Influenza virus small RNAs

描述（由申请人提供）：流感病毒每年通常导致超过30，000名美国人死亡，并有可能导致大流行，可能导致数百万人死亡并造成严重的经济破坏。虽然抗病毒药物存在，但这些药物很容易产生耐药性病毒。更好地了解流感病毒生物学有可能提出新的治疗策略，病毒将难以绕过。 我们最近意外发现流感病毒在感染细胞中表达高水平的约20 nt长的小非编码RNA，这促使我们提出了这项拨款申请。这些RNA不是随机的基因组片段，而是几乎完全来源于病毒基因组RNA（vRNA）片段的5'端。我们假设这些RNA通过破坏细胞先天性抗病毒免疫应答和/或通过在病毒生命周期后期调节从mRNA到vRNA和互补RNA转录的转变在病毒生命周期中发挥关键作用。我们打算充分表征这些小病毒RNA的起源、时间表达和序列同一性，然后严格测试通过引入反义锁核酸（LNA）和/或寡核苷酸阻断这些小RNA是否会抑制病毒生命周期中的一个或多个关键步骤。相反，我们还将通过检测合成RNA分子来测试在病毒生命周期早期过表达这些小RNA的影响。重要的是，一种针对另一种呼吸道病毒RSV的吸入性反义药物目前正在临床试验中，并且有效抑制流感病毒复制的寡核苷酸也可能通过吸入来递送。此外，由于在所有流感病毒vRNA的5'端发现的13-nt序列是不变的，因此靶向该区域的有效反义药物可能不会选择抗性突变。总之，我们目前的初步数据表明，第一次，流感病毒在感染细胞中表达高水平的特定群体的小病毒RNA。我们寻求资金来进一步表征这些小病毒RNA的身份和功能，最重要的是，分析它们作为抗病毒药物靶点的潜力。 公共卫生相关性：我们最近发现流感病毒在感染细胞中表达非常高水平的小RNA分子。我们试图确定这些RNA在病毒生命周期中的作用，并评估其作为新型抗病毒药物靶点的潜力。

DESCRIPTION (provided by applicant): Influenza virus routinely kills over 30,000 Americans per year and has the potential to cause a pandemic that could kill millions and cause severe economic disruption. Although anti-viral drugs exist, these give rise to resistant viruses fairly readily. A better understanding of influenza virus biology has the potential to suggest novel treatment strategies that the virus would find difficult to circumvent. This grant application is prompted by our recent, unexpected discovery that influenza virus expresses high levels of small, ~20 nt long non-coding RNAs in infected cells. These RNAs are not random genome fragments, but instead derive almost entirely from the 5' end of the viral genomic RNA (vRNA) segments. We hypothesize that these RNAs play a key role in the viral life cycle by disrupting cellular innate antiviral immune responses and/or by regulating the transition from mRNA to vRNA and complementary RNA transcription late in the viral life cycle. We intend to fully characterize the origin, temporal expression and sequence identity of these small viral RNAs and to then rigorously test whether blocking these small RNAs, by introducing antisense locked nucleic acid (LNA) and/or antagomir oligonucleotides, will inhibit one or more key steps in the viral life cycle. Conversely, we will also test the effect of overexpressing these small RNAs early in the viral life cycle by transfecting synthetic RNA molecules. Importantly, an inhaled antisense drug that targets another respiratory virus, RSV, is currently in clinical trials, and it is likely that an oligonucleotide that inhibits influenza virus replication effectively could also be delivered by inhalation. Moreover, because the 13-nt sequence found at the 5' end of all influenza virus vRNAs is invariant, it is likely that an effective antisense drug targeted to this region would not select for resistance mutations. In conclusion, we present initial data demonstrating, for the first time, that influenza virus expresses a high level of a specific population of small viral RNAs in infected cells. We seek funds to further characterize the identity and functions of these small viral RNAs and, most importantly, to analyze their potential as targets for antiviral drugs. PUBLIC HEALTH RELEVANCE: We have recently discovered that influenza virus expresses very high levels of small RNA molecules in infected cells. We seek to define the role of these RNAs in the viral life cycle and to evaluate their potential as novel antiviral drug targets.