Neuraminidase activity provides a practical read-out for a high throughput influenza antiviral screening assay.
Neuraminidase activity provides a practical read-out for a high throughput influenza antiviral screening assay.
复制标题
神经氨酸酶活性为高通量流感抗病毒筛查测定法提供了实用的读出。
DOI：
10.1186/1743-422x-5-109
复制
发表时间：
2008-09-26
期刊：
影响因子：
4.8
通讯作者：
Li, Min
中科院分区：
文献类型：
--
作者：
Eichelberger, Maryna C.;Hassantoufighi, Arash;Wu, Meng;Li, Min
对常用抗病毒药物具有抗性的流感毒株的出现突出了开发靶向病毒基因产物或宿主机制的新化合物的需要，所述病毒基因产物或宿主机制对于有效的病毒复制是必不可少的。鉴定潜在抗病毒化合物的现有测定通常使用靶向特定病毒复制步骤的高通量筛选测定。为了拓宽抗病毒药物的研究，可以进行基于细胞的复制测定，但这些通常是劳动密集型的，并且具有有限的通量。我们对传统的病毒中和试验进行了改造，开发出一种实用的、基于细胞的高通量筛选试验。该检测方法使用病毒神经氨酸酶（NA）作为读数来量化流感复制，从而提供了一种快速且灵敏的检测方法。除了鉴定靶向病毒或宿主因子的抑制剂外，该测定还允许同时评价药物毒性。许多已知的流感抑制剂，包括靶向M2离子通道的金刚烷胺、靶向NA的扎那米韦、靶向IMP脱氢酶的利巴韦林和靶向蛋白激酶A/C的双吲哚基马来酰亚胺，都证明了抗病毒活性。通过比较IC 50与野生型病毒的IC 50来鉴定金刚烷胺耐药株。在使用NA作为复制读数的加速病毒抑制试验中，可以很容易地鉴定出对广泛靶点具有特异性的抗病毒药。该检测适用于高通量筛选，以鉴定潜在的抗病毒药，或可用于鉴定耐药流感病毒株。
The emergence of influenza strains that are resistant to commonly used antivirals has highlighted the need to develop new compounds that target viral gene products or host mechanisms that are essential for effective virus replication. Existing assays to identify potential antiviral compounds often use high throughput screening assays that target specific viral replication steps. To broaden the search for antivirals, cell-based replication assays can be performed, but these are often labor intensive and have limited throughput. We have adapted a traditional virus neutralization assay to develop a practical, cell-based, high throughput screening assay. This assay uses viral neuraminidase (NA) as a read-out to quantify influenza replication, thereby offering an assay that is both rapid and sensitive. In addition to identification of inhibitors that target either viral or host factors, the assay allows simultaneous evaluation of drug toxicity. Antiviral activity was demonstrated for a number of known influenza inhibitors including amantadine that targets the M2 ion channel, zanamivir that targets NA, ribavirin that targets IMP dehydrogenase, and bis-indolyl maleimide that targets protein kinase A/C. Amantadine-resistant strains were identified by comparing IC50 with that of the wild-type virus. Antivirals with specificity for a broad range of targets are easily identified in an accelerated viral inhibition assay that uses NA as a read-out of replication. This assay is suitable for high throughput screening to identify potential antivirals or can be used to identify drug-resistant influenza strains.