Discovery of new antiviral methylase, protease, and helicase inhibitors of corona-, flavi-, and alphaviruses

发现冠状病毒、黄病毒和甲病毒的新型抗病毒甲基化酶、蛋白酶和解旋酶抑制剂

基本信息

批准号：
10513923
负责人：
THOMAS TUSCHL
金额：
$ 570.11万
依托单位：
HACKENSACK UNIVERSITY MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-05-16 至 2025-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10513923
关键词：
2019-nCoV Aerosols Alphavirus Animal Model Animals Antiviral Agents Biochemical Biological Assay Biological Availability COVID-19 COVID-19 outbreak Caspase Cell Membrane Permeability Cells Cessation of life Characteristics Chemicals Chikungunya virus China Collection Combined Modality Therapy Complex Consultations Contracts Coronavirus Development Disease Disease Outbreaks Distant Drug Kinetics Enzymes Family Flaviviridae Future Goals Human In Vitro Individual Industry Infection Innate Immune Response Institutes Laboratories Lead Libraries Medicine Metabolic Methyltransferase Middle East Respiratory Syndrome Coronavirus Monitor Mutation Names Oral Patients Peptide Hydrolases Pharmaceutical Chemistry Pharmaceutical Preparations Pharmacology Polyproteins Preparation Program Development Property Protein Biosynthesis RNA Helicase RNA Viruses Recombinant Proteins Recording of previous events Reporting Resources Rice SARS-CoV-2 protease SH2D3A gene SH2D3C gene Series Solubility Specificity Structure Surface Therapeutic Togaviridae Toxic effect Universities Viral Viral Pneumonia Viral Proteins Virus Virus Replication ZIKA Zoonoses assay development base clinical candidate clinical development combat design drug development drug resistant virus falls helicase high throughput screening improved in vivo inhibitor insight lead optimization lead series luminescence member metropolitan mosquito-borne novel operation pandemic disease programs resistance mutation small molecule structural biology virology

项目摘要

Coronavirus Disease 2019 (COVID-19) represents an ongoing pandemic caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) and a current death toll of about 5 million. There are only limited antiviral treatment options for patients who contracted the disease. In March 2020, we initiated collaborative small molecule drug development including the laboratories of Dr. Tuschl (high-throughput screening (HTS) assay development), Dr. Glickman (HTS operations), Dr. Patel (structural biology), Dr. Rice (virology), and the Tri- Institutional Therapeutics Discovery Institute (medicinal chemistry) focusing on the two SARS-CoV-2 proteases, NSP3/PLpro and NSP5/3CLpro, required for viral polyprotein processing, the cap N7-G methyltransferase (MTase) NSP14, required for efficient viral protein synthesis and escape from host cell innate immune response, and the RNA helicase NSP13 important for viral replication. We developed robust, HTS fluorescent-substrate-based assays for monitoring PLpro, 3CLpro, and NSP13 activities as well as a luminescence-based assay for monitoring NSP14 N7-G-MTase activity. By HTS of the diverse library of 430,000 compounds available at Rockefeller University (RU), we identified hits in all assays, validated hits, and pursued the most promising non-covalent hits in medicinal chemistry programs. Our lead compounds for NSP14 and PLpro show IC50 values of 2 nM and 100 nM, respectively, and cell-based inhibition of SARS-CoV-2 replication with EC50 values of 80 nM and 7 µM, without cellular toxicity. Here, we propose further lead optimization to improve inhibitor potency and drug-like properties, as well as apply our HTS expertise to target proteases, MTases, and helicases of other RNA viruses with pandemic potential, including MERS, flavi- and alphaviruses. Our specific aims are: (1) Development of SARS-CoV-2 NSP14 and PLpro clinical candidates. Guided by structural insights from inhibitor-PLpro and NSP14 complexes, we will further optimize inhibitor potency and drug-like characteristics with industry-quality medicinal chemistry support from TDI and this program, and the goal to develop 3-5 advanced leads that are subjected to in vitro and in vivo DMPK/ADME-Tox studies utilizing all scientific cores designated for hit-to-lead and lead optimization (2) Develop antiviral lead series against MTases, proteases and helicases of select members of the Coronaviridae, Flaviviridae and Togaviridae families. We will adapt and develop HTS assays using recombinant proteins and identify covalent and non-covalent inhibitors in the RU drug collection and accompany the medicinal chemistry lead development to provide 3-5 leads per target for comprehensive DMPK/ADME-Tox analysis. (3) Monitor the selectivity of novel small molecule antivirals in biochemical assays (using increasingly distant viral and human recombinant proteins) and cell-based viral propagation, as well as resistance mutation identification. We will express recombinant proteins or exploit assays from other projects and scientific cores to define specificity of viral enzyme and replication inhibition and viral mutational escape.

