; Chikungunya virus; Clinical Trials; Collaborations; Collection; Containment; Coronavirus; Data; Dedications; Dengue Virus; Development; Disease; Disease model; Drug Evaluation; Drug Kinetics; Ebola virus; Encephalitis Viruses; Energy Transfer; Enterovirus 68; Exonuclease; Exposure to; Extramural Activities; Family; Family Picornaviridae; Filovirus; Flaviviridae; Flavivirus; Fostering; Goals; Hospitalization; Human; Infection; Investigational Drugs; Investigational New Drug Application; Kunjin virus; Lead; Length; Libraries; Liquid substance; Liver; Luciferases; Lung; Methyltransferase; Modeling; Monitor; Mutation; Nanovirus; National Center for Advancing Translational Sciences; Neurons; Nucleocapsid; Oral; Orthobunyavirus; Papain; Paramyxovirus; Paxlovid; Peptide Hydrolases; Pharmaceutical Chemistry; Physiological; Play; Pneumovirus; Preparation; Property; RNA; RNA-Directed RNA Polymerase; Reader; Recombinants; Reporter; Research Personnel; Role; SARS-CoV-2 B.1.1.529; SARS-CoV-2 inhibitor; SARS-CoV-2 variant; Safety; Serotyping; Small Intestines; Source; Structure; Structure of parenchyma of lung; Structure-Activity Relationship; System; Technology; Testing; Therapeutic; Therapeutic Intervention; Tissue Model; Togaviridae; Toxicology; Triage; United States Food and Drug Administration; United States National Institutes of Health; Validation; Vascular Endothelium; Venezuelan Equine Encephalitis Virus; Venezuelan Equine Encephalomyelitis; Viral; Viral Load result; Viral Physiology; Viral Proteins; Virus; Virus Diseases; Virus Inhibitors; West Nile virus; Work; Zika Virus; anti-viral efficacy; antiviral drug development; assay development; base; chikungunya; clinical development; combat; counterscreen; design; drug development; drug discovery; drug metabolism; efficacy validation; endonuclease; future pandemic; high throughput screening; high-throughput drug screening; human coronavirus; improved; in silico; in vitro Assay; in vivo; inhibitor; interest; lead optimization; member; mutant; nanoluciferase; novel; novel coronavirus; pandemic disease; pandemic potential; pandemic virus; permissiveness; pre-clinical; preclinical development; prevent; programs; respiratory virus; screening; small molecule; small molecule libraries; therapeutic target; variants of concern; vascular bed; viral outbreak; viral pandemic; virology; working group

1) The Target-Based Assays 1 (Martinez) Group has identified viral targets amenable for therapeutic intervention via small molecule modulation, including endonucleases, exonuclease, proteases, methyltransferases, and nucleocapsid-RNA interactions. Candidate compounds resulting from each assay will be screened in live-virus assays by APP colleagues, and in silico approaches are used to support screening efforts. These viral targets include: Inhibitors of Bunyaviral endonucleases: High-throughput Foster resonance energy transfer (FRET)-based biochemical assays for RVFV, LACV, HANV, and PTV endonucleases have been developed and screened against small molecule libraries. Cell-based Split-luciferase cellular thermal shift assay (CETSA) have been developed to further characterize inhibitor-endonuclease engagement. Inhibitors of viral proteases: High-throughput FRET-based biochemical assays for flaviviral NS2B-NS3 (WNV, ZIKV, DENV2, and YWF), Alphaviral NSP2 (CHKV), and Picornaviral 2A and 3C (EN68 and E71) proteases have been developed and screened. Counterscreen assays against host proteases have also been implemented to support hit triage. A complementary cell-base protease assay using a Split Nanoluciferase reporter has been developed for the above proteases to further characterize compound activity in cells. Inhibitors of viral Methyltransferases: A high-throughput screening platform utilizing recombinant Mtases from families including Flaviviral, Henipaviral and Coronaviral Mtases is being developed. The assay uses the MtaseGlo system to monitor SAM to SAH conversion. Assays for Flaviviral NS5 (DENV, WNV) and Coronavirus NSP14/10 and NSP16/10 (SARS-CoV2) have been developed and screened. Inhibitors of viral Exonuclease: a FRET-based assay was developed to identify inhibitors of Coronavirus NSP14/10 exonuclease activity. We have screened NCATS-sourced libraries and to support hit triage, we are developing target