Abstract
Multiplexed small molecule inhibitors covalently bound to Sepharose beads (MIBs) were used to capture functional kinases in luminal, HER2-enriched and triple negative (basal-like and claudin-low) breast cancer cell lines and tumors. Kinase MIB-binding profiles at baseline without perturbation proteomically distinguished the four breast cancer subtypes. Understudied kinases, whose disease associations and pharmacology are generally unexplored, were highly represented in MIB-binding taxonomies and are integrated into signaling subnetworks with kinases that have been previously well characterized in breast cancer. Computationally it was possible to define subtypes using profiles of less than 50 of the more than 300 kinases bound to MIBs that included understudied as well as metabolic and lipid kinases. Furthermore, analysis of MIB-binding profiles established potential functional annotations for these understudied kinases. Thus, comprehensive MIBs-based capture of kinases provides a unique proteomics-based method for integration of poorly characterized kinases of the understudied kinome into functional subnetworks in breast cancer cells and tumors that is not possible using genomic strategies. The MIB-binding profiles readily defined subtype-selective differential adaptive kinome reprogramming in response to targeted kinase inhibition, demonstrating how MIB profiles can be used in determining dynamic kinome changes that result in subtype selective phenotypic state changes.
Highlights
In 2014 the NIH established an initiative entitled Illuminating the Druggable Genome (IDG) to determine the function of understudied proteins including kinases encoded in the human genome
Multiplexed small molecule inhibitors covalently bound to Sepharose beads (MIBs) were used to capture functional kinases in luminal, HER2-enriched and triple negative breast cancer cell lines and tumors
We focused on exploring the integration of understudied kinases into kinase networks within the context of breast cancer, which has three primary subtypes that include luminal as well as the majority of HER2+ breast cancers along with triple negative breast cancer (TNBC), that can itself be broken into basal-like and claudinlow subtypes [4]
Summary
In 2014 the NIH established an initiative entitled Illuminating the Druggable Genome (IDG) to determine the function of understudied proteins including kinases encoded in the human genome (https://commonfund.nih.gov/idg/index). To categorize our understanding of proteins in the human genome the IDG Knowledge Management Center (KMC) (http://targetcentral.ws/index) has developed a set of criteria for target development level (TDL) for druggable proteins such as kinases, G proteincoupled receptors and ion channels [1] This knowledge base was used to categorize the 634 kinases in the human genome as Tclin (50 kinases), Tchem (390 kinases), Tbio (163 kinases) and Tdark (31 kinases). Additional methods characterizing proteomic taxonomies are needed to understand signaling networks, of protein kinases due to their high druggability potential Important for this analysis of the cancer kinome is a characterization of understudied kinases (including Tbio and Tdark), which represent a third of the kinome and lack essential functional characterization as well as molecular tools for their manipulation and study [3]. These understudied kinases need to be functionally integrated into kinase networks for a global understanding of kinome dynamics to be achieved both at baseline and in response to perturbation
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