Small-molecule inhibitors of flavivirus entry

登革热病毒和其他黄病毒，包括西尼罗河病毒、蜱传脑炎病毒和黄热病病毒，是广泛传播的蚊子或蜱传人类病原体。世界上约40%的人口生活在登革热传播风险很大的地区，每年可能有多达1亿人感染登革热，经历登革热和可能致命的严重登革热。开发一种对所有四种登革热血清型都有效的疫苗一直很困难，因为感染四种血清型中的一种不会导致对其他三种中的任何一种产生保护性免疫，并可能导致患严重的、危及生命的疾病的风险增加，特别是在儿童中。防治登革热的抗病毒药物具有降低登革热相关疾病严重程度和减少传播的潜力，因此受到关注。该提案将通过开发机制特征良好的登革病毒进入小分子抑制剂，并利用这些抑制剂在体外和体内验证登革热E蛋白作为抗病毒靶点，从而有助于开发登革热抗病毒药物。我们之前已经确定了三种结构不同的先导化合物系列和抑制剂，它们对登革热病毒的进入表现出一系列的活性。在这项工作中，我们将结合x射线晶体学、药物化学、直接蛋白质-小分子亲和测量、单病毒粒子融合和活细胞成像测定以及耐药性研究来优化我们化合物的药理活性，并阐明与有效的泛血清型登革热抑制相对应的生化和结构作用机制。这些努力的关键是定量分析，类似于酶活性测量，用于研究病毒包膜蛋白的生化功能，并使我们能够将小分子抑制剂的生化机制与细胞进入途径中它们阻断感染的步骤联系起来。优化的登革进入抑制剂将在登革感染和发病机制的小鼠模型中进行测试。总的来说，我们的努力旨在生产可以作为登革热药物发现工作的临床前候选化合物。更广泛地说，这项工作为开发其他包膜病毒的小分子抑制剂作为潜在抗病毒药物的合理方法建立了模板

Dengue virus and other flaviviruses including West Nile virus, tick-borne encephalitis virus, and Yellow Fever virus, are widespread, mosquito- or tick-borne human pathogens. About 40% of the world's population lives in areas with substantial risk of dengue transmission, and as many as 100 million people may be infected annually, experiencing dengue fever and potentially lethal severe dengue. Development of a vaccine effective against all four dengue serotypes has been difficult because infection with one ofthe four serotypes does not lead to protective immunity against any of the other three and may lead to enhanced risk of severe, life-threatening illness, especially in children. Antivirals against dengue are of interest due to their potential to reduce the severity of dengue-associated disease as well as to reduce transmission. This proposal will contribute to efforts to develop dengue antivirals by developing mechanistically well-characterized small molecule inhibitors of dengue entry and using these to validate the dengue E protein as an antiviral target in vitro and in vivo. We have previously identified three structurally distinct lead compound series and inhibitors that exhibit a spectrum of activities against dengue virus entry. In the proposed work, we will use a combination of x-ray crystallography, medicinal chemistry, direct protein-small molecule affinity measurements, single-virion fusion and live-cell imaging assays, and resistance studies to optimize the pharmacological activity of our compounds and to elucidate the biochemical and structural mechanisms of action that correspond to potent, pan-serotype inhibition of dengue. Key to these efforts are quantitative assays, analogous to enzymatic activity measurements, for studying the biochemical functions of the viral envelope protein and that allow us to link the biochemical mechanism of small-molecule inhibitors with the step(s) in the cell-entry pathway at which they block infection. Optimized dengue entry inhibitors will be tested in a murine model of dengue infection and pathogenesis. Collectively, our efforts are aimed at producing compounds that can be advanced as preclinical candidates for dengue drug discovery efforts. More broadly, this work establishes the template for a rational approach to the development of small molecule inhibitors of other enveloped viruses as potential antivirals