LAUSR

Covalent EGFR family inhibitors bind to and induce the degradation of the pseudokinase TRIB2 to kill cancer cells. Targeting pseudokinases with kinase inhibitors Pseudokinases are structurally similar to kinases but lack catalytic activity; instead, pseudokinases typically function as scaffolds, often promoting the degradation of substrate proteins by bringing them into close proximity with ubiquitin ligases. Two studies explored the structures and protein interactions of the pseudokinases TRIB1 (Jamieson et al.) and TRIB2 (Foulkes et al.). Their findings reveal new insights into the structural regulation of TRIB proteins and show that these proteins, which are implicated in leukemia and other cancers, can bind to clinically approved kinase inhibitors. Binding by these drugs caused structural changes in the TRIB proteins that deprotected them from degradation upon interacting with ubiquitin ligases, indicating that these drugs might be repurposed or redesigned to perturb the function of TRIBs in cancer patients. A major challenge associated with biochemical and cellular analysis of pseudokinases is a lack of target-validated small-molecule compounds with which to probe function. Tribbles 2 (TRIB2) is a cancer-associated pseudokinase with a diverse interactome, including the canonical AKT signaling module. There is substantial evidence that human TRIB2 promotes survival and drug resistance in solid tumors and blood cancers and therefore is of interest as a therapeutic target. The unusual TRIB2 pseudokinase domain contains a unique cysteine-rich C-helix and interacts with a conserved peptide motif in its own carboxyl-terminal tail, which also supports its interaction with E3 ubiquitin ligases. We found that TRIB2 is a target of previously described small-molecule protein kinase inhibitors, which were originally designed to inhibit the canonical kinase domains of epidermal growth factor receptor tyrosine kinase family members. Using a thermal shift assay, we discovered TRIB2-binding compounds within the Published Kinase Inhibitor Set (PKIS) and used a drug repurposing approach to classify compounds that either stabilized or destabilized TRIB2 in vitro. TRIB2 destabilizing agents, including the covalent drug afatinib, led to rapid TRIB2 degradation in human AML cancer cells, eliciting tractable effects on signaling and survival. Our data reveal new drug leads for the development of TRIB2-degrading compounds, which will also be invaluable for unraveling the cellular mechanisms of TRIB2-based signaling. Our study highlights that small molecule–induced protein down-regulation through drug “off-targets” might be relevant for other inhibitors that serendipitously target pseudokinases.