Small molecule-induced degradation of dengue proteins as an antiviral strategy

项目概要/摘要 登革病毒（DENV）是一种具有高度生物医学意义的病原体， 对策尽管使用直接作用抗病毒药物（DAA）组合的靶向化疗 已被证明对丙型肝炎病毒感染和艾滋病毒非常成功，努力开发类似药物， DENV没有成功。依赖RNA复制的登革病毒遗传多样性 缺乏校对功能的聚合酶使其难以开发，从而带来了额外的挑战 疫苗和抗病毒药物具有广谱覆盖一个病毒物种内的所有基因型， 当DAA用作单一疗法时，抗病毒耐药性的快速发展。 最近开发的用于小分子诱导的特异性蛋白质降解的方法依赖于嵌合蛋白质。 分子（“PROTAC”、“去乙酰亚胺”、“降解剂”），其具有与 结合E3泛素连接酶（例如，cereblon，VHL）。小分子结合导致泛素化， 目标的蛋白酶体降解。这导致了事件驱动而不是占领驱动的药理学 从而有效地从细胞中去除靶点并功能性地消除蛋白质的所有功能。 由于药理学活性不需要恒定的、化学计量的靶点参与， 亲和配体可以是有效的降解剂。此外，这种作用机制可以有更高的天然屏障 与传统的抑制剂相比，它更容易产生耐药性，这在癌症生物学领域已经得到证实。虽然这些 潜在的优势对于抗病毒药物的开发是有吸引力的，但目前还不清楚它们可以在多大程度上被开发出来。 以获得显著的抗病毒效果。特别是，强病毒表达和病毒的定位 在专门的膜上或附近的过程（及其效应物）可以限制DENV和其他病毒的易感性。 病毒对这种药理学策略的影响。在这里，我们建议探索我们是否可以成功地部署 针对三种必需DENV蛋白的靶向蛋白降解：核心、NS 4 B和NS 5。因为有 目前没有批准的抗DENV药物，迫切需要找到新的药理学策略来靶向 这个病毒。从已知的抑制剂作为降解剂开发的靶向配体开始，我们将开发和 验证抗病毒降解剂。然后，我们将使用这些工具，系统地探索潜在的点， 降解剂和常规抑制剂在亲和力、效力、选择性、降解持续时间方面的差异 行动和抵抗的敏感性。我们还将优化经验证的抗病毒降解剂，以测试 这种体内抗病毒的方法。总体目标是验证这些目标中的一个或多个的退化， 具有高天然耐药性屏障的抗病毒策略，并将一流的降解剂作为 开发抗病毒药物。为了实现这一目标，我们还将建立重要的概念验证和 为更广泛地开发针对其他病毒病原体的抗病毒降解剂奠定了基础。

PROJECT SUMMARY / ABSTRACT Dengue virus (DENV) is a pathogen of high biomedical significance against which we lack effective countermeasures. Although targeted chemotherapy using combinations of direct-acting antivirals (DAAs) has proven highly successful against hepatitis C virus infection and HIV, efforts to develop analogous drugs against DENV have not been successful. The genetic diversity of DENV due to replication by an RNA-dependent RNA polymerase that lacks proofreading function presents additional challenges by making it difficult to develop vaccines and antivirals with broad-spectrum coverage of all genotypes within one viral species and facilitating the rapid development of antiviral resistance when DAAs are used as monotherapies. Recently developed methods for small molecule-induced degradation of specific proteins rely on chimeric molecules (“PROTACs,” “degronimids,” “degraders”) that have a target-specific ligand linked to a moiety that binds an E3 ubiquitin ligase (e.g., cereblon, VHL). Small molecule-binding leads to ubiquitination and proteasomal degradation of the target. This results in event-driven rather than occupancy-driven pharmacology leading to efficient removal of the target from the cell and functional ablation of all of the protein's functions. Since pharmacological activity does not require constant, stoichiometric engagement of the target, even modest affinity ligands can be effective degraders. In addition, this mechanism of action can have higher natural barriers to resistance than conventional inhibitors, as has been demonstrated in the cancer biology field. While these potential advantages are attractive for antivirals development, it remains unclear the extent to which they can be leveraged to attain significant antiviral effects. In particular, strong viral expression and localization of viral processes (and their effectors) on or near specialized membranes may limit the susceptibility of DENV and other viruses to this pharmacological strategy. Here we propose to explore whether we can successfully deploy targeted protein degradation against three essential DENV proteins: core, NS4B, and NS5. As there are currently no approved anti-DENV drugs, there is an urgent need to find new pharmacological strategies to target this virus. Starting with known inhibitors as targeting ligands for degrader development, we will develop and validate antiviral degraders. We will then use these as tools to systematically explore potential points of differentiation between degraders and conventional inhibitors in terms of affinity, potency, selectivity, duration of action and susceptibility to resistance. We will also optimize validated antiviral degraders to test the efficacy of this antiviral approach in vivo. The overall goal is to validate degradation of one or more of these targets as an antiviral strategy with high natural barrier to resistance and to advance first-in-class degraders as leads for the development of antivirals. In pursuit of this goal, we will also establish important proof of concept and the foundation for more broadly developing antiviral degraders against other viral pathogens.