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

项目摘要/摘要 登革病毒(DENV)是一种具有很高生物医学意义的病原体，但我们对其缺乏有效的治疗 对策。尽管使用直接作用抗病毒药物(DAA)组合的靶向化疗具有 事实证明，在对抗丙型肝炎病毒感染和艾滋病毒方面非常成功，努力开发类似的药物来对抗 DENV尚未成功。依赖RNA的RNA复制引起的DENV的遗传多样性 缺乏校对功能的聚合酶带来了额外的挑战，使其难以开发 疫苗和抗病毒药物，广谱覆盖一个病毒物种内的所有基因类型，并促进 当DAA作为单一疗法使用时，抗病毒耐药性迅速发展。 最近开发的小分子诱导特定蛋白质降解的方法依赖于嵌合体 分子(“PROTAC”、“DECRONIMID”、“DECHERDER”)具有目标特定的配体，连接到部分 结合E3泛素连接酶(例如，Cereblon，VHL)。小分子结合导致泛素化和 靶标的蛋白酶体降解。这导致了事件驱动而不是占用驱动的药理学 从而有效地将靶标从细胞中移除，并对蛋白质的所有功能进行功能性消融。 由于药理活性不需要持续的、化学计量比的靶标参与，即使是适度的 亲和配体可以是有效的降解剂。此外，这种作用机制可能具有更高的自然屏障。 对常规抑制剂的抗药性，已在癌症生物学领域得到证明。而这些 潜在的优势对抗病毒药物的开发是有吸引力的，但目前还不清楚它们能在多大程度上 以达到显著的抗病毒效果。尤其是强病毒表达和病毒的定位 特化膜上或其附近的过程(及其效应器)可能会限制DENV和其他 病毒对这一药理策略的影响。在这里，我们建议探索我们是否可以成功地部署 针对三种必需的DENV蛋白：CORE、NS4B和NS5的靶向蛋白质降解。就像有 目前还没有批准的抗DENV药物，迫切需要寻找新的药理策略来靶向 这种病毒。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来系统地探索潜在的 降解剂和常规抑制剂在亲和力、效力、选择性、持续时间等方面的差异 对抗性的作用和敏感性。我们还将优化经过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降级情况 具有高天然抵抗力的抗病毒战略，并将一流降解物作为领先的 抗病毒药物的开发。在追求这一目标的过程中，我们还将建立重要的概念证明和 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.