Drug target validation of the enterovirus D68 2A protease

肠道病毒 D68 2A 蛋白酶的药物靶点验证

• 批准号: 10543627
• 负责人: Jun Wang
• 金额: $ 54.06万
• 依托单位: RUTGERS, THE STATE UNIV OF N.J.
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-22 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10543627
• 关键词: Drug; target; validation; enterovirus; D68

Enterovirus D68 (EV-D68), a viral pathogen associated with moderate to severe respiratory illness in children, also infects the nervous system in rare cases and causes neurological complications such as polio-like acute flaccid myelitis (AFM). Despite a significant disease burden, no approved antiviral drugs or vaccines are available for EV-D68. Due to its medical importance, EV-D68 is currently listed as a priority pathogen by the National Institute of Allergy and Infectious Diseases. Because the occurrence of the next EV-D68 outbreak cannot be predicted, the need for a potent, broadly acting, and safe EV-D68 inhibitor is urgent. The objectives of this proposal are to explore novel viral proteins as antiviral drug targets and to develop corresponding chemical probes as tools for target validation. This proposal is built upon our recent discovery that the EV-D68 2A protein exerts cysteine protease activity that is specific for the viral polyprotein VP1-2A junction and its protease activity can be inhibited by an FDA-approved oral drug, telaprevir. We further showed that telaprevir had submicromolar to low micromolar potency against several contemporary human EV-D68 strains from clades A and B in different human cell lines, including human neuronal cells. The antiviral potency of telaprevir (IC50) against EV-D68 was similar to that of HCV, indicating clinical relevance of repurposing telaprevir as an EV-D68 antiviral. The mechanism underlying the inhibition of the EV-D68 virus by telaprevir was independently elucidated using a serial viral passage experiment, leading to the selection of an N84T mutation near the active site of EV-D68 2Apro protein that reduced the potency of telaprevir in enzymatic and cellular antiviral assays. To validate the hypothesis that EV-D68 2Apro is essential for viral replication both in vitro and in vivo, our goals are to develop 2Apro inhibitors with favorable pharmacokinetic properties and to use them as chemical probes to validate EV-D68 2Apro as a viable antiviral drug target in cell culture as well as in a neonatal mouse model of EV-D68-induced paralytic disease. Specifically, we propose to design more potent and selective EV- D68 2Apro inhibitors through structure-activity relationship studies of telaprevir. All designed compounds will be systematically tested in in vitro enzymatic and cellular assays and profiled for pharmacokinetic properties. Prioritized lead compounds will be selected to test in EV-D68 mouse models. The expected outcomes of the proposed research are to validate EV-D68 2Apro as a viable antiviral drug target and demonstrate the in vivo antiviral efficacy of EV-D68 2Apro inhibitors.

肠病毒D68 （EV-D68）是一种与儿童中度至重度呼吸道疾病相关的病毒性病原体，在极少数情况下也会感染神经系统，并导致神经系统并发症，如脊髓灰质炎样急性弛缓性脊髓炎（AFM）。尽管有重大的疾病负担，但没有批准的抗病毒药物或疫苗可用于EV-D68。由于其在医学上的重要性，EV-D68目前被美国国家过敏和传染病研究所列为优先病原体。由于无法预测下一次EV-D68暴发的发生，因此迫切需要一种有效、广泛作用和安全的EV-D68抑制剂。本研究的目的是探索新的病毒蛋白作为抗病毒药物靶点，并开发相应的化学探针作为靶点验证的工具。这一建议是基于我们最近的发现，即EV-D68 2A蛋白对病毒多蛋白VP1-2A连接具有特异性的半胱氨酸蛋白酶活性，并且其蛋白酶活性可以被fda批准的口服药物telaprevir抑制。我们进一步发现，在不同的人类细胞系（包括人类神经元细胞）中，telaprevir对来自A枝和B枝的几种当代人类EV-D68株具有亚微摩尔到低微摩尔的效力。telaprevir对EV-D68的抗病毒效力（IC50）与HCV相似，表明将telaprevir重新用作EV-D68抗病毒药物具有临床意义。特拉韦对EV-D68病毒的抑制机制是通过一系列病毒传代实验独立阐明的，导致在EV-D68 2Apro蛋白活性位点附近选择了一个N84T突变，该突变降低了特拉韦在酶和细胞抗病毒试验中的效力。为了验证EV-D68 2Apro在体外和体内对病毒复制至关重要的假设，我们的目标是开发具有良好药代动力学特性的2Apro抑制剂，并将其作为化学探针来验证EV-D68 2Apro在细胞培养和EV-D68诱导的麻痹性疾病的新生小鼠模型中作为可行的抗病毒药物靶点。具体来说，我们建议通过对telaprevir的构效关系研究来设计更有效和选择性的EV- D68 2Apro抑制剂。所有设计的化合物都将在体外酶和细胞分析中进行系统测试，并对其药代动力学特性进行分析。将选择优先的先导化合物在EV-D68小鼠模型中进行测试。本研究的预期结果是验证EV-D68 2Apro是一个可行的抗病毒药物靶点，并证明EV-D68 2Apro抑制剂的体内抗病毒效果。