title: Phosphoproteome Profiling Reveals Molecular Mechanisms of Growth-Factor-Mediated Kinase Inhibitor Resistance in EGFR-Overexpressing Cancer Cells.
abstract: Although substantial progress has been made regarding the use of molecularly targeted cancer therapies, resistance almost invariably develops and presents a major clinical challenge. The tumor microenvironment can rescue cancer cells from kinase inhibitors by growth-factor-mediated induction of pro-survival pathways. Here we show that epidermal growth factor receptor (EGFR) inhibition by Gefitinib is counteracted by growth factors, notably FGF2, and we assessed the global molecular consequences of this resistance at the proteome and phosphoproteome level in A431 cells. Tandem mass tag peptide labeling and quantitative mass spectrometry allowed the identification and quantification of 22 000 phosphopeptides and 8800 proteins in biological triplicates without missing values. The data show that FGF2 protects the cells from the antiproliferative effect of Gefitinib and largely prevents reprogramming of the proteome and phosphoproteome. Simultaneous EGFR/FGFR or EGFR/GSG2 (Haspin) inhibition overcomes this resistance, and the phosphoproteomic experiments further prioritized the RAS/MEK/ERK as well as the PI3K/mTOR axis for combination treatment. Consequently, the MEK inhibitor Trametinib prevented FGF2-mediated survival of EGFR inhibitor-resistant cells when used in combination with Gefitinib. Surprisingly, the PI3K/mTOR inhibitor Omipalisib reversed resistance mediated by all four growth factors tested, making it an interesting candidate for mitigating the effects of the tumor microenvironment.
authors: Chair for Proteomics and Bioanalytics, Technical University of Munich , 85354 Freising, Germany.|German Cancer Consortium (DKTK) , 69120 Heidelberg, Germany.|German Cancer Research Center (DKFZ) , 69120 Heidelberg, Germany.
authors: Heiner
authors: H
authors: Koch
authors: Chair for Proteomics and Bioanalytics, Technical University of Munich , 85354 Freising, Germany.
authors: Mathias
authors: M
authors: Wilhelm
authors: Chair for Proteomics and Bioanalytics, Technical University of Munich , 85354 Freising, Germany.|Center for Integrated Protein Science Munich (CIPSM) , 81377 Munich, Germany.
authors: Benjamin
authors: B
authors: Ruprecht
authors: Proteomics and Signal Transduction, Max-Planck-Institute of Biochemistry , 82152 Martinsried, Germany.
authors: Scarlet
authors: S
authors: Beck
authors: Chair for Proteomics and Bioanalytics, Technical University of Munich , 85354 Freising, Germany.|Department of Oncology, University of Oxford , OX3 7DQ Oxford, United Kingdom.
authors: Martin
authors: M
authors: Frejno
authors: Chair for Proteomics and Bioanalytics, Technical University of Munich , 85354 Freising, Germany.|German Cancer Consortium (DKTK) , 69120 Heidelberg, Germany.|German Cancer Research Center (DKFZ) , 69120 Heidelberg, Germany.
authors: Susan
authors: S
authors: Klaeger
authors: Chair for Proteomics and Bioanalytics, Technical University of Munich , 85354 Freising, Germany.|German Cancer Consortium (DKTK) , 69120 Heidelberg, Germany.|German Cancer Research Center (DKFZ) , 69120 Heidelberg, Germany.|Center for Integrated Protein Science Munich (CIPSM) , 81377 Munich, Germany.|Bavarian Biomolecular Mass Spectrometry Center, Technische Universität München , 85354 Freising, Germany.
authors: Bernhard
authors: B
authors: Kuster
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology; D045744:Cell Line, Tumor; D019008:Drug Resistance, Neoplasm / Q000187:drug effects*; D066246:ErbB Receptors / Q000037:antagonists & inhibitors / Q000235:genetics*; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology*; D000077156:Gefitinib; D006801:Humans; D009369:Neoplasms / Q000378:metabolism* / Q000473:pathology; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010748:Phosphopeptides / Q000032:analysis*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D020543:Proteome / Q000032:analysis*; D040901:Proteomics; D011728:Pyridones / Q000494:pharmacology / Q000627:therapeutic use; D011744:Pyrimidinones / Q000494:pharmacology / Q000627:therapeutic use; D011799:Quinazolines / Q000494:pharmacology
chemical_list: D000970:Antineoplastic Agents; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010748:Phosphopeptides; D047428:Protein Kinase Inhibitors; D020543:Proteome; D011728:Pyridones; D011744:Pyrimidinones; D011799:Quinazolines; D016222:Fibroblast Growth Factor 2; C560077:trametinib; C512478:EGFR protein, human; D066246:ErbB Receptors; D000077156:Gefitinib
keywords: Drug resistance; EGFR; kinase inhibitors; mass spectrometry; phosphoproteomics; quantitative proteomics; tumor microenvironment