Design of SARS-CoV-2 PLpro Inhibitors for COVID-19 Antiviral Therapy Leveraging Binding Cooperativity

J Med Chem. 2022 Feb 24;65(4):2940-2955. doi: 10.1021/acs.jmedchem.1c01307. Epub 2021 Oct 19.


Antiviral agents that complement vaccination are urgently needed to end the COVID-19 pandemic. The SARS-CoV-2 papain-like protease (PLpro), one of only two essential cysteine proteases that regulate viral replication, also dysregulates host immune sensing by binding and deubiquitination of host protein substrates. PLpro is a promising therapeutic target, albeit challenging owing to featureless P1 and P2 sites recognizing glycine. To overcome this challenge, we leveraged the cooperativity of multiple shallow binding sites on the PLpro surface, yielding novel 2-phenylthiophenes with nanomolar inhibitory potency. New cocrystal structures confirmed that ligand binding induces new interactions with PLpro: by closing of the BL2 loop of PLpro forming a novel "BL2 groove" and by mimicking the binding interaction of ubiquitin with Glu167 of PLpro. Together, this binding cooperativity translates to the most potent PLpro inhibitors reported to date, with slow off-rates, improved binding affinities, and low micromolar antiviral potency in SARS-CoV-2-infected human cells.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antiviral Agents / chemical synthesis
  • Antiviral Agents / chemistry
  • Antiviral Agents / pharmacology*
  • Binding Sites / drug effects
  • COVID-19 / metabolism
  • COVID-19 Drug Treatment*
  • Coronavirus Papain-Like Proteases / antagonists & inhibitors*
  • Coronavirus Papain-Like Proteases / isolation & purification
  • Coronavirus Papain-Like Proteases / metabolism
  • Crystallography, X-Ray
  • Cysteine Proteinase Inhibitors / chemical synthesis
  • Cysteine Proteinase Inhibitors / chemistry
  • Cysteine Proteinase Inhibitors / pharmacology*
  • Humans
  • Microbial Sensitivity Tests
  • Microsomes, Liver / chemistry
  • Microsomes, Liver / metabolism
  • Models, Molecular
  • Pandemics
  • Surface Plasmon Resonance
  • Tumor Cells, Cultured


  • Antiviral Agents
  • Cysteine Proteinase Inhibitors
  • Coronavirus Papain-Like Proteases
  • papain-like protease, SARS-CoV-2