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Identification of Small Molecule Inhibitors of Influenza a Virus by Chemical Genetics: (English)

Identification of Small Molecule Inhibitors of Influenza a Virus by Chemical Genetics: (English)

          
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About the Book

This dissertation, "Identification of Small Molecule Inhibitors of Influenza A Virus by Chemical Genetics" by Lai-shan, Lau, 劉麗珊, was obtained from The University of Hong Kong (Pokfulam, Hong Kong) and is being sold pursuant to Creative Commons: Attribution 3.0 Hong Kong License. The content of this dissertation has not been altered in any way. We have altered the formatting in order to facilitate the ease of printing and reading of the dissertation. All rights not granted by the above license are retained by the author. Abstract: Abstract of thesis entitled "Identification of small molecule inhibitors of influenza A viruses by chemical genetics" Submitted by LAU Lai Shan for the degree of Master of Philosophy at The University of Hong Kong in August 2007 With the next influenza pandemic expected to hit the society on catastrophic scale, researchers in infectious diseases and public health are working against the clock trying to discover the next vaccine or antiviral that could save millions of lives. Frequent cases of avian H5N1 virus infecting humans and incidents of Tamiflu resistance in hospitals have alarmed the public who are urged to prepare for the worst. Based on the concept of chemical genetics, we screened 50,240 structurally diverse small molecules in influenza cellular infection models employing Madin-Darby Canine Kidney (MDCK) cells and influenza A/WSN/33[H1N1] using a colorimetric MTT (3-[4,5-dimethylth-iazol-2-yl]-2,5-diphenyltetrazolium bromide) high-throughput screening (HTS) assay in 384-well format. Of the bioactive compounds that exhibited anti-viral activity, 950 were selected. The EC (median 50 effective concentration) of the majority of the 950 compounds determined by plaque reduction assay (PRA) was found to be at low micromolar range. Without further modifications, two selected compounds 5733569 (EC: 0.21 μM 0.03μM) and 50 5847422 (EC: 1.87 μM 0.03 μM), exhibited potent antiviral activity. Compound 505847422 is active solely against the human H1N1 influenza viruses while compound 5733569 inhibits H1N1, H3N2, and H5N1 influenza viruses. In order to elucidate the molecular target of these two compounds, the compounds were used to raise resistant mutant viruses. Resistant mutants of both compounds have at least 50 fold less sensitivity to their corresponding inhibitor. In an analysis of the whole genome of the wild type and mutant viruses, a single substitution mutation was detected in viral nucleoprotein (NP) in both mutants (5733569 mutant: Tyr289His; 5847422 mutant: Ser50Asn). Recombinant virus carrying the corresponding mutation in NP confers the resistance to its inhibitor, this demonstrates the phenotypic resistance of mutant virus against inhibitors 5733569 and 5847422 was solely due to the corresponding mutation. In immuno-staining study, compound 5733569 was found to inhibit the nuclear import of the NP or vRNP while compound 5847422 was detected to perturb the nuclear export of the NP or vRNP. Our data indicate that these two compounds target the nuclear trafficking of influenza viruses and that a viral nucleoprotein is a possible target for the development of novel antiviral agents. We anticipate that application of chemical genetics will help us to probe most, if not all, major pathogenic pathways in viral infections, and the selected compounds can be used as leads in the development of antiviral therapeutics. (391 words) DOI: 10.5353/th_b3963441 Subjects: Influenza A virus Pharmacogenetics Biochemical genetics


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Product Details
  • ISBN-13: 9781374678224
  • Publisher: Open Dissertation Press
  • Publisher Imprint: Open Dissertation Press
  • Height: 279 mm
  • No of Pages: 206
  • Spine Width: 11 mm
  • Width: 216 mm
  • ISBN-10: 1374678228
  • Publisher Date: 27 Jan 2017
  • Binding: Paperback
  • Language: English
  • Series Title: English
  • Weight: 490 gr


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