Hepatitis C Virus NS3/4A Protease Inhibitors Incorporating Flexible P2 Quinoxalines Target Drug Resistant Viral Variants

A substrate envelope-guided design
strategy is reported for improving
the resistance profile of HCV NS3/4A protease inhibitors. Analogues
of 5172-mcP1P3 were designed by incorporating diverse quinoxalines
at the P2 position that predominantly interact with the invariant
catalytic triad of the protease. Exploration of structure–activity
relationships showed that inhibitors with small hydrophobic substituents
at the 3-position of P2 quinoxaline maintain better potency against
drug resistant variants, likely due to reduced interactions with residues
in the S2 subsite. In contrast, inhibitors with larger groups at this
position were highly susceptible to mutations at Arg155, Ala156, and
Asp168. Excitingly, several inhibitors exhibited exceptional potency
profiles with EC50 values ≤5 nM against major drug
resistant HCV variants. These findings support that inhibitors designed
to interact with evolutionarily constrained regions of the protease,
while avoiding interactions with residues not essential for substrate
recognition, are less likely to be susceptible to drug resistance.