Monoacidic Inhibitors of the Kelch-like ECH-Associated Protein 1: Nuclear Factor Erythroid 2‑Related Factor 2 (KEAP1:NRF2) Protein–Protein Interaction with High Cell Potency Identified by Fragment-Based Discovery

KEAP1 is the key regulator of the NRF2-mediated cytoprotective response, and increasingly recognized as a target for diseases involving oxidative stress. Pharmacological intervention has focused on molecules that decrease NRF2-ubiquitination through covalent modification of KEAP1 cysteine residues, but such electrophilic compounds lack selectivity and may be associated with off-target toxicity. We report here the first use of a fragment-based approach to directly target the KEAP1 Kelch–NRF2 interaction. X-ray crystallographic screening identified three distinct “hot-spots” for fragment binding within the NRF2 binding pocket of KEAP1, allowing progression of a weak fragment hit to molecules with nanomolar affinity for KEAP1 while maintaining drug-like properties. This work resulted in a promising lead compound which exhibits tight and selective binding to KEAP1, and activates the NRF2 antioxidant response in cellular and <i>in vivo</i> models, thereby providing a high quality chemical probe to explore the therapeutic potential of disrupting the KEAP1–NRF2 interaction