Receptor-directed small-molecule inhibitors of New World hemorrhagic fever mammarenavirus entry

项目总结 哺乳动物病毒在世界各地的啮齿动物种群中是地方性的，人畜共患传播可导致严重的 危及生命的出血热。在美洲，包括Junín和Machupo在内的五种哺乳动物病毒 病毒(分别为JUNV和MACV)可引起病毒性出血热。在没有FDA许可的抗病毒药物的情况下 无论是治疗方法还是疫苗，这些病毒都构成重大的公共卫生问题，并威胁到国家安全。这个 致病性新世界乳癌病毒(NWMs)利用人转铁蛋白受体1(HTfR1)进入 进入人类细胞。病毒包膜糖蛋白的GP1亚基与hTfR1的顶端结构域结合，a 不参与与主要hTfR1配体结合的区域。GP1和hTfR1之间的这种相互作用 因此，它是广泛抑制NWM感染的潜在治疗靶点。基于结构 在MACV包膜糖蛋白GP1亚单位与hTfR1的复合体中，我们发现了一个新的可用药部位 HTfR1的顶端结构域。一项基于对接的虚拟筛选活动确定了28个匹配项 随后确定了在细胞培养中抗JUNV感染的抗病毒活性。在此基础上 评估，我们选择了两个化学上不同的分子，它们对JUNV表现出很强的活性，用于进一步 对价。我们假设靶向hTfR1的顶端结构域袋与病毒GP1相互作用 将广泛抑制所有已知致病NWM的感染，从而保护表达hTfR1的小鼠免受 与JUNV感染相关的致命性疾病。为了探索这一假设，我们将追求以下具体内容 目标。目的1.鉴定针对可药物hTfR1顶端结构域的有效和广谱活性化合物 与NWM GP1交互的站点。我们初步研究中确定的两种不同的化学支架将 作为设计类似物的基础，以定义结构-活性关系(SARS)，对 广泛的抗病毒活性。我们将实施一项迭代活动，包括分子建模、医学 显示包膜的天然JUNV和伪型病毒的化学和体外抗病毒试验 NWMS的糖蛋白。我们的战略将包括对类药物特性进行优化。目标2.评估 优化的候选铅在hTfR1小鼠体内的药代动力学(PK)和疗效。两个优化 来自每个化学支架的化合物将根据它们的体外效力选择在动物身上进行评估， 选择性、抑制广度和类药物特性。我们将评估口服生物利用度，确定 最大耐受剂量，并建立小鼠体内候选铅的PK曲线。这些研究的结果 将在后续疗效中指导有关剂量水平、治疗频率和持续时间的决定 HTfR1小鼠致死性JUNV感染模型的研究。除了改善患者的生存结果外， JUNV挑战hTfR1小鼠，我们将测量治疗对病毒载量、组织病理学和 疾病的严重程度和持续时间。我们的长期目标是开发一种新的宿主导向的小分子疗法 一种可以用来治疗NWM出血热的药物。

PROJECT SUMMARY Mammarenaviruses are endemic in rodent populations worldwide and zoonotic transmission can lead to severe life-threatening hemorrhagic fever. In the Americas, five mammarenavirus species, including Junín and Machupo viruses (JUNV and MACV, respectively), cause viral hemorrhagic fever. In the absence of FDA-licensed antiviral therapies or vaccines, these viruses pose a significant public health concern and threaten national security. The pathogenic New World mammarenaviruses (NWMs) utilize the human transferrin receptor 1 (hTfR1) for entry into human cells. The GP1 subunit of the virus envelope glycoprotein binds to the apical domain of hTfR1, a region that is not involved in the binding to the major hTfR1 ligands. This interaction between GP1 and hTfR1 therefore represents a potential therapeutic target for broad inhibition of NWM infection. Based on the structure of the MACV envelope glycoprotein GP1 subunit in complex with hTfR1, we identified a novel druggable site in the apical domain of hTfR1. A docking-based virtual screening campaign identified 28 hits that were subsequently characterized to determine antiviral activity against JUNV infection in cell culture. Based on this assessment, we selected two chemically distinct molecules displaying strong activity against JUNV for further consideration. We hypothesize that targeting the apical domain pocket of hTfR1 that interacts with viral GP1 will broadly inhibit infection by all known pathogenic NWMs and thereby protect mice expressing hTfR1 from lethal disease associated with JUNV infection. To explore this hypothesis, we will pursue the following specific aims. Aim 1. Identify potent and broadly active compounds targeting the druggable hTfR1 apical domain site that interacts with NWM GP1. The two distinct chemical scaffolds identified in our preliminary studies will serve as the foundation for the design of analogs to define structure-activity relationships (SARs) important for broad antiviral activity. We will implement an iterative campaign involving molecular modeling, medicinal chemistry and in vitro antiviral testing against native JUNV and pseudotyped viruses displaying the envelope glycoprotein of NWMs. Our strategy will include optimization for drug-like properties. Aim 2. Evaluate pharmacokinetics (PK) and efficacy of optimized lead candidates in hTfR1 mice. Two optimized compounds from each chemical scaffold will be chosen for evaluation in animals based on their in vitro potency, selectivity, breadth of inhibition and drug-like properties. We will assess oral bioavailability, determine the maximum tolerated dose and establish PK profiles for the lead candidates in mice. Results from these studies will guide decisions regarding dosing levels and the frequency and duration of treatment in subsequent efficacy studies in a validated hTfR1 mouse model of lethal JUNV infection. In addition to improved survival outcome in JUNV-challenged hTfR1 mice, we will measure the impact of treatments on viral loads, histopathology, and disease severity and duration. Our long-term goal is to develop a novel host-directed small-molecule therapeutic agent that could be deployed to treat NWM hemorrhagic fevers.