Expression of t-DARPP mediates trastuzumab resistance in breast cancer cells

Clin Cancer Res. 2008 Jul 15;14(14):4564-71. doi: 10.1158/1078-0432.CCR-08-0121. Epub 2008 Jun 25.

Abstract

Purpose: We have investigated the role of t-DARPP in trastuzumab resistance in ERBB2-amplified and overexpressed breast cancer cell lines.

Experimental design: We have used the HR-5 and HR-6 trastuzumab-resistant cells that were established from tumors that recurred in the presence of trastuzumab therapy following xenografts of BT-474 cells in nude mice. In addition, SKBR-3 cells, engineered for stable expression of t-DARPP, and HCC-1569 cells, which have constitutive expression of t-DARPP and are de novo resistant to trastuzumab, were used.

Results: We reported > or =15-fold up-regulation of mRNA and protein levels of t-DARPP in HR-5 and HR-6 cells compared with their progenitor BT-474 trastuzumab-sensitive cells. The t-DARPP expression was not regulated by changes in its promoter DNA methylation levels. The SKBR-3 cells stably expressing t-DARPP developed resistance to trastuzumab compared with their parental cells and empty vector controls (P < 0.01). The trastuzumab-resistant cell lines showed a significant increase in pAKT (Ser(473)) and BCL2 protein levels. The small interfering RNA knockdown of t-DARPP in all trastuzumab-resistant cells led to a significant reduction in ERBB2, pAKT (Ser(473)), and BCL2 protein levels with a significant decrease in cell viability (P < or = 0.001) and an increase in cleaved caspase-3 levels, indicating the progression of these cells toward apoptosis. The t-DARPP protein was associated with both heat shock protein 90 and ERBB2 forming a potential protein complex. This association may play a role in regulating ERBB2 protein in trastuzumab-resistant cells.

Conclusion: We conclude that t-DARPP is a novel molecular target that can mediate the therapeutic resistance to trastuzumab in breast cancer cells.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Antibodies, Monoclonal / therapeutic use*
  • Antibodies, Monoclonal, Humanized
  • Antineoplastic Agents / therapeutic use*
  • Apoptosis / drug effects
  • Apoptosis / physiology
  • Blotting, Western
  • Breast Neoplasms / drug therapy
  • Breast Neoplasms / genetics
  • Breast Neoplasms / metabolism*
  • Cell Line, Tumor
  • Dopamine and cAMP-Regulated Phosphoprotein 32 / biosynthesis*
  • Drug Resistance, Neoplasm / physiology*
  • Female
  • Gene Expression
  • Humans
  • Immunoprecipitation
  • In Situ Nick-End Labeling
  • Mice
  • Protein Isoforms / biosynthesis
  • RNA, Small Interfering
  • Receptor, ErbB-2 / biosynthesis
  • Reverse Transcriptase Polymerase Chain Reaction
  • Trastuzumab
  • Xenograft Model Antitumor Assays

Substances

  • Antibodies, Monoclonal
  • Antibodies, Monoclonal, Humanized
  • Antineoplastic Agents
  • Dopamine and cAMP-Regulated Phosphoprotein 32
  • Protein Isoforms
  • RNA, Small Interfering
  • ERBB2 protein, human
  • Receptor, ErbB-2
  • Trastuzumab