Open Source Repurposing Reveals Broad-Spectrum Antiviral Activity of Diphenylureas

Abstract

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.

Keywords: broadband antiviral compounds; pandemic preparedness; repurposing.

Figure 1

Structures of TIE-2 inhibitors compound 1–4.

Figure 2

The dose–response activity of the three TIE-2 inhibitors from the Merck Mini Library. The results from the primary screening are displayed in (A) against HAdV and (B) against RVFV in A549 cells. Ñ = visual compound precipitation; * = visual host cell toxicity. (C) Activity against SARS-CoV-2 in VeroE6 cells using plaque reduction assay. * = visual host cell toxicity.

Figure 3

The dose–response antiviral activity of compounds 1 and 4 (Figure 1) and remdesivir as a positive control against SARS-CoV-2 in Hela3ACE2 cells. Graphs show antiviral activity measured with a SARS-CoV-2 immunofluorescence signal leading to identification of infected cells with 0% activity equals 100% infected cells (blue curve), total cells per well in SARS-CoV-2 infected cell test with 0% activity equaling no change vs. control (yellow curve), total cells per well in HeLa-ACE2 uninfected cell control (red curve).

Figure 4

Dose–response antiviral activity against SARS-CoV-2 of compound 4 (Figure 1) and remdesivir as a positive control in Calu3 cells. The graphs show antiviral activity measured with a SARS-CoV-2 immunofluorescence signal, leading to the identification of infected cells with 0% activity equaling 100% infected cells (green curve), total cells per well in the SARS-CoV-2 infected cell test with 0% activity equaling no change vs. the control (yellow curve), and total cells per well in the HeLa-ACE2 uninfected cell control (red curve).

Figure 5

Antiviral activity dose response curves for compound 1 and compound 4 against HAdV, dengue virus, and influenza virus.

Figure 6

Activity in human primary mucociliary airway stem cell-based SARS-CoV-2 infection model for compound 4 compared to remdesivir and molnupiravir tested at 10 µM.

Figure 7

In vivo antiviral efficacy of compound 4 against SARS-CoV-2. Mice exhibited increased survival (A), reduced body weight loss (B), and reduced viral load in lungs (C) after oral treatment b.i.d. with 60 mg/kg compound 4 in 0.5% methocel/0.25% Tween-20/water in comparison to placebo. EIDD-2801 (molnupiravir) dosed same way was used as positive control. (ns p > 0.05, * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001).