Epigenetic therapy can rescue trastuzumab resistance in HER2 over-expressing breast cancers.
Authors: Amanika Kumar, Murli Narayan, Vince Schulz, David Tuck, Lyndsay Harris
Journal: Cancer Research (2008)

AACR Annual Meeting-- Apr 12-16, 2008; San Diego, CA

2623 

A number of HER2 over-expressing breast cancers are resistant to trastuzumab, a monoclonal antibody targeting HER2. Our lab has generated cell lines resistant to trastuzumab (BT474\_R and SkBr3\_R) by serial passage through trastuzumab. We have shown that resistant lines and resistant primary tumor samples show an overall decrease of HER2 amplicon gene expression. It is known that histone modifications play a role in regulating gene expression. Histone deacetylase inhibitors (HDACi) have the ability to alter gene expression, and their role in novel combination chemotherapy to treat breast cancers is being explored in the clinic currently. We chose to explore the effect of Trichostatin A, an HDACi, on HER2 amplicon gene expression and response to trastuzumab in HER2 overexpressing cell lines.    Five commercially available breast cancer cell lines, BT474, SkBr3, MCF7, MD361, and UACC812, and two in vitro generated trastuzumab-resistant breast cancer cells lines (BT474\_R and SkBr3\_R) were used. Cells were treated with TSA and/or Herceptin and examined for cell viability, morphology, and apoptosis. HER2 amplicon gene expression was examined before and after 1, 4, and 24 hours treatment with 1 uM TSA by qRT-PCR. Acid extraction of histones was performed to confirm TSA induced changes in histone acetylation.    All 8 cell lines were sensitive to TSA (IC50 range 51 nM to 330 nM) as demonstrated by cytotoxicity assays. As expected, BT474 and SkBr3 showed sensitivity to trastuzumab while de novo trastuzumab resistant cell lines (MD361, UACC812) and previously generated resistant cell lines showed little or no sensitivity to trastuzumab. In trastuzumab sensitive cell lines, the combination of TSA and trastuzumab led to decreased cell viability, greater than expected with either drug alone. In de novo resistant and secondarily resistant cell lines, TSA treatment enhanced trastuzumab induced growth inhibition. Interestingly, TSA treatment led to a 2-25 fold induction of genes in the HER2 amplicon examined including HER2, PP1RIB1 (DARPP32), GRB7, C17orF37, and STARD3 in both parental and in vitro generated resistant lines. In vitro acquired resistant lines had greater amplicon induction by TSA than parental controls. The induction of HER2 amplicon genes in resistant lines by TSA results in a HER2 amplicon gene expression pattern similar to the gene expression pattern of de novo trastuzumab sensitive cell lines.    HDACi are powerful chemotherapeutic agents that will likely play a role in therapy regimens in the future. We have shown that HER2 amplicon gene expression is altered by TSA treatment and that resistant cell lines show increased induction of HER2 amplicon as compared to trastuzumab sensitive cell lines. We have also shown that TSA changes breast cancer cell sensitivity to trastuzumab. Further exploration into the epigenetic mechanisms of trastuzumab resistance and into the application of novel HDACi is warranted.