Development of a Cell-Based Luciferase Complementation Assay for Identification of SARS-CoV-2 3CLpro Inhibitors.

Viruses
Jonathan M O RawsonWei-Shau Hu

Abstract

The 3C-like protease (3CLpro) of SARS-CoV-2 is considered an excellent target for COVID-19 antiviral drug development because it is essential for viral replication and has a cleavage specificity distinct from human proteases. However, drug development for 3CLpro has been hindered by a lack of cell-based reporter assays that can be performed in a BSL-2 setting. Current efforts to identify 3CLpro inhibitors largely rely upon in vitro screening, which fails to account for cell permeability and cytotoxicity of compounds, or assays involving replication-competent virus, which must be performed in a BSL-3 facility. To address these limitations, we have developed a novel cell-based luciferase complementation reporter assay to identify inhibitors of SARS-CoV-2 3CLpro in a BSL-2 setting. The assay is based on a lentiviral vector that co-expresses 3CLpro and two luciferase fragments linked together by a 3CLpro cleavage site. 3CLpro-mediated cleavage results in a loss of complementation and low luciferase activity, whereas inhibition of 3CLpro results in 10-fold higher levels of luciferase activity. The luciferase reporter assay can easily distinguish true 3CLpro inhibition from cytotoxicity, a powerful feature that should reduce false po...Continue Reading

References

Jan 1, 1990·Journal of Enzyme Inhibition·T SasakiT Murachi
Sep 27, 1996·Journal of Medicinal Chemistry·Z LiJ C Powers
Dec 13, 2006·Biochemistry·Jennifer BarrilaErnesto Freire
Sep 18, 2008·Bioorganic & Medicinal Chemistry Letters·Arun K GhoshAndrew D Mesecar
Mar 19, 2020·The New England Journal of Medicine·Bin CaoChen Wang
May 16, 2020·Journal of Biomolecular Structure & Dynamics·Muhammad Tahir KhanDong-Qing Wei
May 24, 2020·The New England Journal of Medicine·John H BeigelUNKNOWN ACTT-1 Study Group Members
Jun 2, 2020·International Journal of Infectious Diseases : IJID : Official Publication of the International Society for Infectious Diseases·Sandra De MeyerSandra Ciesek
Jul 15, 2020·The New England Journal of Medicine·Lisa A JacksonUNKNOWN mRNA-1273 Study Group
Jul 17, 2020·Antimicrobial Agents and Chemotherapy·Hui-Chen HungJohn Tsu-An Hsu
Aug 3, 2020·Bioorganic & Medicinal Chemistry Letters·Sven Ullrich, Christoph Nitsche
Aug 5, 2020·Science Translational Medicine·Athri D RathnayakeKyeong-Ok Chang
Aug 8, 2020·Antimicrobial Agents and Chemotherapy·Natalia Fintelman-RodriguesThiago Moreno L Souza
Aug 14, 2020·Nature·Mark J MulliganKathrin U Jansen
Aug 15, 2020·BioRxiv : the Preprint Server for Biology·Sho IketaniDavid D Ho
Aug 21, 2020·MBio·Shin-Ichiro HattoriHiroaki Mitsuya
Aug 22, 2020·JAMA : the Journal of the American Medical Association·Christoph D SpinnerUNKNOWN GS-US-540-5774 Investigators
Aug 28, 2020·Journal of Virology·Heather M FroggattNicholas S Heaton
Aug 29, 2020·Nature Communications·Wayne VuongM Joanne Lemieux
Sep 2, 2020·BioRxiv : the Preprint Server for Biology·Samuel J ResnickAlejandro Chavez
Sep 11, 2020·BioRxiv : the Preprint Server for Biology·Nir DraymanSavaş Tay
Oct 17, 2020·ACS Pharmacology & Translational Science·Wei ZhuWei Zheng
Oct 29, 2020·The New England Journal of Medicine·Peter ChenUNKNOWN BLAZE-1 Investigators
Oct 30, 2020·Nature Reviews. Microbiology·Philip V'kovskiVolker Thiel
Nov 18, 2020·BioRxiv : the Preprint Server for Biology·Seyad Arad MoghadasiReuben S Harris
Dec 3, 2020·The New England Journal of Medicine·UNKNOWN WHO Solidarity Trial ConsortiumSoumya Swaminathan
Dec 4, 2020·The New England Journal of Medicine·Alicia T WidgeUNKNOWN mRNA-1273 Study Group
Dec 22, 2020·Frontiers in Molecular Biosciences·Wen CuiHaitao Yang

❮ Previous
Next ❯

Methods Mentioned

BETA
transfections
Assay
transfection
fluorescence
flow cytometry

Related Concepts

Related Feeds

Antivirals (ASM)

Antivirals are medications that are used specifically for treating viral infections. Discover the latest research on antivirals here.

Antivirals

Antivirals are medications that are used specifically for treating viral infections. Discover the latest research on antivirals here.