| Assay Method Information | |
| | Antiviral activity assay |
| Description: | To further substantiate the enzymatic inhibition results in vitro, we evaluated whether these compounds could prevent viral replication in cell-based assays. As shown in Fig. 4a, quantitative real-time RT PCR (qRT PCR) demonstrated that, among these compounds, ebselen and N3 showed the strongest antiviral effects at a concentration of 10 μM treatment in SARS-CoV-2-infected Vero cells. We performed a plaque-reduction assay (Extended Data Fig. 8) to further assess the efficacy of these two compounds in protecting cells. Ebselen and N3 displayed inhibition against SARS-CoV-2 with individual half-maximal effective concentration (EC50) values of 4.67 μM and 16.77 μM, respectively (Fig. 4b, c). The dose response curves suggest that both of these compounds may be able to penetrate the cellular membrane to access their targets. Ebselen is an organoselenium compound with anti-inflammatory, anti-oxidant and cytoprotective properties. This compound has previously been investigated for the treatment of multiple diseases, including bipolar disorders26 and hearing loss27,28. Ebselen has extremely low cytotoxicity (the median lethal dose in rats is >4,600 mg kg−1, when taken orally)29, and its safety in humans has been evaluated in a number of clinical trials27,28,30. These data strongly suggest the clinical potential of ebselen for the treatment of coronaviruses. It is also interesting to note that cinanserin displayed moderate inhibition against SARS-CoV-2 with an EC50 value of 20.61 μM, as shown from qRT PCR analysis (Extended Data Fig. 4). This value is superior to that in the enzymatic inhibition assay, which suggests that cinanserin might have multidrug targets in preventing viral infection. In further studies, the selection and characterization of drug-resistant mutants will help to clarify the mode of action of cinanserin. |
| Affinity data for this assay | |
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