Small molecule inhibitors targeting the SARS-CoV-2 pathogenicity factor Nsp1

摘要 新冠肺炎疫情给人类生命和世界经济造成了巨大危害。差不多两年后 其病原体SARS-CoV-2的出现，目前可用的抗病毒药物仍然很少。目前的形势 新病毒变种的出现可能会提供抗病毒治疗药物，从而使病情进一步恶化 而疫苗在未来的效果就不那么好了。这要求制定多样化的抗病毒战略，旨在 以尽可能多的不同病毒途径为目标。在这里，我们第一次探索创造 病毒蛋白Nsp1的抑制剂，用于对抗SARS-CoV-2感染。Nsp1是一种主要的毒力因子， 通过限制宿主基因的表达来抑制抗病毒信号。我们最近确认了九个推定的 通过定量高通量筛选Nsp1的抑制物发挥作用。我们提出了一种深入的结构性和 对已鉴定的化合物进行功能表征，以探索它们开发成强效化合物的能力 抗病毒药物。在目标1中，我们将使用核磁共振波谱来确定药效团和结合部位(S) Nsp1上的小分子抑制剂。目标1的目标是确定最小配体结构特征 对于抑制Nsp1来说是必要的。我们还将测试商业上可用的类似物，以询问哪种化学物质 部分成分可以增加结合和抑制作用。在目标2中，我们将探索Nsp1的哪些功能是目标 小分子抑制。通过结合一系列互补的生化和基于细胞的分析，我们将 询问小分子抑制对核糖体结合、信使核糖核酸降解和信使核糖核酸的影响 翻译。目标2的目标是确定抑制作用的机制，以帮助未来的化合物优化。通过 通过药物靶向Nsp1的选择性功能，我们也可能获得新的生物学洞察力 冠状病毒宿主关闭途径。总体而言，这些研究应该提供对结构和机制的洞察 SARS-CoV-2 Nsp1潜在小分子抑制剂的研究，为今后的化学优化奠定了基础 先导化合物，目标是开发新的有效的抗冠状病毒药物。

SUMMARY The COVID-19 pandemic has caused great harm to human life and the worldwide economy. Almost 2 years after the emergence of its etiological agent SARS-CoV-2, there are still very few available antiviral drugs. The situation is further aggravated by the emergence of new viral variants that might render available antiviral therapeutics and vaccines less effective in the future. This calls for the development of diverse antiviral strategies, aimed at targeting as many different viral pathways as possible. Here we explore for the first time the possibility to create inhibitors of the viral protein Nsp1 to fight SARS-CoV-2 infection. Nsp1 is a major virulence factor that functions by restricting host gene expression to inhibit antiviral signaling. We have recently identified nine putative inhibitors of Nsp1 function by quantitative high throughput screening. We propose an in-depth structural and functional characterization of the identified compounds to explore their capability to be developed into potent antiviral drugs. In aim 1 we will use NMR spectroscopy to identify the pharmacophore and binding site(s) of the small molecule inhibitors on Nsp1. It is the goal of aim 1 to determine the minimal ligand structural features necessary for Nsp1 inhibition. We will also test commercially available analogs to interrogate which chemical moieties can increase binding and inhibition. In aim 2 we will explore which functions of Nsp1 are targeted by small molecule inhibition. By combining an array of complementary biochemical and cell-based assays, we will interrogate the effect of small molecule inhibition on ribosome binding, mRNA degradation, and mRNA translation. It is the goal of aim 2 to identify the mechanism of inhibition to aid future compound optimization. By pharmacologically targeting selective functions of Nsp1, we might also gain new biological insight into the coronaviral host-shutoff pathway. Overall, these studies should provide insight into the structure and mechanism of potential small molecule inhibitors of SARS-CoV-2 Nsp1, laying the foundation for future chemical optimization of lead compounds with the goal to develop new potent anti-coronaviral drugs.