Chemical Biological Probes of the KEAP1/NRF2 Interaction

Kelch ECH-Associated Protein 1 (KEAP1) is an adaptor protein which functions as a negative regulator to Nuclear factor (erythroid-derived 2)-like 2 (NRF2). KEAP1 complexes NRF2 with the E3 ubiquitin ligase Cullin-3 (Cul3). When the body is placed under stress, either electrophilic or oxidative stress, KEAP1 releases NRF2, which translocates to the nucleus and binds to antioxidant response elements (AREs) in DNA promotor regions of genes to upregulate several phase II cytoprotective enzymes. Due to NRF2’s function, and its many pharmacological implications, KEAP1 has become a popular drug target. To date, all clinical NRF2 activators utilize strong electrophilic molecules to bind directly to KEAP1 and allosterically release NRF2. While this mechanism of action produces the desired physiological effect, the electrophilic activators may be non-selective. This trait produces difficulty in pinpointing if observed effects are solely due to NRF2 activation or activation of multiple biological systems by reactive electrophiles. To contribute to the understanding of this issue, we have performed a structure activity relationship on a series of non-electrophilic molecules based on a reported naphthalene-based activator that induces NRF2’s transcriptional activity through direct inhibition of the KEAP1/NRF2 interaction. An unstudied use of KEAP1 involves using the adaptor protein to attempt to access its complexed E3 ligand Cul3. The technologies of Proteolysis Targeting Chimeras (PROTACs) have been developed to use E3 ligases to target and degrade proteins that are not under their regulatory domain. These PROTACs can be more desirable than traditional inhibitors in that they function via catalytic degradation instead of occupancy derived inhibition. A bottle neck in the development of PROTAC technologies, however, is the lack of small molecules that selectively engage E3 ligases. To this issue, we set out to develop PROTACs targeting Cul3 through KEAP1 using our non-electrophilic probes as the KEAP1 ligand.