Abstract
Due to our interest in the chemical space of organic dyes to identify potential small-molecule inhibitors (SMIs) for protein-protein interactions (PPIs), we initiated a screen of such compounds to assess their inhibitory activity against the interaction between SARS-CoV-2 spike protein and its cognate receptor ACE2, which is the first critical step initiating the viral attachment and entry of this coronavirus responsible for the ongoing COVID-19 pandemic. As part of this, we found that methylene blue, a tricyclic phenothiazine compound approved by the FDA for the treatment of methemoglobinemia and used for other medical applications (including the inactivation of viruses in blood products prior to transfusion when activated by light), inhibits this interaction. We confirmed that it does so in a concentration-dependent manner with a low micromolar half-maximal inhibitory concentration (IC50 = 3 μM) in our protein-based ELISA-type setup, while chloroquine, siramesine, and suramin showed no inhibitory activity in this assay. Erythrosine B, which we have shown before to be a promiscuous SMI of PPIs, also inhibited this interaction. Methylene blue inhibited the entry of a SARS-CoV-2 spike bearing pseudovirus into ACE2-expressing cells with similar IC50 (3.5 μM). Hence, this PPI inhibitory activity could contribute to its antiviral activity against SARS-CoV-2 even in the absence of light by blocking its attachment to ACE2-expressing cells and making this inexpensive and widely available drug potentially useful in the prevention and treatment of COVID-19 as an oral or inhaled medication.
Highlights
Severe acute respiratory syndrome-coronavirus 2 (SARS-CoV2), a novel betacoronavirus and the most recent one of the seven coronaviruses (CoVs) known to infect humans, is responsible for COVID-19, which has been declared a pandemic by the World Health Organization in March 2020 and continues to spread worldwide (Liu et al, 2020; Matheson and Lehner, 2020; Moore and June, 2020)
As part of our work to identify small-molecule inhibitor (SMI) for co-signaling protein-protein interaction (PPI) that are essential for the activation and control of immune cells, we discovered that the chemical space of organic dyes, which is rich in strong protein binders, can offer a useful starting point
Results here confirm again the usefulness of our strategy to rely on the chemical space of organic dyes, known to contain strong protein binders, as a starting platform to identify SMI scaffolds for PPI inhibition
Summary
Severe acute respiratory syndrome-coronavirus 2 (SARS-CoV2), a novel betacoronavirus and the most recent one of the seven coronaviruses (CoVs) known to infect humans, is responsible for COVID-19, which has been declared a pandemic by the World Health Organization in March 2020 and continues to spread worldwide (Liu et al, 2020; Matheson and Lehner, 2020; Moore and June, 2020). CoVs use their glycosylated spike (S) protein to bind to their cognate cell surface receptors and initiate membrane fusion and virus entry For both SARS-CoV and SARS-CoV2, the S protein mediates entry into cells by binding to angiotensin converting enzyme 2 (ACE2) via its receptorbinding domain (RBD) followed by proteolytic activation by human proteases (Lan et al, 2020; Matheson and Lehner, 2020; Shang et al, 2020; Sivaraman et al, 2020). Antibodies can be quite effective PPI inhibitors, and they are highly target-specific and relatively stable in vivo They cannot reach intracellular targets and, as all other protein therapies, are hindered by problems such as low solubility, propensity for immunogenicity, long elimination half-lives, lack of oral bioavailability, product heterogeneity, and possible manufacturing and storage stability issues. There are >40 PPIs targeted by SMIs that are in preclinical development, and two such SMIs are approved for clinical use (venetoclax and lifitegrast) (Arkin and Wells, 2004; Milroy et al, 2014; Scott et al, 2016; Bojadzic and Buchwald, 2018)
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