Identification of a novel allosteric binding site on the catalytic domain of NF-κB inducing kinase (NIK)†

NF-κB inducing kinase (NIK) is the central regulatory component of noncanonical NF-κB signalling and has been implicated in a variety of cancers and immune disorders. While NIK has been pursued as a target for such diseases through the design of orthosteric inhibitors, these inhibitors have not resulted in an approved drug. To develop new modalities for NIK-targeting by small molecules, we recently reported a class of chromanol fragments that bind to an unknown allosteric site on the catalytic domain of NIK. Here we report the design of a covalent probe to identify the location of this allosteric binding site. Acrylamide probe 2 (K_d: 24.5 μM) was determined to specifically adduct C573 out of 11 total cysteines on the catalytic domain of NIK, thereby identifying the allosteric binding site of our developed ligands.