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Exploring the Coronavirus Exoribonuclease as an Antiviral Target

探索冠状病毒外核糖核酸酶作为抗病毒靶点

基本信息

批准号：
10238324
负责人：
SANDRA K WELLER
金额：
$ 46.37万
依托单位：
UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
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：
2021-07-19 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10238324
关键词：
2019-nCoV 5&apos -exoribonuclease Active Sites Acute Address Antiviral Agents Asia Binding Binding Sites Biochemical Biological Biological Assay COVID-19 pandemic Catalytic Domain Cells Cessation of life China Chiroptera Collection Complex Coronavirus Coronavirus Infections Custom Defect Development Disease Outbreaks Drug Synergism Drug Targeting Drug resistance Economics Ensure Enzyme Inhibitor Drugs Enzymes Evaluation Excision Exonuclease Exoribonucleases FDA Emergency Use Authorization Family Fluorescence Future Generations Genetic Transcription Genetic study Genome Guanosine Hepatitis C virus Herpesviridae Human Impairment Lead Libraries Magnesium Mediating Methyltransferase Middle East Respiratory Syndrome Middle East Respiratory Syndrome Coronavirus Morbidity - disease rate Murine hepatitis virus Mutagenesis Mutation Nidovirales Nonstructural Protein Outcome Pathogenicity Permeability Pharmaceutical Preparations Pharmacology Phenotype Positioning Attribute Production Proteins Public Health RNA RNA Viruses RNA chemical synthesis RNA replication RNA-Directed RNA Polymerase Reading Recording of previous events Reporting Resistance Respiratory syncytial virus Ribavirin Ribonucleosides Role SARS coronavirus Saudi Arabia Set protein Severe Acute Respiratory Syndrome Structure Testing Therapeutic Time Toxic effect Vaccines Viral Viral Genome Virus Virus Replication Work Zoonoses analog antiviral nucleoside analog base betacoronavirus biodefense biophysical techniques chemical resource cofactor cytotoxicity drug development efficacy study human coronavirus improved in vivo inhibitor/antagonist lead candidate lead series magnesium ion member metallicity mortality mutant novel novel therapeutics nuclease nucleoside analog pandemic disease pathogen programs remdesivir respiratory screening small molecule small molecule inhibitor synergism viral RNA zoonotic coronavirus

项目摘要

Over the last 20 years, three major zoonotic coronavirus (CoV) infections have emerged all causing acute respiratory illness, leading to significant morbidity and mortality. SARS-CoV-1 emerged in Asia in late 2002 while Middle East Respiratory Syndrome (MERS-CoV) was first reported in Saudi Arabia in 2012. In late 2019, SARS-CoV-2 was reported in China and has now spread globally causing over 673,000 deaths in less than eight months. The currently-raging COVID-19 pandemic presents an urgent need to explore new targets and approaches. Many RNA viruses such as Hepatitis C virus (HCV) and Respiratory Syncytial virus (RSV) can be treated with ribavirin and other broad-spectrum antiviral nucleoside analogues. Ribavirin and other nucleoside analogues are misincorporated into progeny genomes by the virally encoded RNA-dependent RNA-polymerase (RdRp), resulting in lethal mutagenesis. Interestingly, ribavirin has minimal effect against SARS or MERS due to unique aspects of CoV replication. The 30kb CoV genomes are the largest of all RNA viral genomes, more than three times the typical size. All CoVs encode 16 non-structural proteins (nsps) required for the production of progeny RNA. Nsp14 contains an exoribonuclease domain (ExoN), which has been shown to ensure replication fidelity of the large genome and provide resistance to drugs like ribavirin. ExoN, a conserved 3’ to 5’ proofreading exoribonuclease, removes misincorporated ribavirin, rendering it ineffective. Active site mutants of human CoV ExoN result in severe defects in viral RNA synthesis. Thus, ExoN inhibitors are expected to be effective anti- coronavirus agents either as monotherapy or in synergistic combination with nucleoside analogues. This proposal aims to discover first-in-class ExoN inhibitors for further drug development. Our current program is focused on developing broad-spectrum small molecule inhibitors of essential viral exonucleases as antiviral drugs. These exonucleases, like CoV ExoN, possess an acidic active site containing dual magnesium ions that coordinate substrate binding and catalyze bond cleavage. This ExoN key structural motif presents an excellent opportunity to expand our antiviral program to CoVs. We have synthesized hundreds of herpesvirus exonuclease inhibitors tailored to the bi-metallic binding site, which will be evaluated for activity against bacterially expressed ExoN. We propose that inhibitors of ExoN proof-reading activity would exert strong antiviral effects as single agents and would powerfully synergize with ribonucleoside analogs such as ribavirin and remdesivir. Key outcomes of the work would be (1) establishing the druggability of the active site of ExoN for small molecule inhibitors, (2) determining the antiviral activity produced by direct ExoN inhibition, (3) evaluating the potential synergism between ExoN inhibitors and nucleoside analogs and (4) identifying lead candidates for follow on in vivo drug development efforts.

