title: Multiplex imaging and cellular target identification of kinase inhibitors via an affinity-based proteome profiling approach.
abstract: MLN8237 is a highly potent and presumably selective inhibitor of Aurora kinase A (AKA) and has shown promising antitumor activities. Like other kinase inhibitors which target the ATP-binding site of kinases, MLN8237 might be expected to have potential cellular off-targets. Herein, we report the first photoaffinity-based, small molecule AKA probe capable of both live-cell imaging of AKA activities and in situ proteome profiling of potential off-targets of MLN8237 (including AKA-associating proteins). By using two mutually compatible, bioorthogonal reactions (copper-catalyzed azide-alkyne cycloaddition chemistry and TCO-tetrazine ligation), we demostrate small molecule-based multiplex bioimaging for simultaneous in situ monitoring of two important cell-cycle regulating kinases (AKA and CDK1). A broad range of proteins, as potential off-targets of MLN8237 and AKA's-interacting partners, is subsequently identified by affinity-based proteome profiling coupled with large-scale LC-MS/MS analysis. From these studies, we discover novel AKA interactions which were further validated by cell-based immunoprecipitation (IP) experiments.
authors.affiliation: Department of Chemistry, National University of Singapore, 3 Science Drive 3, Singapore 117543.
authors.forename: Ying
authors.initials: Y
authors.lastname: Su
authors.affiliation: Department of Chemistry, National University of Singapore, 3 Science Drive 3, Singapore 117543.
authors.forename: Sijun
authors.initials: S
authors.lastname: Pan
authors.affiliation: Department of Chemistry, National University of Singapore, 3 Science Drive 3, Singapore 117543.
authors.forename: Zhengqiu
authors.initials: Z
authors.lastname: Li
authors.affiliation: Department of Chemistry, National University of Singapore, 3 Science Drive 3, Singapore 117543.
authors.forename: Lin
authors.initials: L
authors.lastname: Li
authors.affiliation: Department of Chemistry, National University of Singapore, 3 Science Drive 3, Singapore 117543.
authors.forename: Xiaoyuan
authors.initials: X
authors.lastname: Wu
authors.affiliation: School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551.
authors.forename: Piliang
authors.initials: P
authors.lastname: Hao
authors.affiliation: School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551.
authors.forename: Siu Kwan
authors.initials: SK
authors.lastname: Sze
authors.affiliation: Department of Chemistry, National University of Singapore, 3 Science Drive 3, Singapore 117543.
authors.forename: Shao Q
authors.initials: SQ
authors.lastname: Yao
mesh_terms: D002846:Chromatography, Affinity; D002853:Chromatography, Liquid; D006367:HeLa Cells; D006801:Humans; D057054:Molecular Imaging / Q000379:methods*; D015335:Molecular Probes / Q000378:metabolism; D058990:Molecular Targeted Therapy; D047428:Protein Kinase Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D020543:Proteome / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*; D053719:Tandem Mass Spectrometry
chemical_list: D015335:Molecular Probes; D047428:Protein Kinase Inhibitors; D020543:Proteome