Microarray analysis of tamoxifen resistance in breast cancer.

Abstract

Tamoxifen remains a widely used hormone therapy for pre and postmenopausal women with hormone receptor positive breast cancer in both adjuvant and metastatic disease settings. Resistance to this well tolerated and cost-effective drug limits its use. Only an improved understanding of the mechanisms of tamoxifen resistance will provide the basis for overcoming this phenomenon. Expression profiles from tamoxifen-resistant and sensitive MCF7 derived breast cancer cell lines were prepared, using Affymetrix HG_U133A cDNA microarrays. The data generated was analysed to identify novel pathways and genes associated with tamoxifen resistance or sensitivity. Selected genes, whose expression correlates with response to tamoxifen, were validated using RT-PCR in cell lines and following this, in situ hybridisation and immunohistochemistry on cell lines. Functional analyses of these genes were carried out: genes that were down-regulated in tamoxifen resistant MCF7 cells (HRASLS3, CTSD, CAXII) were selectively knocked down using RNA interference. Cell lines stably over-expressing genes upregulated in the tamoxifen resistant MCF7s (ATP1B1, SOCS2, NR4A2) were selected. These manipulated cells were subsequently tested for their response to anti-oestrogen treatment. Another major marker in breast cancer is the ERBB2 proto-oncogene; overexpressed in 20% of breast carcinomas, it is associated with poor prognosis and hormone resistance. The transcriptional deregulation of ERBB2 in breast cancer may in part be mediated by the transcription factors AP-2 and . Previous studies have shown that ERBB2 expression is repressed by oestrogen activated oestrogen receptor and that AP-2 binding sites within the ERBB2 promoter and the intronic enhancer are required for this oestrogenic repression. Using RNA interference, AP-2 and were successfully knocked down in breast cancer cell lines MCF7, T47D and ZR75-1. These have been used to investigate the effect of AP-2 loss on ERBB2 expression in hormonally manipulated cells.