Growth and molecular interactions of the anti-EGFR antibody cetuximab and the DNA cross-linking agent cisplatin in gefitinib-resistant MDA-MB-468 cells: new prospects in the treatment of triple-negative/basal-like breast cancer

Int J Oncol. 2008 Dec;33(6):1165-76.


Three prominent hallmarks of triple-negative/basal-like breast carcinomas, a subtype of breast cancer gene phenotype associated with poor relapse-free and overall survival, are overexpression of the epidermal growth factor receptor (EGFR), hyperactivation of the MEK/ERK transduction pathway and high sensitivity to DNA-damaging agents. The cytotoxic interaction between EGFR inhibitors (monoclonal antibodies such as cetuximab and small molecule tyrosine kinase inhibitors such as gefitinib) and DNA cross-linking agents (e.g. platinum derivatives) might represent a promising combination for the treatment of triple-negative/basal-like breast tumors that are dependent upon EGFR/MEK/ERK signaling. We evaluated the growth and molecular interactions of the anti-EGFR antibody cetuximab (erbitux) and the DNA cross-linking agent cisplatin (cis-diammedichloroplatinum; CDDP) in the gefitinib-resistant MDA-MB-468 breast cancer cell line, an in vitro model system that shows many of the recurrent basal-like molecular abnormalities including ER-PR-HER2-negative status, TP53 deficiency, EGFR overexpression, PTEN loss and constitutive activation of the MEK/ERK pathway. Unlike other basal-like breast cancer models, MDA-MB-468 cells do not carry mutations of the key DNA repair gene BRCA1. Concurrent treatment with sub-optimal doses of cetuximab significantly enhanced CDDP-induced apoptotic cell death. However, an isobologram-based mathematical assessment of the nature of the interaction revealed a loss of synergism when employing a high-dose of cetuximab. Since BRCA1 depletion has been found to decrease DNA damage repair and cell survival in MDA-MB-468 cells when treated with DNA-damaging drugs, we employed ELISA-based quantitative analyses to measure BRCA1 protein levels in CDDP+/- cetuximab-treated cells. Cetuximab as single agent was as efficient as CDDP at increasing BRCA1 protein expression. Interestingly, cetuximab co-exposure significantly antagonized the ability of CDDP to up-regulate BRCA1 expression. Low-scale phosphor-proteomic approaches [i.e. phospho-receptor tyrosine kinase (RTK) and phospho-mitogen-activated protein kinases (MAPKs) Array Proteome Profiler capable of simultaneously identifying the relative levels of phosphorylation of 42 different RTKs and 23 different MAPKs and other serine/threonine kinases, respectively] revealed the ability of Cetuximab, as single agent, to paradoxically induce hyper-phosphorylation of EGFR while concomitantly deactivating p42/44 (ERK1/ERK2) MAPK. Unexpectedly, ELISA-based quantitative analyses of EGFR protein content demonstrated that simultaneous exposure to cetuximab and optimal doses of CDDP completely depleted EGFR protein in MDA-MB-468 cells. Although these findings preclinically support, at least in part, ongoing clinical trials for 'triple-negative/basal-like' metastatic breast cancer patients who are receiving either cetuximab alone versus cetuximab plus carboplatin (, the unexpected ability of CDDP to promote a complete depletion of the cetuximab target EGFR further suggests that treatment schedules, cetuximab/CDDP doses and BRCA1 status should be carefully considered when combining anti-EGFR antibodies and platinum derivatives in triple-negative/basal-like breast carcinomas.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antibodies, Monoclonal / administration & dosage
  • Antibodies, Monoclonal, Humanized
  • Antineoplastic Combined Chemotherapy Protocols / pharmacology*
  • Apoptosis / drug effects*
  • BRCA1 Protein / metabolism
  • Breast Neoplasms / enzymology
  • Breast Neoplasms / metabolism
  • Breast Neoplasms / pathology*
  • Cell Line, Tumor
  • Cell Proliferation / drug effects*
  • Cetuximab
  • Cisplatin / administration & dosage
  • Cross-Linking Reagents / administration & dosage
  • Dose-Response Relationship, Drug
  • Drug Resistance, Neoplasm*
  • Drug Synergism
  • ErbB Receptors / antagonists & inhibitors*
  • ErbB Receptors / metabolism
  • Female
  • Gefitinib
  • Humans
  • Mitogen-Activated Protein Kinase Kinases / metabolism
  • PTEN Phosphohydrolase / deficiency
  • Phosphorylation
  • Protein Kinase Inhibitors / administration & dosage
  • Quinazolines / pharmacology*
  • Receptor, ErbB-2 / deficiency
  • Receptors, Estrogen / deficiency
  • Receptors, Progesterone / deficiency
  • Tumor Suppressor Protein p53 / deficiency
  • Up-Regulation


  • Antibodies, Monoclonal
  • Antibodies, Monoclonal, Humanized
  • BRCA1 Protein
  • BRCA1 protein, human
  • Cross-Linking Reagents
  • Protein Kinase Inhibitors
  • Quinazolines
  • Receptors, Estrogen
  • Receptors, Progesterone
  • TP53 protein, human
  • Tumor Suppressor Protein p53
  • EGFR protein, human
  • ERBB2 protein, human
  • ErbB Receptors
  • Receptor, ErbB-2
  • Mitogen-Activated Protein Kinase Kinases
  • PTEN Phosphohydrolase
  • PTEN protein, human
  • Cetuximab
  • Cisplatin
  • Gefitinib

Associated data