engagement assays 2) The Target-Based Assays 2 (Kales) group has developed assays to facilitate the study of mutations in key CoV-2 therapeutic targets and are leveraging those technologies and platforms to target other viral families of pandemic potential, including Alphaviruses. The team has initiated the design and optimization of assays related to the following: Inhibitors of SARS-CoV-2 PLPro Protease: the team has screened 91K compounds to date, and is currently developing a cell-based nanoluciferase assay to assess cellular efficacy of lead compounds and the effects of novel CoV-2 PLPro mutations. Inhibitors of SARS-CoV-2 3CL/MPro Protease: two enzymatic assays have been developed to screen for novel CoV-2 3CL inhibitors, and to assess effects of mutations on current therapeutics, including Paxlovid. The team is studying mutant forms of SARS-CoV-2 3CL to assess activity and efficacy of existing therapeutics. Non-Nucleoside Inhibitors (NNIs) of SARS-CoV-2 RNA-dependent RNA Polymerase: several HTS-amenable biochemical assays and multiple secondary assays have been developed to support screens for inhibitors against recombinant CoV-2 RdRp. Nearly 100K compounds have been screened to date. Inhibitors of Chikungunya virus (CHIKV) nsp3/macrodomain: several activity and binding-based assays towards nsp3/macrodomain of Chikungunya and Venezuelan Equine Encephalitis (VEEV) have been developed. 3) The Advanced Models and Cell-Based Discovery (Lee) group works across a large panel of BSL-2 and BSL-3 viruses in both 2D and 3D assays for antiviral screening and development. Live viruses: More than 25 species of full-length or isolate human viruses + 1 minigenome system have been onboarded across 11 viral families, including Bunyavirales, Arenaviridae JUNV, Peribunyaviridae CEV, LACV, Phenuiviridae RVFV-MP12 and PTV (strains -Balliet and -Adames), Nairoviridae HAZV; Coronaviridae SARS-CoV-2 (previous and current variants of concern), HCoV-OC43/GFP, HCoV-229E, HCoV-NL63; Filoviridae EBOV (minigenome system); Flaviviridae DENV (serotypes 1-4), KUNV, YFV-17D, ZIKV (strains -MR766 and -PRVAB59); Paramyxoviridae PIV-3-GFP, PIV-1-GFP; Picornaviridae EV-D68; Pneumoviridae RSV; Togaviridae CHIKV 181/25/nluc, MAYV/nluc, ONNV-GFP, RRV-GFP, SINV-GFP (neuroinvasive and non-neuroinvasive strains), and VEEV-Tc83-GFP. HTS Assays: the team has 22 live virus 1536w or 384w HTS assays developed and 8 in development across 23 viral species, including Coronaviridae, Phenuiviridae (Bunyavirales), Peribunyaviridae (Bunyavirales), Arenaviridae (Bunyavirales), Flaviviridae, Paramyxoviridae, Picornaviridae and Togaviridae. In addition, the team continues to optimize and support the BSL-3 high-containment lab on NIH main campus containing liquid handlers, an automated confocal plate reader and an integrated screening system to support APP screening pipelines. 3D Live Virus Assays: the team continues to utilize air liquid interface (ALI) lung models including an ALI bronchial model and an ALI alveolar model to investigate antivirals against SARS-CoV-2 as well as for examining differences in SARS-CoV-2 variant infectivity. Additional work includes: o ALI lung tissue mimics to include infectivity screening with a panel of additional respiratory viruses ( o SARS-CoV-2 contemporary omicron variant infectivity differences in lung and small intestinal tissue models o Neuronal spheroid infection and disease models for encephalitis viruses, including RVFV, CHIKV, CEV, JCV, LACV, MAYV, SINV, and VEEV o Liver spheroid model that is permissive to a panel of APP viruses including RVFV, CHIKV, DENV, and YFV o Brain endothelial vascular bed model that recapitulates damage induced by exposure to the viral protein NS1 from the flavivirus JEV 4) The Medicinal Chemistry (Shamim) group is currently optimizing a lead for Papain like-protease, an undrugged target for SARS-CoV-2. Med-chem efforts are directed towards improving enzymatic inhibition, antiviral activity, and pharmacokinetic (PK) properties. A reporter-based luciferase enzymatic assay and cell-based SARS-CoV-2 live virus nano luciferase reporter are being employed to drive the SAR in a structure-based discovery campaign. The medicinal chemistry team is also working towards targeting the cap snatching endonuclease for bunyavirus.