在过去的20年里，出现了三种主要的人畜共患冠状病毒（CoV）感染，呼吸道疾病，导致严重的发病率和死亡率。SARS-CoV-1于2002年底在亚洲出现而中东呼吸综合征（MERS-CoV）于2012年首次在沙特阿拉伯报告。2019年末， SARS-CoV-2在中国报告，目前已在全球蔓延，在不到8年的时间里造成673，000多人死亡个月当前肆虐的COVID-19大流行迫切需要探索新的目标，接近。许多RNA病毒，如丙型肝炎病毒（HCV）和呼吸道合胞病毒（RSV），可以被感染。用利巴韦林和其他广谱抗病毒核苷类似物治疗。利巴韦林和其他核苷类似物被病毒编码的依赖RNA的RNA聚合酶错误地整合到后代基因组中（RdRp），导致致死性诱变。有趣的是，利巴韦林对SARS或MERS的作用最小，冠状病毒复制的独特之处30 kb的CoV基因组是所有RNA病毒基因组中最大的，超过是一般尺寸的三倍所有CoV编码16种非结构蛋白（nsps），这些蛋白是产生子代RNA。Nsp 14含有核糖核酸外切酶结构域（ExoN），已显示其确保复制大基因组的保真度，并提供对利巴韦林等药物的耐药性。ExoN，保守的3'至5'校对核糖核酸外切酶去除错误掺入的利巴韦林，使其无效。人冠状病毒的活性位点突变体 ExoN导致病毒RNA合成的严重缺陷。因此，预期ExoN抑制剂是有效的抗肿瘤药物。冠状病毒药物作为单一疗法或与核苷类似物协同组合。这该提案旨在发现一流的ExoN抑制剂，用于进一步的药物开发。我们目前的项目集中在开发广谱的小分子抑制剂，核酸外切酶作为抗病毒药物。这些核酸外切酶，如CoV ExoN，具有酸性活性位点，双镁离子，协调底物结合并催化键断裂。ExoN的关键结构 motif提供了一个极好的机会，将我们的抗病毒计划扩展到CoV。我们已经合成了数百种的疱疹病毒核酸外切酶抑制剂定制的双金属结合位点，这将被评估的活性针对细菌表达的ExoN。我们认为，ExoN校对活性的抑制剂将发挥强大的作用，作为单一药物的抗病毒作用，并将与核糖核苷类似物如利巴韦林和Remdesivir。这项工作的主要成果将是（1）建立ExoN活性位点的药物可药性对于小分子抑制剂，（2）测定通过直接ExoN抑制产生的抗病毒活性，（3）评估ExoN抑制剂和核苷类似物之间的潜在协同作用，以及（4）鉴定先导化合物用于后续体内药物开发工作的候选物。