Evans-blue for COVID-19

Effective SARS-CoV-2 antiviral drugs are desperately needed. The SARS-CoV-2 main protease (Mpro) appears as an attractive target for drug development. We show that the existing pharmacopeia contains many drugs with potential for therapeutic repurposing as selective and potent inhibitors of SARS-CoV-2 Mpro. We screened a collection of ~6,070 drugs with a previous history of use in humans for compounds that inhibit the activity of Mpro in vitro and found ~50 compounds with activity against Mpro. Subsequent dose validation studies demonstrated 8 dose responsive hits with an IC50 ≤ 50 μM. Hits from our screen are enriched with hepatitis C NS3/4A protease targeting drugs including boceprevir, ciluprevir. narlaprevir, and telaprevir. This work suggests previous large-scale commercial drug development initiatives targeting hepatitis C NS3/4A viral protease should be revisited because some previous lead compounds may be more potent against SARS-CoV-2 Mpro than boceprevir and suitable for rapid repurposing.

ABSTRACTInhibitors of the protein-protein interaction (PPI) between the SARS-CoV-2 spike protein and ACE2, which acts as a ligand-receptor pair that initiates the viral attachment and cellular entry of this coronavirus causing the ongoing COVID-19 pandemic, are of considerable interest as potential antiviral agents. While blockade of such PPIs with small molecules is more challenging than with antibodies, small-molecule inhibitors (SMIs) might offer alternatives that are less strain- and mutation-sensitive, suitable for oral or inhaled administration, and more controllable / less immunogenic. Here, we report the identification of SMIs of this PPI by screening our compound-library that is focused on the chemical space of organic dyes. Among promising candidates identified, several dyes (Congo red, direct violet 1, Evans blue) and novel drug-like compounds (DRI-C23041, DRI-C91005) inhibited the interaction of hACE2 with the spike proteins of SARS-CoV-2 as well as SARS-CoV with low micromolar activity in our cell-free ELISA-type assays (IC50s of 0.2-3.0 μM); whereas, control compounds, such as sunset yellow FCF, chloroquine, and suramin, showed no activity. Protein thermal shift assays indicated that the SMIs identified here bind SARS-CoV-2-S and not ACE2. Selected promising compounds inhibited the entry of a SARS-CoV-2-S expressing pseudovirus into ACE2-expressing cells in concentration-dependent manner with low micromolar IC50s (6-30 μM). This provides proof-of-principle evidence for the feasibility of small-molecule inhibition of PPIs critical for coronavirus attachment/entry and serves as a first guide in the search for SMI-based alternative antiviral therapies for the prevention and treatment of diseases caused by coronaviruses in general and COVID-19 in particular.