Most flaviviruses are arthropod-borne viruses, transmitted by either ticks or mosquitoes, and cause morbidity and mortality worldwide. They are endemic in many countries and have recently emerged in new regions, such as the Zika virus (ZIKV) in South-and Central America, the West Nile virus (WNV) in North America, and the Yellow fever virus (YFV) in Brazil and many African countries, highlighting the need for preparedness. Currently, there are no antiviral drugs available to treat flavivirus infections. We have previously discovered a broad-spectrum antiviral compound, benzavir-2, with potent antiviral activity against both DNA- and RNA-viruses. Our purpose was to investigate the inhibitory activity of benzavir-2 against flaviviruses. We used a ZIKV ZsGreen-expressing vector, two lineages of wild-type ZIKV, and other medically important flaviviruses. Benzavir-2 inhibited ZIKV derived reporter gene expression with an EC₅₀ value of 0.8 ± 0.1 µM. Furthermore, ZIKV plaque formation, progeny virus production, and viral RNA expression were strongly inhibited. In addition, 2.5 µM of benzavir-2 reduced infection in vitro in three to five orders of magnitude for five other flaviviruses: WNV, YFV, the tick-borne encephalitis virus, Japanese encephalitis virus, and dengue virus. In conclusion, benzavir-2 was a potent inhibitor of flavivirus infection, which supported the broad-spectrum antiviral activity of benzavir-2.

The pandemic threat from newly emerging viral diseases constitutes a major unsolved issue for global health. Antiviral therapy can play an important role in treating and preventing the spread of unprecedented viral infections. A repository of compounds exhibiting broad-spectrum antiviral activity against a series of different viral families would be an invaluable asset to be prepared for future pandemic threats. Utilizing an open innovation crowd-sourcing paradigm, we were able to identify a compound class of diphenylureas that exhibits in vitro antiviral activity against multiple viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), adenovirus, dengue virus, herpes, and influenza viruses. Compound 4 among the series exhibits strong activity against dengue virus, a growing global health problem with high medical need and no approved antiviral drug. The compounds are active against SARS-CoV-2 in a primary human stem cell-based mucociliary airway epithelium model and also active in vivo, as shown in a murine SARS-CoV-2 infection model. These results demonstrate the potential of the chemical class as antivirals on the one hand and the power of open innovation, crowd-sourcing, and repurposing on the other hand.