Development of direct-acting flavivirus inhibitors

摘要 开发有效和广谱的直接作用抗病毒药物（DAA）是第一线防御的关键 对抗由黄病毒引起的疾病（例如，登革热病毒，黄热病病毒，西尼罗河病毒，日本 脑炎病毒、寨卡病毒和蜱传脑炎病毒）。为了促进全球健康， 这一建议的前提是联合收割机的专业知识和尖端技术的学术和 制药部门为药物鉴定和商业化建立一个强有力的环境。的 针对黄病毒的广谱DAA的设想的开发过程分为三个阶段（目的 1 - 3）。第一个目标需要鉴定和验证保守的黄病毒靶标和命中物 （由Discovery Core B和Enzymology Core C驱动。已鉴定的依赖RNA的RNA 聚合酶（RdRp）命中将作为MedChem Core进一步优化的化学起点 D.此外，与Discovery Core B合作，我们的目标是鉴定新的结构保守的 以下关键病毒复制酶的结合口袋：非结构蛋白5（NS5）RdRp、NS3 解旋酶和NS2B-NS3蛋白酶。靶向这些位点的化合物将使用虚拟对接进行探索， 基于靶向小分子和片段的高通量筛选和DNA编码的化学文库 (DEL)卡位将在酶测定中验证靶位点。我们将确定关键氨基酸的作用 通过蛋白质工程，酸在酶活性、病毒复制和适应性中的作用。至少有两到四个人 然后，系列将在目标2（命中领先）中进行优化和测试。将对化合物进行表征， 在生物化学和基于细胞的病毒复制测定中的活性，有利于体外药物吸收， 分布、代谢、排泄和毒性（ADME-TOX）、作用机制和抗性。我们 异质黄病毒组将评估不同细胞类型中的广谱抗病毒活性水平。 虽然我们的目标是开发广谱抗黄病毒药物，但几种黄病毒引起显著的人类感染。 因此，高度选择性的抑制剂也将在适当的情况下被追求。铅化合物将 与项目1、2、3和5共享，以评估其他病毒家族的广度。至少两个引线 在目标2中选择的化合物将在目标3中测试体内功效。在一次概念验证之后， 在体内研究中，我们预期递送一种对至少一种黄病毒有活性的黄病毒抑制剂，或至多递送一种对至少一种黄病毒有活性的黄病毒抑制剂。 针对本提案中描述的所有黄病毒，准备启动新药临床试验申请（IND）/ 临床试验授权（CTA）-启动研究，随后如果获得批准，则进入临床开发 并开始第一阶段研究。

ABSTRACT Development of potent and broad-spectrum direct acting antivirals (DAAs) is key as a first-line defense against diseases caused by flaviviruses (e.g., dengue virus, yellow fever virus, West Nile virus, Japanese encephalitis virus, Zika virus and tick-borne encephalitis virus). To promote global health, the underlying premise of this proposal is to combine expertise and cutting-edge technology of academic and pharmaceutical sectors to establish a robust environment for drug identification and commercialization. The envisioned development process of broad-spectrum DAAs against flaviviruses is divided in three stages (aim 1–3). The first aim entails the identification and validation of conserved flavivirus targets and hits (driven by Discovery Core B and the Enzymology Core C. Already identified RNA-dependent RNA polymerase (RdRp) hits will serve as the chemical starting point for further optimization by MedChem Core D. In addition, in collaboration with the Discovery Core B, we aim to identify novel structurally conserved binding pockets of the following key viral replication enzymes: nonstructural protein 5 (NS5) RdRp, NS3 helicase, and NS2B-NS3 protease. Compounds targeting those sites will be explored using virtual docking, targeted small-molecule and fragment-based high- throughput screens and DNA-encoded chemical library (DEL) screens. Target sites will be validated in enzymatic assays. We will determine the role of key amino acids in enzyme activity, viral replication and fitness through protein engineering. At least two to four hit series will then be optimized and tested in aim 2 (Hit to Lead). Compounds will be characterized for activity in biochemical and cell-based viral replication assays, for favorable in vitro drug absorption, distribution, metabolism, excretion and toxicity (ADME-TOX), mechanism of action and resistance. Our heterogeneous flavivirus panel will assess the level of broad-spectrum antiviral activity in diverse cell types. While we aim to develop broad-spectrum anti-flavivirus drugs, several flaviviruses cause significant human disease globally, so highly selective inhibitors will also be pursued as appropriate. Lead compounds will be shared with Projects 1, 2, 3, and 5 to evaluate breadth across other viral families. At least two lead compounds selected in aim 2 will be tested in aim 3 for in vivo efficacy. After proof of concept in an in vivo study, we anticipate to deliver one Flavivirus inhibitor active against at least one flavivirus, or at best against all flaviviruses described in this proposal, ready to start Investigational New Drug Application (IND)/ Clinical Trial Authorization (CTA)-enabling studies, and subsequently if approved, enter clinical development and start Phase1 studies.