Rational Design of Hybrid SARS-CoV-2 Main Protease Inhibitors Guided by the Superimposed Cocrystal Structures with the Peptidomimetic Inhibitors GC-376, Telaprevir, and Boceprevir

Abstract

SARS-CoV-2 main protease (M pro ) is a cysteine protease that mediates the cleavage of viral polyproteins and is a validated antiviral drug target. M pro is highly conserved among all seven human coronaviruses, with certain M pro inhibitors having broad-spectrum antiviral activity. In this study, we designed two hybrid inhibitors UAWJ9-36-1 and UAWJ9-36-3 based on the superimposed X-ray crystal structures of SARS-CoV-2 M pro with GC-376, telaprevir, and boceprevir. Both UAWJ9-36-1 and UAWJ9-36-3 showed potent binding and enzymatic inhibition against the M pro ’s from SARS-CoV-2, SARS-CoV, MERS-CoV, HCoV-OC43, HCoV-NL63, HCoV-229E, and HCoV-HKU1. Cell-based Flip-GFP M pro assay results show that UAWJ9-36-1 and UAWJ9-36-3 inhibited the intracellular protease activity of SARS-CoV-2 M pro . In addition, UAWJ9-36-1 and UAWJ9-36-3 had potent antiviral activity against SARS-CoV-2, HCoV-OC43, HCoV-NL63, and HCoV-229E, with UAWJ9-36-3 being more potent than GC-376 in inhibiting SARS-CoV-2. Selectivity profiling revealed that UAWJ9-36-1 and UAWJ9-36-3 had an improved selectivity index over that of GC-376 against host cysteine proteases calpain I and cathepsin L, but not cathepsin K. The X-ray crystal structures of SARS-CoV-2 M pro with UAWJ9-36-1 and UAWJ9-36-3 were both solved at 1.9 Å, which validated our design hypothesis. Overall, hybrid inhibitors UAWJ9-36-1 and UAWJ9-36-3 are promising candidates to be further developed as broad-spectrum coronavirus antivirals.

Inhibition of the main protease of SARS-CoV-2 (Mpro) by repurposing/designing drug-like substances and utilizing nature’s toolbox of bioactive compounds

“…Xia et al [27] have used superposition of the crystal structures of inhibitors GC376, telaprevir and boceprevir to design two novel hybrid inhibitors, which combine the chemical groups of their parent compounds that result in the most interactions and most favorable binding. The designed inhibitors are UAWJ9-36-1 , as a hybrid of GC376 and telaprevir, and UAWJ9-36-3 , as a hybrid of GC376 and boceprevir.…”

A Review of the Current Landscape of SARS-CoV-2 Main Protease Inhibitors: Have We Hit the Bullseye Yet?

“…This assay is based on the thermal stabilization of a protein when it binds to a protein. A ΔTm shift of up to 18 °C has been observed for some SARS-CoV-2 inhibitors [ 51 , 70 , 73 , 92 , 93 ]. However, the thermal shift assay might not be suitable for analyzing non-covalent M-pro inhibitors [ 90 ].…”

Structural, Electronic, and Electrostatic Determinants for Inhibitor Binding to Subsites S1 and S2 in SARS-CoV-2 Main Protease

“…The hepatitis C virus clinical protease inhibitor boceprevir and the feline peritonitis virus protease inhibitor GC-376 were initially considered for drug repurposing, leading to the rational design of hybrid inhibitors. 26 − 28 In addition, noncovalent competitive and allosteric inhibitors have captured interest due to the availability of high-throughput virtual and experimental screening of large compound libraries that leverage new advances in supercomputing and fast X-ray crystallographic screening. 15 , 29 − 32 Compounds can be fed into structure-based drug design pipelines and chemically modified to improve their potency to inhibit SARS-CoV-2 M pro .…”