Novel antiviral activities of tumor necrosis factor-alpha

描述（由申请人提供）：肿瘤坏死因子-a （TNF1）具有有效的直接抗病毒活性，但由于其全身毒性作用，尚未被用作抗病毒治疗。我们观察到，用低水平TNF1处理的原代人肝细胞和肝癌细胞在体外抵抗丙型肝炎病毒（HCV）感染。这种作用依赖于TNF1与其受体的相互作用，并且在缺乏可检测到的IFN1或IFN2基因表达时可见。Sindbis病毒是一种感染Huh-7肝癌细胞的甲病毒，对TNF1预处理不敏感。然而，TNF1与IFN2协同作用可阻断Sindbis病毒感染。我们建议使用最先进的技术来鉴定tnf - 1单独或与IFN2联合发挥抗病毒作用的关键介质。我们提出三个具体目标。在第一个目标中，我们将使用药物抑制剂以及TNF1信号转导的主要负调节因子，通过TNF1与其受体结合激活的主要途径来测试信号转导的要求。在第二个目标中，我们将使用新的病毒学和细胞生物学工具来确定在TNF1治疗后抑制HCV感染的阶段。在第三个目标中，我们将使用微阵列和下一代测序方法来定义TNF1对转录组的影响，确定可能介导TNF1处理细胞中抗病毒状态发展的候选基因，并通过敲低和过表达研究来测试这些基因的重要性。本研究的长期目标是确定介导TNF1抗病毒活性的特定细胞途径，并确定是否有可能将这些活性从TNF1的全身毒性作用中分离出来。

DESCRIPTION (provided by applicant): Tumor necrosis factor-a (TNF1) has potent, direct antiviral activity but has not been exploited as an antiviral therapy because of its systemic toxic effects. We have observed that primary human liver cells and hepatoma cells treated with low levels of TNF1 are resistant to infection with the hepatitis C virus (HCV) in vitro. This effect is dependent on TNF1 interaction with its receptor and is seen in the absence of detectable IFN1 or IFN2 gene expression. Sindbis virus, an alphavirus, infects Huh-7 hepatoma cells and is insensitive to TNF1 pretreatment. However, TNF1 synergizes with IFN2 to block Sindbis virus infection. We propose to use state- of-the-art techniques to identify key mediators of TNF1's antiviral effects alone and in combination with IFN2. We propose three Specific Aims. In the first Aim, we will test the requirements for signal transduction through the major pathways activated by TNF1 binding to its receptor, using pharmacologic inhibitors as well as dominant negative regulators of TNF1 signal transduction. In the second Aim, we will use novel virological and cell biological tools to define the stages in HCV infection that are inhibited following TNF1 treatment. In the third Aim, we will use microarray and next-generation sequencing approaches to define the effects of TNF1 on the transcriptome, identify candidate genes that may mediate the development of an antiviral state in TNF1- treated cells, and test the importance of these genes by knockdown and overexpression studies. The long-term goals of this research are to identify specific cellular pathways that mediate TNF1's antiviral activity, and to determine whether it is possible to isolate these activities from TNF1's systemic toxic effects. PUBLIC HEALTH RELEVANCE: Hepatitis C virus infection remains a major public health problem and can cause liver cancer and cirrhosis, and is the leading indication for liver transplantation in the United States and Europe. We have found that tumor necrosis factor-1 (TNF1) can make certain cells resistant to infection with hepatitis C virus as well as Sindbis virus, an alphavirus. However, TNF1 is highly toxic and is not suitable as a treatment for viral diseases. We propose to use a variety of techniques to determine how TNF1 mediates its antiviral activity with the hope of isolating the antiviral effects of TNF1 from its toxic effects.