LAUSR

Thermodynamic parameters were determined for structurally-related inhibitors of HCV NS3 protease to assess how binding entropies and enthalpies vary with incremental changes at the P2 and P3 inhibitor subsites. Changing the heterocyclic substituent at P2 from a pyridyl to a 7-methoxy-2-phenyl-4-quinolyl group leads to a 710-fold increase in affinity. Annelating a benzene ring onto a pyridine ring leads to quinoline-derived inhibitors having higher affinities, but the individual enthalpy and entropy contributions are markedly different for each ligand pair. Introducing a phenyl group at C2 of the heterocyclic ring at P2 uniformly leads to higher affinity analogs with more favorable binding entropies, while adding a methoxy group at C7 of the quinoline ring at P2 provides derivatives with more favorable binding enthalpies. Significant enthalpy/entropy compensation is observed for structural changes made to inhibitors lacking a 2-phenyl substituent, whereas favorable changes in both binding enthalpies and entropies accompany structural modifications when a 2-phenyl group is present. Overall, binding energetics of inhibitors having a 2-phenyl-4-quinolyl group at P2 are dominated by entropic effects, whereas binding of the corresponding norphenyl analogs are primarily enthalpy driven. Notably, the reversal from an entropy driven association to an enthalpy driven one for this set of inhibitors also correlates with alternate binding modes. When the steric bulk of the side chain at P3 is increased from a hydrogen atom to a tert-butyl group, there is a 770-fold improvement in affinity. The 30-fold increase resulting from the first methyl group is solely the consequence of a more favorable change in entropy, whereas subsequent additions of methyl groups leads to modest increases in affinity that arise primarily from incremental improvements in binding enthalpies accompanied with smaller favorable entropic contributions.