title: Abrogating phosphorylation of eIF4B is required for EGFR and mTOR inhibitor synergy in triple-negative breast cancer.
abstract: Triple-negative breast cancer (TNBC) patients suffer from a highly malignant and aggressive disease. They have a high rate of relapse and often develop resistance to standard chemotherapy. Many TNBCs have elevated epidermal growth factor receptor (EGFR) but are resistant to EGFR inhibitors as monotherapy. In this study, we sought to find a combination therapy that could sensitize TNBC to EGFR inhibitors. Phospho-mass spectrometry was performed on the TNBC cell line, BT20, treated with 0.5 μM gefitinib. Immunoblotting measured protein levels and phosphorylation. Colony formation and growth assays analyzed the treatment on cell proliferation, while MTT assays determined the synergistic effect of inhibitor combination. A Dual-Luciferase reporter gene plasmid measured translation. All statistical analysis was done on CalucuSyn and GraphPad Prism using ANOVAs. Phospho-proteomics identified the mTOR pathway to be of interest in EGFR inhibitor resistance. In our studies, combining gefitinib and temsirolimus decreased cell growth and survival in a synergistic manner. Our data identified eIF4B, as a potentially key fragile point in EGFR and mTOR inhibitor synergy. Decreased eIF4B phosphorylation correlated with drops in growth, viability, clonogenic survival, and cap-dependent translation. Taken together, these data suggest EGFR and mTOR inhibitors abrogate growth, viability, and survival via disruption of eIF4B phosphorylation leading to decreased translation in TNBC cell lines. Further, including an mTOR inhibitor along with an EGFR inhibitor in TNBC with increased EGFR expression should be further explored. Additionally, translational regulation may play an important role in regulating EGFR and mTOR inhibitor synergy and warrant further investigation.
authors.affiliation: Departments of Oncology, Wayne State University, 4100 John R. St, Detroit, MI, 48201, USA.
authors.forename: Julie M
authors.initials: JM
authors.lastname: Madden
authors.forename: Kelly L
authors.initials: KL
authors.lastname: Mueller
authors.forename: Aliccia
authors.initials: A
authors.lastname: Bollig-Fischer
authors.forename: Paul
authors.initials: P
authors.lastname: Stemmer
authors.forename: Raymond R
authors.initials: RR
authors.lastname: Mattingly
authors.forename: Julie L
authors.initials: JL
authors.lastname: Boerner
mesh_terms: D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology*; D002460:Cell Line; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects / Q000235:genetics; D004357:Drug Synergism; D066246:ErbB Receptors / Q000037:antagonists & inhibitors* / Q000378:metabolism; D039642:Eukaryotic Initiation Factors / Q000378:metabolism*; D005260:Female; D000077156:Gefitinib; D057809:HEK293 Cells; D006801:Humans; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000008:administration & dosage / Q000494:pharmacology; D011799:Quinazolines / Q000008:administration & dosage / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D020123:Sirolimus / Q000008:administration & dosage / Q000031:analogs & derivatives / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D064726:Triple Negative Breast Neoplasms / Q000188:drug therapy* / Q000378:metabolism* / Q000473:pathology
chemical_list: D039642:Eukaryotic Initiation Factors; D047428:Protein Kinase Inhibitors; D011799:Quinazolines; C031799:eIF-4B; C401859:temsirolimus; C546842:MTOR protein, human; C512478:EGFR protein, human; D066246:ErbB Receptors; D058570:TOR Serine-Threonine Kinases; D000077156:Gefitinib; D020123:Sirolimus