SARS-CoV-2 3CLpro mutations selected in a VSV-based system confer resistance to nirmatrelvir, ensitrelvir, and GC376
et al., Science Translational Medicine,
SARS-CoV-2 3CLpro mutations selected in a VSV-based system confer resistance to nirmatrelvir, ensitrelvir, and..
, Science Translational Medicine, doi:10.1126/scitranslmed.abq7360 (In Vitro)
and In Silico
study showing selection of resistant mutations with nirmatrelvir use.
Several mutations were identified that confer resistance to 3CLpro
inhibitors nirmatrelvir, ensitrelvir, and GC376. Authors note that most of these have already been found in existing SARS-CoV-2 sequences.
Authors argue for highly selective use because extensive, unselective use is expected to rapidly lead to emergence of drug resistance.
Heilmann et al., 11 Jan 2023, peer-reviewed, 15 authors.
In Vitro studies are an important part of preclinical research, however results may be very different in vivo.
Abstract: S C I E N C E T R A N S L AT I O N A L M E D I C I N E | R E S E A R C H A R T I C L E
C O R O N AV I R U S
SARS-CoV-2 3CLpro mutations selected in a VSV-based
system confer resistance to nirmatrelvir, ensitrelvir,
Emmanuel Heilmann1†*, Francesco Costacurta1†, Seyed Arad Moghadasi2, Chengjin Ye3,
Matteo Pavan4, Davide Bassani4, Andre Volland1, Claudia Ascher5,
Alexander Kurt Hermann Weiss5, David Bante1, Reuben S. Harris2,6,7, Stefano Moro4,
Bernhard Rupp8,9, Luis Martinez-Sobrido3, Dorothee von Laer1*
Copyright © 2023 The
for the Advancement
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License 4.0 (CC BY).
Protease inhibitors are among the most powerful antiviral drugs. Nirmatrelvir is the first protease inhibitor specifically developed against the SARS-CoV-2 protease 3CLpro that has been licensed for clinical use. To identify
mutations that confer resistance to this protease inhibitor, we engineered a chimeric vesicular stomatitis virus
(VSV) that expressed a polyprotein composed of the VSV glycoprotein (G), the SARS-CoV-2 3CLpro, and the VSV
polymerase (L). Viral replication was thus dependent on the autocatalytic processing of this precursor protein by
3CLpro and release of the functional viral proteins G and L, and replication of this chimeric VSV was effectively
inhibited by nirmatrelvir. Using this system, we applied nirmatrelvir to select for resistance mutations. Resistance was confirmed by retesting nirmatrelvir against the selected mutations in additional VSV-based
systems, in an independently developed cellular system, in a biochemical assay, and in a recombinant SARSCoV-2 system. We demonstrate that some mutants are cross-resistant to ensitrelvir and GC376, whereas
others are less resistant to these compounds. Furthermore, we found that most of these resistance mutations
already existed in SARS-CoV-2 sequences that have been deposited in the NCBI and GISAID databases, indicating
that these mutations were present in circulating SARS-CoV-2 strains.
In late 2019, the zoonotic transmission of a new coronavirus, severe
acute respiratory syndrome coronavirus 2 (SARS-CoV-2), into the
human population (1) led to worldwide efforts to find effective
treatments against the various pathologies caused by the virus. Inhibitors of viral enzymes, such as proteases, have proven to be highly
potent drugs in the treatment of HIV and hepatitis C virus infections. However, resistant viruses rapidly emerge unless the protease
inhibitors are given in combination with other directly acting antivirals (2, 3). SARS-CoV-2 encodes two proteases. The 3-chymotrypsin–like protease (3CLpro) cleaves 11 sites in the viral polyproteins
pp1a and pp1ab and is also referred to as the main protease or nonstructural protein 5, indicating that it cleaves more sites than the
second protease and its location within the polyproteins, respectively (4). The second viral protease, papain-like protease (PLpro),
cleaves three additional sites in pp1a and pp1ab (5). Thus, both
Institute of Virology, Medical University of Innsbruck, Innsbruck, 6020, Austria.
Department of Biochemistry, Molecular Biology and Biophysics, Institute for Molecular Virology, University of Minnesota, Minneapolis, MN 55455, USA. 3Texas Biomedical Research Institute, San Antonio, TX 78229, USA. 4Molecular Modeling
Section (MMS), Department of..
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