Kinase Activity Profiling in Soft Tissue Sarcoma
Arguably, the 518 protein kinases which make up the human kinome constitute the most tractable group of new cancer targets. However, despite its well-established ‘druggability’ and central position within key signaling networks, very few kinases have been studied in detail. In sarcoma, alterations in select kinases are thought to causally contribute to disease initiation, progression and therapeutic resistance (eg. PDGFRA, CDK4, ALK). We hypothesize that additional kinases display altered activity in sarcoma, and that these kinases represent immense diagnostic, prognostic and therapeutic value. Until recently, technical hurdles have prevented kinome-wide activity assessment and study; the understudied kinome may contain more than 400 kinases. We have developed multiplex inhibitor beads and protein mass spectrometry (MIB/MS) to isolate and quantify ~275 protein kinases in a single mass spectrometry run. Kinase capture is highly reproducible and is a function of kinase affinity for different immobilized inhibitors—activated kinases preferentially bind and inactive kinases do not. Here, we will use MIB/MS to define the activation state of the kinome across 50 soft tissue human sarcomas (AIM#1) and 16 well-characterized human sarcoma cell lines (AIM#2). Statistical, computational and informatic analyses will look for minimal kinase signatures that classify and diagnose human sarcoma. The data will also be interpreted within the larger protein-protein interaction network, revealing signaling hubs that represent new therapeutic opportunities. Quite simply, the proposed experiments will capture and quantify 80% of the expressed kinome across more than 50 sarcoma tumor samples, resulting in new diagnostics and therapeutic targets.