Identification and characterization of novel drugs that target the Influenza

该项目的总体目标是确定针对流感病毒复制的新疗法。每年流感流行造成的全球健康负担，加上流感病毒高致病性毒株的出现，突出表明迫切需要新的有效治疗方法。目前用于治疗流感的药物（金刚烷胺和神经氨酸酶抑制剂）的主要问题是产生耐药性突变，从而使治疗效果失效。已发表的证据表明，靶向流感病毒RNA依赖性RNA聚合酶（RdRp）是一种合理的抗病毒治疗方法。RdRp负责许多功能，包括5'cap识别，内切酶活性，复制，转录和聚腺苷化。最近，冷冻电镜重构研究确定了支链核糖核蛋白（RNPs）结构作为假定的复制中间物，并提出了一种通过RNP模板上的第二聚合酶活性进行病毒复制的机制。第二种聚合酶活性被认为是聚合酶复合物的功能。显然，RdRp提供了多个功能域，可以作为抗病毒药物治疗的靶点。先前的研究表明，聚合酶复合体PB1亚基保守区域的突变会产生失活的RNA聚合酶。我们假设，特异性靶向聚合酶复合物的化合物可能会减少逃逸突变的频率，或促进不适合复制的逃逸突变。我们最近从以前的HTS筛选中发现了潜在的hit化合物，这些化合物在RdRp瞬态测定中显著抑制流感病毒聚合酶活性。在CPE试验中，这些hit化合物对三种不同的流感病毒株有效。我们建议对这些化合物进行表征，并利用新型小分子文库的高通量筛选（HTS）来鉴定抗聚合酶候选化合物，对其进行化学优化，并确定其在流感小鼠模型中的有效性。

The overall goal of this project is to identify new therapies that target influenza virus replication. The global health burden of annual influenza epidemics coupled with the emergence of highly pathogenic strains of influenza virus has highlighted the urgent need for new effective treatments A primary concern with the current drugs (amantadines and neuraminidase inhibitors) used to treat influenza is the development of resistance mutations that negate therapeutic benefit. Published evidence suggests that targeting the influenza virus RNA dependent RNA polymerase (RdRp) is a rational approach for antiviral therapy. The RdRp is responsible for a number of functions including 5'cap recognition, endonuclease activity, replication, transcription, and polyadenylation. Recently, cryo-EM reconstitution studies identified branchedribonucleoproteins (RNPs) structures as putative replication interhiediates and suggested a mechanism for viral replication by a second polymerase activity on the RNP template . The second polymerase activity is believed to be a function ofthe polymerase complex. Clearly, the RdRp provides multiple functional domains that could be targets for antiviral drug therapy. Previous studies showed that mutations in the conserved regions of PB1 subunit of the polymerase complex produce inactive RNA polymerase. We hypothesize that compounds that specifically target the polymerase complex might reduce the frequency of escape mutations, or promote escape mutants that are unfit for replication. We have recently identified potential hit compounds from previous HTS screens that significantly inhibit the influenza virus polymerase activity in an RdRp transient assay. These hit compounds were effective against three different strains of influenza viruses in CPE assays. We propose to characterize these compounds and use high-throughput screening (HTS) of novel small molecule libraries to identify anti-polymerase candidate compounds, chemically optimize them, and establish their effectiveness in an influenza mouse model. \