Synergism between Ad-E2F1 and celecoxib results in increased rates of apoptotic cell death in breast cancer cells

Abstract

3181 Background: E2F1 is a transcription factor important in the regulation of cell cycle and apoptosis. Adenoviral mediated delivery of E2F1 (Ad-E2F1) has been shown to promote apoptosis in many cancer cells. Celecoxib, one of the selective cyclooxygenase 2 (COX-2) inhibitors, has also been shown to potentiate cancer cell apoptosis. We sought to determine whether the combination of Ad-E2F1 and celecoxib, targeting different cellular pathways, could result in synergism allowing for increased rates of apoptotic cell death in breast cancer cells in vitro. Methods: We utilized MCF7/Her18 cells (HER-2/neu-positive) and MDA-MB-436 (HER-2/neu-negative) breast cancer cells. We treated cells with Ad-E2F1 at a multiplicity of infection (MOI) of 1000 for MCF7/Her18 and an MOI of 500 for MDA-MB-436 cells. Celecoxib was added to the culture media to obtain concentrations of 30 uM for Her18 cells and 50 uM for 436 cells. We determined cell survival by trypan blue exclusion following treatment of cells with phosphate buffered saline (control), adenovirus with the luciferase reporter gene, Ad-E2F1, celecoxib, and the combination of Ad-E2F1 with celecoxib. Flow cytometry was performed to assess changes in cell cycle profile and to assess for apoptotic cell populations. Western blot analysis was performed to determine changes in protein levels. Results: The combination of Ad-E2F1 and celecoxib showed a marked reduction in cell survival over either treatment alone in both cell lines (p<0.05). In addition, there was a significant increase in apoptosis in both breast cancer cell lines with the combination therapy. On western blot, Akt, phosphorylated Akt, and beta-catenin levels were decreased in the combination group compared to controls. Conclusions: The combination therapy of Ad-E2F1 and celecoxib were synergistic in reducing cell survival in HER-2 positive and negative breast cancer cells. Decreased cell survival correlated with increased apoptosis and was associated with downregulation of Akt and beta-catenin. These data show that celecoxib and E2F1 cooperate to result in increased rates of cell death in breast cancer cells and may be a novel treatment option for breast cancer patients who fail standard therapy. No significant financial relationships to disclose.