Optimization of direct small molecule inhibitors of Keap1-Nrf2 protein-protein interaction

The Keap1-Nrf2-ARE signaling system that regulates the transcription and subsequent expression of cellular cytoprotective proteins and enzymes plays a crucial role in preventing pathological conditions exacerbated by the overproduction of oxidative stress. Thus, a noncovalent inhibitor that directly interrupts the Keap1-Nrf2 protein-protein interaction (PPI) and eventually leads to stimulation of related antioxidant responses has attracted a great deal of attention as a preventive and therapeutic agent for the treatment of oxidative stress-related diseases. Considering that the cellular potency or drug-like properties of direct inhibitors have remained as significant factors to be overcome, we have focused on discovering diverse classes of potent small molecule inhibitors based on known compounds via two different rational approaches. In this study, we found a series of 1,4-diamino phenyl and naphthalene scaffolds containing an oxygen-linked substituent that could be modified, contributing to greater structural diversity of the core moiety. The SAR studies revealed that O-linked derivatives displayed potent inhibitory activity in the submicromolar or nanomolar range. Among them, compound 120 is the most promising noncovalent inhibitor, with 64.5 nM in the fluorescent polarization (FP) assay and 14.2 nM in a time-resolved fluorescence resonance energy transfer (TR-FRET) assay. In addition, we designed three different scaffolds as direct inhibitors of Keap1-Nrf2 PPI via a molecular hybridization strategy, and identified THIQ and N-phenylglycine analogs which exhibited good potency for a preliminary SAR study. As a result, THIQ analog 139c was found to be the most potent with an IC50 value of 0.37 μM among a series of THIQ compounds developed by our group and other groups. Among the N-phenylglycine derivatives, compound 191b is the most active with an IC50 value of 183.4 nM in the FP assay and 107.5 nM in a TR-FRET assay. The optimized new scaffolds may serve as a promising starting point for further biological evaluation and rational optimization as a Nrf2 activator.Ph.D.Includes bibliographical reference