We assessed the
in vitro
antiviral activity of remdesivir and its parent nucleoside GS-441524, molnupiravir and its parent nucleoside EIDD-1931 and the viral protease inhibitor nirmatrelvir against the ancestral SARS-CoV2 strain and the five variants of concern including Omicron. VeroE6-GFP cells were pre-treated overnight with serial dilutions of the compounds before infection. The GFP signal was determined by high-content imaging on day 4 post-infection. All molecules have equipotent antiviral activity against the ancestral virus and the VOCs Alpha, Beta, Gamma, Delta and Omicron. These findings are in line with the observation that the target proteins of these antivirals (respectively the viral RNA dependent RNA polymerase and the viral main protease Mpro) are highly conserved.
Global health is threatened by emerging viral infections, which largely lack effective vaccines or therapies. Targeting host pathways that are exploited by multiple viruses could offer broad-spectrum solutions. We previously reported that AAK1 and GAK, kinase regulators of the host adaptor proteins AP1 and AP2, are essential for hepatitis C virus (HCV) infection, but the underlying mechanism and relevance to other viruses or in vivo infections remained unknown. Here, we have discovered that AP1 and AP2 cotraffic with HCV particles in live cells. Moreover, we found that multiple viruses, including dengue and Ebola, exploit AAK1 and GAK during entry and infectious virus production. In cultured cells, treatment with sunitinib and erlotinib, approved anticancer drugs that inhibit AAK1 or GAK activity, or with more selective compounds inhibited intracellular trafficking of HCV and multiple unrelated RNA viruses with a high barrier to resistance. In murine models of dengue and Ebola infection, sunitinib/erlotinib combination protected against morbidity and mortality. We validated sunitinib- and erlotinib-mediated inhibition of AAK1 and GAK activity as an important mechanism of antiviral action. Additionally, we revealed potential roles for additional kinase targets. These findings advance our understanding of virus-host interactions and establish a proof of principle for a repurposed, host-targeted approach to combat emerging viruses.
Drugs targeting SARS-CoV-2 could have saved millions of lives during the COVID-19
pandemic, and it is now crucial to develop inhibitors of coronavirus replication in
preparation for future outbreaks. We explored two virtual screening strategies to find
inhibitors of the SARS-CoV-2 main protease in ultralarge chemical libraries. First,
structure-based docking was used to screen a diverse library of 235 million virtual
compounds against the active site. One hundred top-ranked compounds were tested in
binding and enzymatic assays. Second, a fragment discovered by crystallographic
screening was optimized guided by docking of millions of elaborated molecules and
experimental testing of 93 compounds. Three inhibitors were identified in the first
library screen, and five of the selected fragment elaborations showed inhibitory
effects. Crystal structures of target–inhibitor complexes confirmed docking
predictions and guided hit-to-lead optimization, resulting in a noncovalent main
protease inhibitor with nanomolar affinity, a promising in vitro pharmacokinetic
profile, and broad-spectrum antiviral effect in infected cells.
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