Development of PROTACs Targeting Papain-like Protease as Broad-Spectrum Anti-Coronavirus Therapeutics

开发针对木瓜蛋白酶的 PROTAC 作为广谱抗冠状病毒治疗药物

• 批准号: 10527571
• 负责人: Shiqing Xu
• 金额: $ 22.18万
• 依托单位: TEXAS A&M UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-26 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10527571
• 关键词: 2019-nCoV; Active Sites; Address; Antiviral Agents; Binding; Binding Sites; Biological Assay; COVID-19; COVID-19 pandemic; COVID-19 therapeutics; COVID-19 vaccine; Cells; Cessation of life; Chemicals; Complex; Coronavirus; Development; Disadvantaged; Disease; Disease Outbreaks; Drug Targeting; Event; Fingers; Future; Genetic Transcription; Goals; Health Promotion; Human; Immunization; In Vitro; Infection; Innate Immune Response; Knowledge; Lead; Length; Ligand Binding; Ligands; Link; Mass Spectrum Analysis; Membrane; Mission; Mutation; Nature; Nonstructural Protein; Papain; Paralysed; Pathway interactions; Peptide Hydrolases; Pharmaceutical Preparations; Play; Property; Protac; Protease Inhibitor; Proteins; Public Health; RNA Viruses; Research; Resistance; Role; SARS-CoV-2 B.1.617.2; SARS-CoV-2 P.1; SARS-CoV-2 protease; Site; System; Technology; Testing; Thumb structure; Ubiquitination; United States; United States National Institutes of Health; Vaccines; Variant; Viral; Viral Proteins; Virus Replication; antiviral drug development; base; combat; coronavirus therapeutics; cytotoxicity; drug development; drug discovery; global health emergency; human coronavirus; improved; inhibitor; insight; multicatalytic endopeptidase complex; new technology; novel coronavirus; pathogen; protein function; protein protein interaction; recruit; small molecule; targeted treatment; technology development; tool; ubiquitin-protein ligase; vaccine development; variants of concern

PROJECT SUMMARY/ABSTRACT The current COVID-19 pandemic caused by SARS-CoV-2 has paralyzed much of the world. Encouragingly, a few COVID-19 vaccines have been developed and approved for human immunization. However, existing COVID-19 vaccines target the highly mutable membrane Spike protein of SARS-CoV-2. New viral strains with critical mutations in Spike have emerged such as alpha, beta, delta, and gamma variants that could make current vaccines less effective. To truly overcome the threat posed by SARS-CoV-2 and its emerging variants of concern, it is paramount to develop antiviral drugs that can combat COVID-19 and also be potentially repurposed to combat novel coronaviruses (CoVs) in the future. To address this urgent need, this proposal aims to interface papain-like protease (PLpro) inhibitors/binders and the emerging proteolysis targeting chimera (PROTAC) technology for the development of broad-spectrum anti-CoV PROTACs. PLpro, one of two essential SARS-CoV- 2 proteases, plays a dual role in promoting viral transcription and replication, and antagonizing host innate immune responses. PLpro is encoded by nonstructural protein 3 (nsp3) which is relatively conserved across various CoVs. Thus, PLpro is an attractive target for the development of broad-spectrum anti-CoV drugs. The proposed small-molecule PROTACs are bifunctional small molecules containing two active ligands connected via a chemical linker. One ligand binds specifically to a viral protein target PLpro while the other ligand selectively engages an E3 ubiquitin ligase. The recruitment of the E3 ligase to PLpro facilitates the formation of a ternary complex, leading to ubiquitination and ultimate degradation of PLpro by the ubiquitination-proteasome pathway. As PROTACs regulate protein function by degrading target proteins instead of inhibiting them, they offer many advantages over traditional occupancy-based inhibitors, including (i) sub-stoichiometric activity, (ii) high barrier to resistance, (iii) improved target selectivity, and (iv) that weak binders (no inhibition required) can become potent degraders. On this basis, the overall goal of this application is to validate the degradation of PLpro as a new strategy for the development of broad-spectrum antiviral therapeutics to combat CoVs. In Aim 1, to chemically optimize non-covalent, reversible covalent, and multiple binding-mode PROTACs by linking VHL E3 ligands and PLpro inhibitors/binders, and to evaluate their degradation potency of PLpro in cell-based assays. In Aim 2, to determine the anti-SARS-CoV-2 activity of developed PROTAC molecules. Potent PROTACs (EC50 < 500 nM) will be further tested for their antiviral activities against other CoVs. In vitro DMPK and cytotoxicity studies of potent PROTACs will be conducted. The successful completion of the proposed study will lead to potent anti-CoV PROTACs against COVID-19 that the mechanism of action is fundamentally different from existing antivirals. The developed PROTAC molecules will also have the potential to be repurposed to contain future coronavirus outbreaks.

项目摘要/摘要 当前由SARS-COV-2引起的COVID-19大流行使世界大部分地区瘫痪了。令人鼓舞的是， 已经开发并批准了几种Covid-19-19疫苗用于人类免疫。但是，存在 COVID-19疫苗靶向SARS-COV-2的高度可变的膜尖峰蛋白。新病毒菌株与 出现了尖峰的临界突变，例如α，beta，delta和伽马变体，它们可能使电流产生 疫苗效率较低。为了真正克服SARS-COV-2及其新兴变体所构成的威胁， 开发可以打击Covid-19的抗病毒药是至关重要的 将来战斗新颖的冠状病毒（COV）。为了满足这一迫切需求，该建议旨在接口 木瓜蛋白酶样蛋白酶（PLPRO）抑制剂/粘合剂和新兴蛋白水解靶向嵌合体（Protac） 开发广谱抗COV Protacs的技术。 PLPRO，两个必需的SAR-COV - 之一 2种蛋白酶，在促进病毒转录和复制方面起双重作用，并与宿主与先天作拮抗 免疫反应。 PLPRO由非结构蛋白3（NSP3）编码，该蛋白3（NSP3）相对保守 各种COV。因此，PLPRO是开发广谱抗COV药物的有吸引力的靶标。这 提出的小分子protac是双功能的小分子，该分子包含两个连接的活性配体 通过化学接头。一种配体与病毒蛋白靶PLPRO专门结合，而另一种配体有选择地结合 参与E3泛素连接酶。 E3连接酶以PLPRO的募集促进了三元的形成 复合物，导致PLPRO的泛素化和最终降解，并通过泛素化途径。 由于Protac通过降解靶蛋白而不是抑制它们来调节蛋白质功能，因此它们提供了许多 比传统基于占用的抑制剂的优点，包括（i）子杂化活动，（ii）高障碍 为了抵抗，（iii）提高了目标选择性，并且（iv）弱粘合剂（无需抑制）可能会变成 有效的降级器。在此基础上，此应用的总体目标是验证PLPRO的降解为 开发广谱抗病毒疗法以对抗COV的新战略。在目标1中 通过连接VHL E3，化学优化非共价，可逆的共价和多种结合模式Protac 配体和PLPRO抑制剂/粘合剂，并评估其在基于细胞的测定中PLPRO的降解效力。 在AIM 2中，确定发达的Protac分子的抗SARS-COV-2活性。有效的Protac（EC50 <500 nm）将进一步测试其针对其他COV的抗病毒活性。体外DMPK和细胞毒性 将进行有效的Protac研究。拟议研究的成功完成将导致 有效的抗COV Protac针对COVID-19，即作用机理与 现有的抗病毒药。开发的Protac分子也将有可能重新使用以包含 未来的冠状病毒爆发。