Development of Herceptin resistance in breast cancer cells

Cytometry A. 2004 Feb;57(2):86-93. doi: 10.1002/cyto.a.10095.


Background: Herceptin, a humanized antibody to HER-2, is now utilized in the clinic for metastatic breast cancer treatment. The response rate for HER-2+ patients is only 30% and little is known as to mechanisms of resistance. The mechanism of Herceptin action is also unknown but has been related to cell cycle inhibition.

Methods: The effects of Herceptin and other antibody treatments were determined by cell counting and cell cycle analysis. HER-2 and p27 expression levels were analyzed by flow cytometry and levels of activated AKT were compared by Western blot analysis. Cellular HER-2 and p27 expression was measured by immunofluorescence.

Results: Herceptin treatment of BT-474 cells results in inhibition of cell growth and arrest in the G1 phase. The efficacy of growth arrest was not directly correlated to the binding affinity of antibodies to Her-2. Our laboratory has developed cell lines that are resistant to Herceptin treatment. In resistant cell lines, binding of antibodies is not hindered. However, Herceptin has completely lost the ability to inhibit cell proliferation. Yet, the mouse isotype 4D5 maintains significant inhibitory activity upon Herceptin-resistant clones.

Conclusions: Herceptin binds effectively to Her-2 on the cell surface of Herceptin-resistant cell lines and the level of Her-2 expression on the cell surface is not downregulated. Herceptin resistance is not due to downregulation of levels of AKT protein expression, although, phosphorylation of AKT is enhanced in resistant lines and could have a role in resistance. Resistance appears to correlate with the loss of nuclear expression of the cyclin-dependent kinase inhibitor, p27, as defined by immunofluorescence and flow cytometry studies and cdk-2 binding studies.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Antibodies, Monoclonal / metabolism
  • Antibodies, Monoclonal / pharmacology*
  • Antibodies, Monoclonal / therapeutic use
  • Antibodies, Monoclonal, Humanized
  • Antineoplastic Agents / metabolism
  • Antineoplastic Agents / pharmacology*
  • Antineoplastic Agents / therapeutic use
  • Breast Neoplasms / drug therapy*
  • Breast Neoplasms / metabolism
  • Cell Cycle Proteins / metabolism
  • Cell Line, Tumor
  • Cyclin-Dependent Kinase Inhibitor p27
  • Down-Regulation
  • Drug Resistance, Neoplasm*
  • ErbB Receptors / metabolism
  • Female
  • Flow Cytometry
  • Humans
  • Mice
  • Trastuzumab
  • Tumor Suppressor Proteins / metabolism


  • Antibodies, Monoclonal
  • Antibodies, Monoclonal, Humanized
  • Antineoplastic Agents
  • Cdkn1b protein, mouse
  • Cell Cycle Proteins
  • Tumor Suppressor Proteins
  • Cyclin-Dependent Kinase Inhibitor p27
  • ErbB Receptors
  • Trastuzumab