The Development of Reversible Covalent PROTAC Technology as a New Anti-COVID-19 Strategy

项目摘要/摘要 目前由SARS-CoV-2引起的新冠肺炎大流行是一场全球性的卫生紧急事件。然而，到目前为止， 目前还没有确定有效的靶向药物或疫苗。寻找有效的靶向治疗方案是 最重要的是。SARS-CoV-2是一种包膜的、阳性的RNA病毒。主要蛋白酶(MPRO)，a 半胱氨酸蛋白酶是病毒复制和致病所必需的，是一个吸引人的靶点。 抗SARS-CoV-2的抗病毒药物研究进展传统的抗病毒策略之一是开发高效的 直接与病毒蛋白结合并抑制其功能的亲和配体，如SARS-CoV-2 MPRO(SC2Mpro)。 然而，这些占有率驱动的抑制剂可能会导致几个潜在的问题，如脱靶毒性， 剂量限制毒性和抗药性。因此，迫切需要新的抗病毒策略，以 通过开发替代机制来应对这些挑战，以对抗现有的冠状病毒病原体，如SARS- CoV-2。蛋白水解靶向嵌合体(PROTAC)是一种新兴的靶向蛋白质降解技术 药物发现。PROTAC是事件驱动的双功能小分子，同时与E3接合 泛素连接酶与靶蛋白形成三元复合体，导致泛素化 和目标蛋白的最终降解。与PROTAC相比，PROTAC具有许多潜在的优势 传统的以占有率为基础的抑制剂，包括(1)催化性质，以实现亚化学计量比活性，(2) 增强的靶标选择性；(Iii)高抗药性；和(Iv)取消靶标蛋白的所有功能 以及它的下游蛋白质。在此基础上，这一提议提供了一种创新的反Cov战略：可逆 结合超强可逆共价SC2Mpro抑制剂和SC2Mpro抑制剂优势的共价PROTAC 事件驱动的PROTAC技术。总体目标是验证SC2MPro的降级作为一种新的战略 开发选择性和有效性更高的新冠肺炎药物。目前的建议是建立在 发现几种有效的可逆共价SC2Mpro抑制剂的初步工作(最低IC50；lt；10 NM) 和一种小分子SC2Mpro PROTAC降解剂。在这些令人兴奋的初步研究的鼓舞下，目标是 将通过追求以下目标来实现：(1)开发细胞系统以评估退化 SC2Mpro；(2)针对SC2Mpro的各种高效可逆共价抗冠状病毒PROTAC的开发；(3) SC2Mpro降解能力与抗SARS-CoV-2活性关系的探讨 可逆共价抗冠状病毒PROTACs。成功完成拟议的研究不仅将导致 有效的抗冠状病毒PROTAC具有良好的类药物特性，可潜在地发展到临床前 对新冠肺炎的治疗评价，也将为更广泛地发展抗-AND提供概念验证研究 针对各种冠状病毒的冠状病毒PROTAC。

PROJECT SUMMARY/ABSTRACT The current COVID-19 pandemic caused by SARS-CoV-2 is a global health emergency. However, to date, no effective targeted drug or vaccine has been identified yet. Finding effective targeted treatment options is of paramount importance. SARS-CoV-2 is an enveloped, positive-sensed RNA virus. Main protease (Mpro), a cysteine protease, is essential for viral replication and pathogenesis which represents an attractive target for the development of antiviral drugs against SARS-CoV-2. One of the traditional antiviral strategies is to develop high- affinity ligands that bind directly to viral proteins and inhibit their functions like SARS-CoV-2 Mpro (SC2Mpro). However, these occupancy-driven inhibitors may lead to several potential problems such as off-target toxicity, dose-limiting toxicity, and drug resistance. Thus, there is an urgent need for new antiviral strategies that can address these challenges by exploiting alternative mechanisms to combat existing CoV pathogens like SARS- CoV-2. Proteolytic targeting chimaera (PROTAC) is an emerging technology for targeted protein degradation in drug discovery. PROTACs are event-driven bifunctional small molecules that simultaneously engage an E3 ubiquitin ligase and a target protein to facilitate the formation of a ternary complex, leading to the ubiquitination and ultimate degradation of the target protein. PROTACs have many potential advantages compared to traditional occupancy-based inhibitors, including (i) catalytic nature to allow for sub-stoichiometric activity, (ii) enhanced target selectivity, (iii) high barrier to resistance; and (iv) abrogating all functions of the target protein and its downstream proteins. On this basis, this proposal provides an innovative anti-CoV strategy: reversible covalent PROTACs by combination of the advantages of ultra-potent reversible covalent SC2Mpro inhibitors and event-driven PROTAC technology. The overall goal is to validate degradation of SC2Mpro as a new strategy for developing COVID-19 drugs with improved selectivity and efficacy. The current proposal is built upon the preliminary work on the discovery of several potent reversible covalent SC2Mpro inhibitors (lowest IC50 < 10 nM) and one small-molecule SC2Mpro PROTAC degrader. Encouraged by these exciting preliminary studies, the goal will be achieved by pursuing the following aims: (1) the development of cellular systems to evaluate degradation of SC2Mpro; (2) the development of various potent reversible covalent anti-CoV PROTACs targeting SC2Mpro; (3) the exploration of the relationship between SC2Mpro degradation potencies and anti-SARS-CoV-2 activities of reversible covalent anti-CoV PROTACs. The successful completion of the proposed study will not only lead to potent anti-CoV PROTACs with good drug-like properties that can be potentially advanced to pre-clinical evaluation for treating COVID-19, but also will provide a proof-of-concept study for more broadly developing anti- CoV PROTACs against various coronaviruses.