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Updated: Sep 5, 2025
Detection of SARS-CoV-2 Neutralizing Antibodies using High-Throughput Fluorescent Imaging of Pseudovirus Infection
Published on: June 5, 2021
[Identifying SARS-CoV-2 main protease inhibitors by a novel sandwich-like fluorescence polarization screening assay]
Haohao Yan1, Gangan Yan1, Haiyan Qi1
1Institute for Drug Screening and Evaluation, Wannan Medical College, Wuhu 241002, Anhui, China.
Abstract:
SARS-CoV-2 main protease (Mpro) is responsible for polyprotein cleavage to release non-structural proteins (nsps) for viral genomic RNA replication, and its homologues are absent in human cells. Therefore, Mpro has been regarded as one of the ideal drug targets for the treatment of coronavirus disease 2019 (COVID-19). In this study, we first combined the fluorescence polarization (FP) technique with biotin-avidin system (BAS) to develop a novel sandwich-like FP screening assay for quick discovery of SARS-CoV-2 Mpro inhibitors from a natural product library. With this screening assay, anacardic acid (AA) and 1, 2, 3, 4, 6-O-pentagalloylglucose (PGG) were found to be the competitive inhibitor and mixed-type inhibitor targeting Mpro, respectively. Importantly, our results showed that the majority of the reported Mpro inhibitors are promiscuous cysteine inhibitors that are not specific to Mpro. In summary, this novel sandwich-like FP screening assay is simple, sensitive, and robust, which is ideal for large-scale screening. Natural products AA and PGG will be the promising lead compounds for generating more potent antiviral agents targeting Mpro, and the stringent hit validation at the early stage of drug discovery is urgently needed.
Insights
Researchers developed a novel assay to screen for SARS-CoV-2 Mpro inhibitors. Natural products anacardic acid and pentagalloylglucose were identified as promising lead compounds for COVID-19 drug discovery.
Area of Science:
- Biochemistry
- Drug Discovery
- Virology
Background:
- SARS-CoV-2 main protease (Mpro) is crucial for viral replication and lacks human homologues, making it an ideal drug target for COVID-19.
- Existing Mpro inhibitors often lack specificity, highlighting the need for improved screening methods.
Purpose of the Study:
- To develop a novel, sensitive, and robust screening assay for identifying SARS-CoV-2 Mpro inhibitors.
- To discover novel inhibitors from a natural product library targeting SARS-CoV-2 Mpro.
Main Methods:
- Combined fluorescence polarization (FP) with the biotin-avidin system (BAS) to create a sandwich-like FP assay.
- Screened a natural product library for inhibitors of SARS-CoV-2 Mpro.
Main Results:
- Anacardic acid (AA) was identified as a competitive Mpro inhibitor.
- 1,2,3,4,6-O-pentagalloylglucose (PGG) was identified as a mixed-type Mpro inhibitor.
- Demonstrated that many previously reported Mpro inhibitors are non-specific cysteine inhibitors.
Conclusions:
- The novel sandwich-like FP assay is effective for large-scale screening of Mpro inhibitors.
- AA and PGG are promising lead compounds for developing potent antiviral agents against SARS-CoV-2.
- Emphasized the importance of stringent hit validation in early drug discovery for Mpro inhibitors.
