title: Characterization of the Tyrosine Kinase-Regulated Proteome in Breast Cancer by Combined use of RNA interference (RNAi) and Stable Isotope Labeling with Amino Acids in Cell Culture (SILAC) Quantitative Proteomics.
abstract: Tyrosine kinases (TKs) are central regulators in cellular activities and perturbations of TK signaling contribute to oncogenesis. However, less than half of the TKs have been thoroughly studied and a global functional analysis of their proteomic portrait is lacking. Here we conducted a combined approach of RNA interference (RNAi) and stable isotope labeling with amino acids in cell culture (SILAC)-based quantitative proteomics to decode the TK-regulated proteome and associated signaling dynamics. As a result, a broad proteomic repertoire modulated by TKs was revealed, upon silencing of the 65 TKs expressed in MCF7 breast cancer cells. This yielded 10 new distinctive TK clusters according to similarity in TK-regulated proteome, each characterized by a unique signaling signature in contrast to previous classifications. We provide functional analyses and identify critical pathways for each cluster based on their common downstream targets. Analysis of different breast cancer subtypes showed distinct correlations of each cluster with clinical outcome. From the significantly up- and down-regulated proteins, we identified a number of markers of drug sensitivity and resistance. These data supports the role of TKs in regulating major aspects of cellular activity, but also reveals redundancy in signaling, explaining why kinase inhibitors alone often fail to achieve their clinical aims. The TK-SILACepedia provides a comprehensive resource for studying the global function of TKs in cancer.
authors.affiliation: From the ‡Department of Surgery and Cancer, Division of Cancer, Imperial College London, Hammersmith Hospital Campus, Du Cane Road, London, W12 ONN, UK;
authors.forename: Justin
authors.initials: J
authors.lastname: Stebbing
authors.affiliation: From the ‡Department of Surgery and Cancer, Division of Cancer, Imperial College London, Hammersmith Hospital Campus, Du Cane Road, London, W12 ONN, UK; h.zhang10@imperial.ac.uk.
authors.forename: Hua
authors.initials: H
authors.lastname: Zhang
authors.affiliation: From the ‡Department of Surgery and Cancer, Division of Cancer, Imperial College London, Hammersmith Hospital Campus, Du Cane Road, London, W12 ONN, UK;
authors.forename: Yichen
authors.initials: Y
authors.lastname: Xu
authors.affiliation: From the ‡Department of Surgery and Cancer, Division of Cancer, Imperial College London, Hammersmith Hospital Campus, Du Cane Road, London, W12 ONN, UK;
authors.forename: Arnhild
authors.initials: A
authors.lastname: Grothey
authors.affiliation: §Dundee Cell Products Ltd, James Lindsay Place, Dundee Technopole, Dundee, DD1 5JJ, UK.
authors.forename: Paul
authors.initials: P
authors.lastname: Ajuh
authors.affiliation: From the ‡Department of Surgery and Cancer, Division of Cancer, Imperial College London, Hammersmith Hospital Campus, Du Cane Road, London, W12 ONN, UK;
authors.forename: Nicos
authors.initials: N
authors.lastname: Angelopoulos
authors.affiliation: From the ‡Department of Surgery and Cancer, Division of Cancer, Imperial College London, Hammersmith Hospital Campus, Du Cane Road, London, W12 ONN, UK;
authors.forename: Georgios
authors.initials: G
authors.lastname: Giamas
mesh_terms: D000596:Amino Acids / Q000737:chemistry; D001943:Breast Neoplasms / Q000378:metabolism*; D018929:Cell Culture Techniques; D045744:Cell Line, Tumor; D004317:Doxorubicin / Q000494:pharmacology; D019008:Drug Resistance, Neoplasm; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D007553:Isotope Labeling / Q000379:methods*; D061986:MCF-7 Cells; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods*; D034622:RNA Interference; D015398:Signal Transduction
chemical_list: D000596:Amino Acids; D020543:Proteome; D004317:Doxorubicin; D011505:Protein-Tyrosine Kinases