Abstract
A new series of 13 ritonavir-like inhibitors of human drug-metabolizing CYP3A4 was rationally designed to study the R(2) side-group and R(3) end-group interplay when the R(1) side-group is represented by phenyl. Spectral, functional, and structural characterization showed no improvement in the binding affinity and inhibitory potency of R(1)/R(2)-phenyl inhibitors upon elongation and/or fluorination of R(3)-Boc (tert-butyloxycarbonyl) or its replacement with benzenesulfonyl. When R(3) is pyridine, the impact of R(2)-phenyl-to-indole/naphthalene substitution was multidirectional and highly dependent on side-group stereo configuration. Overall, the R(2)-naphthalene/R(3)-pyridine containing 2f (R/S) was the series lead compound and one of the strongest binders/inhibitors designed thus far (K(s) = 0.009 μM; IC(50) = 0.10 μM). Introduction of a larger biphenyl or fluorene as R(2) did not lead to any improvements. Contrarily, fluorene-containing 13 was the series weakest binder and inhibitor (K(s) = 0.734 μM; IC(50) = 1.32 μM), implying that the fluorene moiety is too large to allow unrestricted access to the active site. The R(2)-biphenyl, however, can switch positions with R(3)-Boc to enable heme ligation. Thus, for small and chemically simple end-groups such as Boc and pyridine, the R(2)/R(3) interplay could lead to conformational rearrangement that would be difficult to foresee without structural information.