Molecular and Structural Mechanism of Pan-Genotypic HCV NS3/4A Protease Inhibition by Glecaprevir

Hepatitis C virus,
causative agent of chronic viral hepatitis,
infects 71 million people worldwide and is divided into seven genotypes
and multiple subtypes with sequence identities between 68 to 82%.
While older generation direct-acting antivirals had varying effectiveness
against different genotypes, the newest NS3/4A protease inhibitors
including glecaprevir (GLE) have pan-genotypic activity. The structural
basis for pan-genotypic inhibition and effects of polymorphisms on
inhibitor potency were not well-known due to lack of crystal structures
of GLE-bound NS3/4A or genotypes other than 1. In this study, we determined
the crystal structures of NS3/4A from genotypes 1a, 3a, 4a, and 5a
in complex with GLE. Comparison with the highly similar grazoprevir
indicated the mechanism of GLE’s drastic improvement in potency.
We found that, while GLE is highly potent against wild-type NS3/4A
of all genotypes, specific resistance-associated substitutions (RASs)
confer orders of magnitude loss in inhibition. Our crystal structures
reveal molecular mechanisms behind pan-genotypic activity of GLE,
including potency loss due to RASs at D168. Our structures permit
for the first time analysis of changes due to polymorphisms among
genotypes, providing insights into design principles that can aid
future drug development and potentially can be extended to other proteins.