Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.680807
Title: Development of a novel cell-based screening platform to identify inhibitors of viral interferon antagonists from clinically important viruses
Author: Vasou, Andri
ISNI:       0000 0004 5917 1904
Awarding Body: University of St Andrews
Current Institution: University of St Andrews
Date of Award: 2016
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Abstract:
All viruses encode for at least one viral interferon (IFN) antagonist, which is used to subvert the cellular IFN response, a powerful antiviral innate immune response. Numerous in vitro and in vivo studies have demonstrated that IFN antagonism is crucial for virus survival, suggesting that viral IFN antagonists could represent promising therapeutic targets. This study focuses on Respiratory Syncytial Virus (RSV), an important human pathogen for which there is no vaccine or virus-specific antiviral drug. RSV encodes two IFN antagonists NS1 and NS2, which play a critical role in RSV replication and pathogenicity. We developed a high-throughput screening (HTS) assay to target NS2 via our A549.pr(ISRE)GFP-RSV/NS2 cell-line, which contains a GFP gene under the control of an IFN-stimulated response element (ISRE) to monitor IFN- signalling pathway. NS2 inhibits the IFN-signalling pathway and hence GFP expression in the A549.pr(ISRE)GFP-RSV/NS2 cell-line by mediating STAT2 degradation. Using a HTS approach, we screened 16,000 compounds to identify small molecules that inhibit NS2 function and therefore relinquish the NS2 imposed block to IFN-signalling, leading to restoration of GFP expression. A total of twenty-eight hits were identified; elimination of false positives left eight hits, four of which (AV-14, -16, -18, -19) are the most promising. These four hit compounds have EC₅₀ values in the single μM range and three of them (AV-14, -16, -18) represent a chemically related series with an indole structure. We demonstrated that the hit compounds specifically inhibit the STAT2 degradation function of NS2, not the function of NS1 or unrelated viral IFN antagonists. At the current time, compounds do not restrict RSV replication in vitro, hence hit optimization is required to improve their potency. Nonetheless, these compounds could be used as chemical tools to determine the unknown mechanism by which NS2 mediates STAT2 degradation and tackle fundamental questions about RSV biology.
Supervisor: Adamson, Catherine S. ; Randall, R. E. Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.680807  DOI: Not available
Keywords: Respiratory Syncytial Virus (RSV) ; RSV antivirals ; Viral interferon (IFN) antagonists ; RSV IFN antagonists ; RSV NS1 ; RSV NS2 ; RSV NS2 inhibitors ; STAT2 degradation ; IFN-signalling pathway ; Cell-based high-throughput screening (HTS) assay ; GFP reporter assay ; QR187.5V28 ; Microbiological assay ; Interferon--Antagonists ; Viral proteins ; Respiratory syncytial virus
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