A Chemproteomics Approach to Query the Degradable Kinome Using a Multi-kinase Degrader
Heterobifunctional molecule that recruits E3 ubiquitin ligases, such as cereblon, for targeted protein degradation is an emerging pharmacological strategy. A major unanswered question is how generally applicable this strategy is to all protein targets. In this study, we designed a multi-kinase degrader by conjugating a highly promiscuous kinase inhibitor with a cereblon-binding ligand, and used quantitative proteomics to discover 28 kinases, including BTK, PTK2, PTK2B, FLT3, AURKA, AURKB, TEC, ULK1, ITK and 9 members of the CDK family, as degradable. This set of kinases is only a fraction of the intracellular targets bound by the degrader, demonstrating that successful degradation requires more than target engagement. The results guided us to develop selective degraders for FLT3 and BTK, with potentials to improve disease treatment. Together, this study demonstrates an efficient approach to triage a gene family of interest to identify readily degradable targets for further studies and pre-clinical developments.
Sample Processing Protocol
Cells were treated with selected compounds, lysed, reduced and alkylated, subjected to Trypsin/LysC digest, labelled with TMT 10plex reagents, pre-fractionated using basic pH reverse phase chromatography, and subjected to LC-MS/MS using a SPS MS3 protocol on a Orbitrap Fusion. Details are provided in the referenced manuscript.
Data Processing Protocol
Data were processed with in house scripts, and statistical analysis carried out using the R framework.
Huang HT, Dobrovolsky D, Paulk J, Yang G, Weisberg EL, Doctor ZM, Buckley DL, Cho JH, Ko E, Jang J, Shi K, Choi HG, Griffin JD, Li Y, Treon SP, Fischer ES, Bradner JE, Tan L, Gray NS. A Chemoproteomic Approach to Query the Degradable Kinome Using a Multi-kinase Degrader. Cell Chem Biol. 2018 25(1):88-99.e6 PubMed: 29129717