TY - JOUR
T1 - Validation and Invalidation of SARS-CoV-2 Papain-like Protease Inhibitors
AU - Ma, Chunlong
AU - Wang, Jun
N1 - Funding Information:
This research was supported by the National Institutes of Health (NIH) (grants AI147325, AI157046, and AI158775) and the Arizona Biomedical Research Centre Young Investigator grant (ADHS18-198859) to J.W.
Publisher Copyright:
© 2022 American Chemical Society
PY - 2022/2/11
Y1 - 2022/2/11
N2 - SARS-CoV-2 encodes two viral cysteine proteases, the main protease (Mpro) and the papain-like protease (PLpro), both of which are validated antiviral drug targets. PLpro is involved in the cleavage of viral polyproteins as well as immune modulation by removing ubiquitin and interferon-stimulated gene product 15 (ISG15) from host proteins. Therefore, targeting PLpro might be a two-pronged approach. Several compounds including YM155, cryptotanshinone, tanshinone I, dihydrotanshinone I, tanshinone IIA, SJB2-043, 6-thioguanine, and 6-mercaptopurine were recently identified as SARS-CoV-2 PLpro inhibitors through high-throughput screenings. In this study, we aim to validate/invalidate the reported PLpro inhibitors using a combination of PLpro target-specific assays including enzymatic FRET assay, thermal shift binding assay (TSA), and cell-based FlipGFP assay. Collectively, our results showed that all compounds tested either did not show binding or led to denaturation of PLpro in the TSA binding assay, which might explain their weak enzymatic inhibition in the FRET assay. In addition, none of the compounds showed cellular PLpro inhibition as revealed by the FlipGFP assay. Therefore, more efforts are needed to search for potent and specific SARS-CoV-2 PLpro inhibitors.
AB - SARS-CoV-2 encodes two viral cysteine proteases, the main protease (Mpro) and the papain-like protease (PLpro), both of which are validated antiviral drug targets. PLpro is involved in the cleavage of viral polyproteins as well as immune modulation by removing ubiquitin and interferon-stimulated gene product 15 (ISG15) from host proteins. Therefore, targeting PLpro might be a two-pronged approach. Several compounds including YM155, cryptotanshinone, tanshinone I, dihydrotanshinone I, tanshinone IIA, SJB2-043, 6-thioguanine, and 6-mercaptopurine were recently identified as SARS-CoV-2 PLpro inhibitors through high-throughput screenings. In this study, we aim to validate/invalidate the reported PLpro inhibitors using a combination of PLpro target-specific assays including enzymatic FRET assay, thermal shift binding assay (TSA), and cell-based FlipGFP assay. Collectively, our results showed that all compounds tested either did not show binding or led to denaturation of PLpro in the TSA binding assay, which might explain their weak enzymatic inhibition in the FRET assay. In addition, none of the compounds showed cellular PLpro inhibition as revealed by the FlipGFP assay. Therefore, more efforts are needed to search for potent and specific SARS-CoV-2 PLpro inhibitors.
KW - FlipGFP
KW - GRL0617
KW - SARS-CoV-2
KW - antiviral
KW - papain-like protease
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U2 - 10.1021/acsptsci.1c00240
DO - 10.1021/acsptsci.1c00240
M3 - Article
AN - SCOPUS:85124150605
SN - 2575-9108
VL - 5
SP - 102
EP - 109
JO - ACS Pharmacology and Translational Science
JF - ACS Pharmacology and Translational Science
IS - 2
ER -