Development and validation of antivirals against Flaviviruses

摘要 黄病毒每年导致数亿人感染，一系列临床症状包括 出血热、脑炎、肝功能衰竭和先天性疾病。目前，尽管进行了广泛的筛查 由学术和药物实验室发起的运动，没有针对这些疾病的特定疗法 病毒和现有疫苗的匮乏。我们的目标是使用创新的筛查方法和新的病毒蛋白 目标是开发口服配方的直接作用抗病毒药物的组合，可以治疗现有的和新出现的 黄病毒。我们将开发针对黄病毒三种基本蛋白的抗病毒药物。首先，鉴于核苷 类似物是一类重要的抗病毒药物，我们将开发口服生物利用型泛抗黄病毒核苷 类比。我们手头有两个正在开发的支架，将筛选新的核苷库 脚手架。由于核苷具有广泛的活性，我们将针对额外的rna测试最活跃的核苷。 集中在我们CAMPP中的新出现病毒(如SARS-CoV-2、EBOV、LASV和SFTSV)的病毒 财团。由于黄病毒是小的RNA病毒，编码的蛋白质很少，我们将开发抗病毒药物来对抗 其中两个没有已知酶活性的必需蛋白：NS4A和NS1。最近，临床批准了 针对远亲丙型肝炎病毒NS5B的抗病毒药物可以阻断这个非酶靶点的感染。我们 已经确定ZIKV NS4A的一种抑制剂，一种重要的非酶病毒蛋白，可能是一种抗病毒药物。 作为我们计划的一部分，我们将使用药物化学和SAR来优化和扩大其对抗 其他黄病毒。NS1为内质中细胞质中的复制复合体提供支架。 此外，细胞外还通过以下途径促进血管渗漏和中枢神经系统的侵袭 与内皮细胞结合，拮抗先天免疫。我们将使用生化分析来鉴定药物 它与黄病毒NS1结合，然后对其细胞内和/或细胞外功能的阻断进行筛选。毒品 阻断NS1的功能活性可能会影响病毒的复制和致病步骤。对于那些 针对NS5、NS4A或NS1的抑制剂，显示出最佳的生物利用度和疗效，体内水平 在细胞EC90值的几倍以上，我们将进入黄病毒感染的动物模型。在……里面 此外，我们将确定耐药性的屏障，首先在体外测试抑制剂的组合，最终在 体内增强药效，防止出现耐药性。

SUMMARY Flaviviruses cause hundreds of millions of infections annually and a range of clinical syndromes including hemorrhagic fever, encephalitis, liver failure, and congenital disease. Currently, and despite extensive screening campaigns by academic and pharmaceutical laboratories, there are no specific therapeutics against these viruses and a paucity of available vaccines. Our goal is to use innovative screening assays and novel viral protein targets to develop combinations of orally-formulated direct acting antivirals that can treat existing and emerging flaviviruses. We will develop antivirals against three essential proteins in flaviviruses. First, given that nucleoside analogs are an important class of antivirals we will develop orally bioavailable pan-antiflaviviral nucleoside analogs. We have in hand two scaffolds we are developing and will screen libraries of nucleosides for new scaffolds. As nucleosides can be broadly active, we will test the most active nucleosides against additional RNA viruses focusing on emerging viruses (e.g., SARS-CoV-2, EBOV, LASV and SFTSV) that are in our CAMPP consortium. Since flaviviruses are small RNA viruses and encode few proteins, we will develop antivirals against two of these essential protens with no known enzymatic activity: NS4A and NS1. Recently, clinically approved antiviral against the distantly related hepatitis C virus NS5B blocks infection of this non-enzymatic target. We have identified an inhibitor of ZIKV NS4A, an essential non-enzymatic viral protein, as a possible antiviral drug. As part of our program, we will use medicinal chemistry and SAR to optimize and broaden its activity against additional flaviviruses. NS1 serves a scaffold for replication complexes in the cytoplasm at the endoplasmic reticulum and also functions extracellularly to promote vascular leakage and central nervous system invasion by binding to endothelial cells and antagonizing innate immunity. We will use biochemical assays to identify drugs that bind flavivirus NS1 and then screen them for blockade of its intracellular and/or extracellular functions. Drugs that can block the functional activity of NS1 may impact viral replication and steps in pathogenesis. For those inhibitors against NS5, NS4A or NS1 that show optimal bioavailability and efficacy profiles, with in vivo levels that at several fold above their cellular EC90 values, we will advance into animal models of flavivirus infection. In addition, we will determine the barrier to resistance, test combinations of inhibitors first in vitro and ultimately in vivo to increase potency and prevent the emergence of resistance.