1）基于靶标的测定1（马丁内斯）组已通过小分子调节鉴定了用于治疗干预的病毒靶标，包括核酸内切酶，外核酸酶，蛋白酶，蛋白酶，甲基转移酶和nucleocapsid-rna相互作用。每种测定法引起的候选化合物将在应用程序同事中进行实时病毒测定法中进行筛选，并使用计算机方法来支持筛查工作。这些病毒靶标包括： BUNYAVIRAL核酸内切酶的抑制剂：针对RVFV，LACV，HANV和PTV内核酸酶的高通量促进寄生共振能量转移（FRET）的生化测定法，并筛选了针对小型分子图书馆。已经开发出基于细胞的分裂酸酶细胞热移测定法（CETSA），以进一步表征抑制剂 - 核酸酶的参与。 病毒蛋白酶的抑制剂：基于高通量的FRET生物化学测定NS2B-NS3（WNV，ZIKV，DENV2和YWF），Alphaviral NSP2（CHKV）（CHKV），以及Picornaviral 2A和3C（EN68和E71）的Protease和3C（EN68和E71）的Protease and Spearted and Specioned and Specioned。还针对宿主蛋白酶进行了反屏幕测定，以支持HIT Triage。已经为上述蛋白酶开发了使用分裂纳米酸酯酶报道器的补充细胞碱碱蛋白酶测定，以进一步表征细胞中的复合活性。 病毒甲基转移酶的抑制剂：正在开发来自Flaviviviral，Henipaviral和冠状病毒MTases的重组MTases的高通量筛查平台。该测定使用mtaseglo系统监视SAM转换。已经开发和筛选了黄素NS5（DENV，WNV）和冠状病毒NSP14/10和NSP16/10（SARS-COV2）的测定法。 病毒外切核酸酶的抑制剂：开发了基于FRET的测定方法，以鉴定冠状病毒NSP14/10外切核酸酶活性的抑制剂。我们已经筛选了NCATS的图书馆并支持HIT Triage，我们正在开发目标参与分析 2）基于目标的测定2（KALES）组开发了测定方法，以促进关键COV-2治疗靶标的突变研究，并利用这些技术和平台来针对其他大流行潜力的病毒家族，包括α病毒。该团队启动了与以下测定的设计和优化： SARS-COV-2 PLPRO蛋白酶的抑制剂：该团队迄今已筛选了91K化合物，目前正在开发一种基于细胞的纳米酸酯酶测定法，以评估铅化合物的细胞功效和新型COV-2 PLPRO突变的影响。 SARS-COV-2 3CL/MPRO蛋白酶的抑制剂：已经开发了两种酶试验来筛选新的COV-2 3CL抑制剂，并评估突变对包括Paxlovid在内的当前治疗剂的影响。该团队正在研究SARS-COV-2 3CL的突变形式，以评估现有治疗剂的活动和功效。 SARS-COV-2 RNA依赖性RNA聚合酶的非核苷抑制剂（NNIS）：已经开发了几种HTS不合同的生化测定和多种次级测定，以支持针对重组COVV-2 RDRP的抑制剂的筛查。迄今为止，已经筛选了将近100K的化合物。 Chikungunya病毒（CHIKV）NSP3/MACRODOMAIN的抑制剂：对Chikungunya和委内瑞拉北马脑炎（VEEV）的几种活性和基于结合的测定法。 3）高级模型和基于细胞的发现组（LEE）组在2D和3D测定中的大型BSL-2和BSL-3病毒中工作，用于抗病毒筛查和发育。 实时病毒：超过25种全长或分离的人类病毒 + 1个微型素组系统已在11个病毒家族中入门，包括Bunyavirales，Arenaviridae Junv，Peribunyaviridae Cev，Lacv，pheiviridaeeae rvffv -Mp12和ptv（strains -balliet and -balliet and -AdahameAse），naiririridae; Coronaviridae SARS-COV-2（以前和当前关注的变体），HCOV-OC43/GFP，HCOV-229E，HCOV-NL63； Filoviridae Ebov（微型元组系统）； flaviviridae denv（血清型1-4），kunv，yfv -17d，zikv（strains -mr766和-prvab59）; Paramyxoviridae PIV-3-GFP，PIV-1-GFP; Picornaviridae EV-D68;肺炎病毒RSV; Togaviridae CHIKV 181/25/NLUC，MASEV/NLUC，ONNV-GFP，RRV-GFP，SINV-GFP（NEUROINOVASIVE和非神经活动菌株）和VEEV-TC83-GFP。 HTS Assays: the team has 22 live virus 1536w or 384w HTS assays developed and 8 in development across 23 viral species, including Coronaviridae, Phenuiviridae (Bunyavirales), Peribunyaviridae (Bunyavirales), Arenaviridae (Bunyavirales), Flaviviridae, Paramyxoviridae, Picornaviridae and togaviridae。此外，该团队继续在NIH主校园，装有液体处理程序，自动共聚焦板读取器和集成的筛选系统上优化和支持BSL-3高控制实验室，以支持应用程序筛选管道。 3D实时病毒测定法：该团队继续利用包括ALI支气管模型和ALI肺泡模型在内的空气液体界面（ALI）肺模型，以研究针对SARS-COV-2的抗病毒药，以检查SARS-COV-2变体感染性的差异。其他工作包括： o ali肺组织模仿，包括通过一系列其他呼吸道病毒进行感染性筛查（ o SARS-COV-2当代OMICROR的变异感染性差异在肺和小肠组织模型中 O脑炎病毒的神经元球体感染和疾病模型，包括RVFV，CHIKV，CEV，JCV，LACV，MASEV，SINV和VEEV o肝球体模型，该模型允许使用APP病毒，包括RVFV，CHIKV，DENV和YFV o大脑内皮血管床模型，该模型概括了暴露于Flavivirus JEV的病毒蛋白NS1引起的损害 4）药物化学（Shamim）组目前正在优化蛋白乳木瓜类似蛋白酶的铅，蛋白酶蛋白酶是SARS-COV-2的未扣靶。 Med-Chem努力旨在改善酶促抑制作用，抗病毒活性和药代动力学（PK）特性。基于记者的荧光素酶酶试验和基于细胞的SARS-COV-2 Live Virus Nano Luciferase Reporter正在使用基于结构的发现活动来推动SAR。药物化学团队还致力于针对Bunyavirus的帽盖扣核酸内切酶。