Breast cancer oestrogen independence mediated by BCAR1 or BCAR3 genes is transmitted through mechanisms distinct from the oestrogen receptor signalling pathway or the epidermal growth factor receptor signalling pathway

Abstract

Introduction: Tamoxifen is effective for endocrine treatment of oestrogen receptor-positive breast cancers but ultimately fails due to the development of resistance. A functional screen in human breast cancer cells identified two BCAR genes causing oestrogen-independent proliferation. The BCAR1 and BCAR3 genes both encode components of intracellular signal transduction, but their direct effect on breast cancer cell proliferation is not known. The aim of this study was to investigate the growth control mediated by these BCAR genes by gene expression profiling.

Methods: We have measured the expression changes induced by overexpression of the BCAR1 or BCAR3 gene in ZR-75-1 cells and have made direct comparisons with the expression changes after cell stimulation with oestrogen or epidermal growth factor (EGF). A comparison with published gene expression data of cell models and breast tumours is made.

Results: Relatively few changes in gene expression were detected in the BCAR-transfected cells, in comparison with the extensive and distinct differences in gene expression induced by oestrogen or EGF. Both BCAR1 and BCAR3 regulate discrete sets of genes in these ZR-75-1-derived cells, indicating that the proliferation signalling proceeds along distinct pathways. Oestrogen-regulated genes in our cell model showed general concordance with reported data of cell models and gene expression association with oestrogen receptor status of breast tumours.

Conclusions: The direct comparison of the expression profiles of BCAR transfectants and oestrogen or EGF-stimulated cells strongly suggests that anti-oestrogen-resistant cell proliferation is not caused by alternative activation of the oestrogen receptor or by the epidermal growth factor receptor signalling pathway.

Figure 1

Expression changes of selected genes in ZR-75-1-derived cell lines, and their relation to reported clinical phenotypes. Average gene expression ratios (log₁₀) in ZR-75-1 cells stimulated with oestrogen for 6 hours (E6hr) or continuously (Econt), of ZR/EGFR cells stimulated with epidermal growth factor for 7 days (EGF), of BCAR1-transfected or BCAR3-transfected ZR-75-1 cells versus unstimulated ZR-75-1 cells are indicated. A selection of 234 sequences was made from the 1006 sequences showing at least once a P value of 0.01 or less and exhibiting at least once a |DE| of more than 1.60. Individual data points of combined swapped and of virtual experiments were only included when either one of the following criteria was met: P < 0.051 or log(error) < 0.1761 or |log(ratio)| > 1.5 × log(error). This procedure eliminates most of the unreliable data points. Genes were hierarchically clustered by using Spotfire. A colour picture was made with TreeView [44]. Increased expression is shown in red and decreased expression is shown in green. Black represents no change and white indicates missing data. In addition, the reported inducing (red) or reducing (green) effects of oestrogen stimulation in cell line models [35–37,39,40] and the correlation (red, positive; green, negative) of individual genes with breast cancer oestrogen receptor (ER) status, BRCA mutation status (BRCA) and prognosis of disease recurrence is indicated [28,41]. Genes investigated with quantitative reverse transcriptase-mediated polymerase chain reaction have been marked with an asterisk. The complete list of 1006 genes with attached information is presented in Additional file 3.