An increase in side-group hydrophobicity largely improves the potency of ritonavir-like inhibitors of CYP3A4.

Identification of structural determinants required for potent inhibition of drug-metabolizing cytochrome P450 3A4 (CYP3A4) could help develop safer drugs and more effective pharmacoenhancers. We utilize a rational inhibitor design to decipher structure-activity relationships in analogues of ritonavir, a highly potent CYP3A4 inhibitor marketed as pharmacoenhancer. Analysis of compounds with the R side-group as phenyl or naphthalene and R as indole or naphthalene in different stereo configuration showed that (i) analogues with the R-naphthalene tend to bind tighter and inhibit CYP3A4 more potently than the R-phenyl/indole containing counterparts; (ii) stereochemistry becomes a more important contributing factor, as the bulky side-groups limit the ability to optimize protein-ligand interactions; (iii) the relationship between the R/R configuration and preferential binding to CYP3A4 is complex and depends on the side-group functionality/interplay and backbone spacing; and (iv) three inhibitors, 5a-b and 7d, were superior to ritonavir (IC of 0.055-0.085 μM vs. 0.130 μM, respectively).