2019 年冠状病毒病 (COVID-19) 是一种由严重急性呼吸系统疾病引起的持续流行病冠状病毒 2 综合征 (SARS-CoV-2) 目前死亡人数约为 500 万人，抗病毒药物有限。 2020 年 3 月，我们启动了小型协作项目。分子药物开发，包括 Tuschl 博士的实验室（高通量筛选 (HTS) 测定）开发）、Glickman 博士（HTS 操作）、Patel 博士（结构生物学）、Rice 博士（病毒学）和 Tri- 机构治疗发现研究所（药物化学）专注于两种 SARS-CoV-2 蛋白酶， NSP3/PLpro 和 NSP5/3CLpro，病毒多蛋白加工所需，帽 N7-G 甲基转移酶 (MTase) NSP14，是有效的病毒蛋白合成和逃避宿主细胞先天免疫反应所必需的， RNA解旋酶NSP13对于病毒复制很重要。我们开发了强大的基于 HTS 荧光底物的检测方法，用于监测 PLpro、3CLpro 和 NSP13 活性以及通过 HTS 监测 NSP14 N7-G-MTase 活性的基于发光的测定。洛克菲勒大学 (RU) 提供的包含 430,000 种化合物的多样化库，我们在所有检测中都鉴定出了命中结果，验证命中，并追求药物化学项目中最有前途的非共价命中。 NSP14和PLpro的化合物显示IC50值分别为2 nM和100 nM，并且基于细胞的抑制 SARS-CoV-2 复制的 EC50 值为 80 nM 和 7 µM，无细胞毒性。在这里，我们建议进一步优化先导化合物以提高抑制剂的效力和类药特性应用我们的 HTS 专业知识来靶向大流行的其他 RNA 病毒的蛋白酶、MTase 和解旋酶我们的具体目标是：(1) SARS-CoV-2 的开发。 NSP14 和 PLpro 临床候选药物以抑制剂 PLpro 和 NSP14 的结构见解为指导。复合物，我们将通过行业品质的药物进一步优化抑制剂效力和药物特性 TDI 和该计划的化学支持，以及开发 3-5 个先进先导化合物的目标利用指定用于先导化合物和先导化合物的所有科学核心进行体外和体内 DMPK/ADME-Tox 研究优化 (2) 开发针对选定成员的 MTase、蛋白酶和解旋酶的抗病毒先导系列我们将使用冠状病毒科、黄病毒科和披膜病毒科的 HTS 检测方法进行调整和开发。重组蛋白并识别 RU 药物集合中的共价和非共价抑制剂并伴随药物化学先导物开发可为每个靶点提供 3-5 个先导物，用于综合 DMPK/ADME-Tox (3) 监测新型小分子抗病毒药物在生化测定中的选择性（使用）越来越远的病毒和人类重组蛋白）和基于细胞的病毒传播，以及抗性突变鉴定。我们将表达其他项目的重组蛋白或检测结果。和科学核心来定义病毒酶的特异性以及复制抑制和病毒突变逃逸。