Quantum Free energy calculation software in combination with the docking tools and clustering tools to “zip” the amount of work required to screen the large libraries of the available chemical compounds lets us search for novel inhibitors against multiple targets.
Recently we performed the experimental tests of 50 top-predicted compounds for their inhibitory activity against a recent circulating and vaccine strain, Influenza A/Wisconsin/67/2005. VIRAPUR screened the compounds for ability to inhibit the formation of viral plaques on a monolayer of tissue culture cells in vitro.We used Oseltamivir, which is inhibitory to Influenza A/Wisconsin/67/2005 in this assay as a positive control.
Drug solutions at 10 μM, and 25 μM drug solutions (QP-F26 through QP-F50) were added to duplicate wells of each 6-well dish which had been seeded the previous day with a known concentration of MDCK cells. Drug solutions were incubated on the cells for one hour and then approximately 100 plaque forming units of Influenza A/Wisconsin/67/2005 was added to each well. Virus was allowed to adsorb to the monolayer for a defined time. After virus incubation, the virus inoculum was suctioned off the monolayer and the appropriate dilute drug solution to which had been added 0.4% agarose was carefully overlaid on the appropriate monolayer. Cultures were incubated for less than 48 hours and monitored for plaque size. Cultures were stained to easily visualize plaques between 40-50 hours of virus incubation. We stained metabolically active cells with (MTT) , 3-[4,5- dimethyltiazol-2-yl]-2,5-diphenyltetrazolium bromide. Plaques which consist of infected cells originating from one infectious virus are very metabolically active in the second day of infection.
The best of the compounds were taken to IC50 measurements and exhibited the in-vitro activities in 100nM range, as shown on the graph below (half of the data is shown):
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