title: Proteomic analysis of imatinib-resistant CML-T1 cells reveals calcium homeostasis as a potential therapeutic target.
abstract: Chronic myeloid leukemia (CML) therapy has markedly improved patient prognosis after introduction of imatinib mesylate for clinical use. However, a subset of patients develops resistance to imatinib and other tyrosine kinase inhibitors (TKIs), mainly due to point mutations in the region encoding the kinase domain of the fused BCR-ABL oncogene. To identify potential therapeutic targets in imatinib‑resistant CML cells, we derived imatinib-resistant CML-T1 human cell line clone (CML-T1/IR) by prolonged exposure to imatinib in growth media. Mutational analysis revealed that the Y235H mutation in BCR-ABL is probably the main cause of CML-T1/IR resistance to imatinib. To identify alternative therapeutic targets for selective elimination of imatinib-resistant cells, we compared the proteome profiles of CML-T1 and CML-T1/IR cells using 2-DE-MS. We identified eight differentially expressed proteins, with strongly upregulated Na+/H+ exchanger regulatory factor 1 (NHERF1) in the resistant cells, suggesting that this protein may influence cytosolic pH, Ca2+ concentration or signaling pathways such as Wnt in CML-T1/IR cells. We tested several compounds including drugs in clinical use that interfere with the aforementioned processes and tested their relative toxicity to CML-T1 and CML-T1/IR cells. Calcium channel blockers, calcium signaling antagonists and modulators of calcium homeostasis, namely thapsigargin, ionomycin, verapamil, carboxyamidotriazole and immunosuppressive drugs cyclosporine A and tacrolimus (FK-506) were selectively toxic to CML-T1/IR cells. The putative cellular targets of these compounds in CML-T1/IR cells are postulated in this study. We propose that Ca2+ homeostasis can be a potential therapeutic target in CML cells resistant to TKIs. We demonstrate that a proteomic approach may be used to characterize a TKI-resistant population of CML cells enabling future individualized treatment options for patients.
authors.affiliation: Institute of Hematology and Blood Transfusion, CZ-12820 Prague 2, Czech Republic.
authors.forename: O
authors.initials: O
authors.lastname: Toman
authors.affiliation: Institute of Hematology and Blood Transfusion, CZ-12820 Prague 2, Czech Republic.
authors.forename: T
authors.initials: T
authors.lastname: Kabickova
authors.affiliation: BIOCEV, First Faculty of Medicine, Charles University in Prague, CZ-25250 Vestec, Czech Republic.
authors.forename: O
authors.initials: O
authors.lastname: Vit
authors.affiliation: Department of Genetics and Microbiology, Faculty of Natural Sciences, Charles University in Prague, CZ-12843 Prague, Czech Republic.
authors.forename: R
authors.initials: R
authors.lastname: Fiser
authors.affiliation: Institute of Hematology and Blood Transfusion, CZ-12820 Prague 2, Czech Republic.
authors.forename: K Machova
authors.initials: KM
authors.lastname: Polakova
authors.affiliation: Institute of Hematology and Blood Transfusion, CZ-12820 Prague 2, Czech Republic.
authors.forename: J
authors.initials: J
authors.lastname: Zach
authors.affiliation: Institute of Hematology and Blood Transfusion, CZ-12820 Prague 2, Czech Republic.
authors.forename: J
authors.initials: J
authors.lastname: Linhartova
authors.affiliation: Institute of Hematology and Blood Transfusion, CZ-12820 Prague 2, Czech Republic.
authors.forename: D
authors.initials: D
authors.lastname: Vyoral
authors.affiliation: Institute of Hematology and Blood Transfusion, CZ-12820 Prague 2, Czech Republic.
authors.forename: J
authors.initials: J
authors.lastname: Petrak
mesh_terms: D002118:Calcium / Q000378:metabolism*; D045744:Cell Line, Tumor; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D016044:Fusion Proteins, bcr-abl / Q000378:metabolism; D006706:Homeostasis / Q000187:drug effects*; D006801:Humans; D000068877:Imatinib Mesylate / Q000494:pharmacology*; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000188:drug therapy* / Q000378:metabolism; D009154:Mutation / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D020543:Proteome / Q000378:metabolism*; D040901:Proteomics / Q000379:methods; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D047428:Protein Kinase Inhibitors; D020543:Proteome; D000068877:Imatinib Mesylate; D016044:Fusion Proteins, bcr-abl; D002118:Calcium