The SARS-CoV-2 replication and transcription complex (RTC) comprising nonstructural protein (nsp) 2-16 plays crucial roles in viral replication, reducing the efficacy of broad-spectrum nucleoside analog drugs such as remdesivir and in evading innate immune responses. Most studies target a specific viral component of the RTC such as the main protease or the RNA-dependent RNA polymerase. In contrast, our strategy is to target multiple conserved domains of the RTC to prevent SARS-CoV-2 genome replication and to create a high barrier to viral resistance or evasion of antiviral drugs. We show that clinically-safe Zn-ejector drugs, disulfiram/ebselen, can target conserved Zn2+-sites in SARS-CoV-2 nsp13 and nsp14 and inhibit nsp13 ATPase and nsp14 exoribonuclease activities. As the SARS-CoV-2 nsp14 domain targeted by disulfiram/ebselen is involved in RNA fidelity control, our strategy allows coupling of the Zn-ejector drug with a broad-spectrum nucleoside analog that would otherwise be excised by the nsp14 proofreading domain. As proof-of-concept, we show that disulfiram, when combined with remdesivir, can synergistically inhibit SARS-CoV-2 replication in Vero E6 cells. We present a mechanism of action and the advantages of our multi-targeting strategy, which can be applied to any type of coronavirus with conserved Zn2+-sites.

中文翻译：

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使用双硫仑和瑞德昔韦协同抑制SARS-CoV-2复制

包含非结构蛋白（nsp）2-16的SARS-CoV-2复制和转录复合体（RTC）在病毒复制，降低广谱核苷类似药物（如瑞德昔韦）的功效以及逃避先天免疫应答中起着至关重要的作用。大多数研究都针对RTC的特定病毒成分，例如主要蛋白酶或RNA依赖性RNA聚合酶。相比之下，我们的策略是针对RTC的多个保守域，以防止SARS-CoV-2基因组复制，并为病毒抗药性或抗病毒药物逃避创造高障碍。我们表明，临床安全的锌喷射药物，双硫仑/依布硒仑，可以靶向SARS-CoV-2 nsp13和nsp14中保守的Zn2 +位点，并抑制nsp13 ATPase和nsp14核糖核酸外切酶活性。由于双硫仑/依布硒仑靶向的SARS-CoV-2 nsp14结构域参与RNA保真度控制，因此我们的策略允许将Zn喷射药物与广谱核苷类似物偶联，否则将被nsp14校对结构域切除。作为概念验证，我们证明了双硫仑与瑞姆昔韦合用可以协同抑制Vero E6细胞中的SARS-CoV-2复制。我们提出了一种作用机理和我们多靶点策略的优势，该策略可以应用于具有保守Zn2 +位点的任何类型的冠状病毒。可以协同抑制SARS-CoV-2在Vero E6细胞中的复制。我们提出了一种作用机理和我们多靶点策略的优势，该策略可以应用于具有保守Zn2 +位点的任何类型的冠状病毒。可以协同抑制SARS-CoV-2在Vero E6细胞中的复制。我们提出了一种作用机理和我们多靶点策略的优势，该策略可以应用于具有保守Zn2 +位点的任何类型的冠状病毒。