Synergistic Inhibition of Influenza a Virus Replication by a Bacterial Protease Inhibitor and a Plant Preparation

International audienceWe screened the NIH MLSCN 100,000 compound library and discovered a novel scaffold that shows sub-micromolar activity against H5N1and H1N1 influenza viruses in vitro. Cheminformatics and medicinal chemistry analyses were performed of the hit compounds and SAR led to the synthesis of several second-generation compounds with potent nanomolar activity and increased polarity for hit-to-lead optimization. Of the second-generation compounds, several met our activity criteria for identification of lead compounds: an efficacy EC50 value of 10 in secondary assays. We screened several lead compounds against 15 influenza A and B viruses in cell culture. They were active against H1N1 and H5N1 viruses, but not against H3N2 and B viruses. Interestingly, neuraminidase assays reveal that this scaffold did not inhibit viral neuraminidase. Real-time q-RT-PCR results revealed that these chemotypes significantly reduced RNA levels as compared to the no drug influenza infected MDCK cells. This suggests that these compounds target an early event in the viral life cycle in agreement with time-of-addition experiments whereby a significant reduction in virus-specific protein synthesis occurred 6 h post-infection in the presence of compound. Indirect immunofluorescence studies suggest these compounds affect the nuclear export of the N proteins from H1N1 virus. To supplement our in vitro studies, we have developed an HPLC procedure to measure the concentration of lead agents in mouse plasma after IP injection. Preliminary pharmacokinetic results indicate that significant plasma levels (μM) were achieved with five of the eight compounds that have been tested. These compounds were retained in the plasma for over 1 h. Because of the nM potency of these compounds, these results suggest that these five compounds are candidates for evaluation of efficacy in animal models. We are in the process of determining the maximally tolerated dose of these five agents. One or more of these will be selected for initial in vivo evaluation and the data will be presented

A novel proteinaceous protease inhibitor from Streptomyces chromophuscus with antiviral activity

International audienc

A novel proteinaceous protease inhibitor from Streptomyces chromophuscus with antiviral activity

International audienc