Development of Broad-Spectrum Antiviral Therapeutics by Destabilizing the Main Protease of Coronaviruses

摘要 目前，全球范围内爆发了2019年冠状病毒病(新冠肺炎)，这是由 严重急性呼吸综合征冠状病毒2(SARS-CoV-2)感染。我们迫切需要有效的 针对SARS-CoV-2和相关冠状病毒的抗病毒治疗，以避免未来可能爆发的疫情。这 一项提案寻求试点资金，以支持药物化学家和 病毒学家开发能够破坏3-类糜蛋白酶(3CLpro)稳定的新型小分子， SARS-CoV-2和许多相关冠状病毒复制所必需的主要病毒蛋白酶。这个 所提出的小分子能够通过宿主细胞的泛素来催化3CLPro的降解。 蛋白酶体系统，它例行地移除受损或未折叠的蛋白质。 所提出的降解剂是两个配体之间带有短链的双功能分子。一个配基结合 对E3泛素连接酶具有高选择性，而另一配体同时与病毒蛋白结合 目标。当结合事件发生时，病毒蛋白与E3泛素连接酶密切接触 复合体，在蛋白酶体中被多泛素化降解。然后，降解器被释放到 继续其对病毒蛋白降解的催化活性。降级器只需要瞬时绑定 来诱导目标病毒蛋白的泛素化，这比传统的小分子 分子抑制剂。 我们选择3CLPro作为选择性降解的主要靶点，是因为它在病毒复制和 选择性抑制剂的可用性，它将作为与3CLPro结合的配体。虽然抑制药物是 3CLPro的存在，降解剂的催化性能将使它们变得更加有效。通过破坏稳定和 破坏病毒蛋白质，而不是化学计量结合到病毒蛋白质，降解者也应该采取行动 比抑制剂还快。3CLPro在包括SARS冠状病毒、MERS冠状病毒和 SARS-CoV-2。所提出的小分子降解剂可以有效地对抗广泛的 冠状病毒谱，包括对抗病毒抑制剂具有抗药性的病毒株。在目标1中，我们将 利用我们的最新研究成果连接E3连接酶和3CLPro配体制备双功能小分子降解剂 制定了两阶段战略。在目标2中，我们将评估3CLPro的降解和抗病毒活性 基于细胞的分析中的降解剂。 PI和Co-Investigator研究小组在双功能小分子降解剂方面经验丰富 分别是发展和抗病毒研究。拟议的研究将产生重要的概念验证 针对SARS-CoV-2和相关冠状病毒的新型抗病毒疗法的开发数据。

Abstract At the moment, there is a worldwide outbreak of coronavirus disease 2019 (COVID-19) that is caused by the infection from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We urgently need effective anti-viral therapeutics against SARS-CoV-2 and related coronaviruses to avoid potential future outbreaks. This proposal seeks pilot funding to support the interdisciplinary collaboration between medicinal chemists and virologists to develop novel small molecules that can destabilize 3-chymotrypsin like protease (3CLpro), the main viral protease that is essential for the replication of SARS-CoV-2 and many related coronaviruses. The proposed small molecules are able to catalyze the degradation of 3CLpro through the host cell’s ubiquitin- proteasome-system, which routinely removes damaged or unfolded proteins. The proposed degraders are bifunctional molecules with a short linker between two ligands. One ligand binds with high selectivity to an E3 ubiquitin ligase while the other ligand simultaneously engages the viral protein target. As the binding event occurs, the viral protein is brought in close contact with the E3 ubiquitin ligase complex and is poly-ubiquitinated for degradation in the proteasome. The degrader is then released to continue its catalytic activity for the degradation of the viral protein. The degrader only needs to bind transiently to the target viral protein to induce its ubiquitination, which offers many advantages over traditional small molecule inhibitors. We selected 3CLpro as the primary target for selective degradation based on its key role in viral replication and the availability of selective inhibitors, which will serve as the ligand that binds to 3CLPro. Although inhibitors for 3CLpro exist, degraders’ catalytic properties will render them much more potent. By destabilizing and destroying the viral protein instead of stoichiometrically binding to the viral protein, degraders should also act faster than inhibitors. 3CLpro is conserved among many coronaviruses including SARS-CoV, MERS-CoV, and SARS-CoV-2. The proposed small molecule degraders can be potent antiviral therapeutics against a broad spectrum of coronaviruses, including viral strains that are resistant to antiviral inhibitors. In aim 1, we will prepare bifunctional small molecule degraders by linking ligands of E3 ligase and 3CLPro using our recently developed two-stage strategy. In aim 2, we will evaluate the degradation and anti-viral activities of 3CLPro degraders in cell-based assays. The research groups of PI and Co-Investigator are highly experienced in bifunctional small molecule degrader development and anti-viral research, respectively. The proposed study will produce important proof-of-concept data for the development of novel antiviral therapeutics against SARS-CoV-2 and related coronaviruses.