t10c12 Conjugated Linoleic Acid Suppresses HER2 Protein and Enhances Apoptosis in SKBr3 Breast Cancer Cells: Possible Role of COX2

Background: HER2-targeted therapy with the monoclonal antibody trastuzumab (Herceptin (R)) has improved disease-free survival for women diagnosed with HER2-positive breast cancers; however, treatment resistance and disease progression are not uncommon. Current data suggest that resistance to treatment in HER2 cancers may be a consequence of NF-kappa B overexpression and increased COX2-derived prostaglandin E2 (PGE(2)). Conjugated linoleic acid (CLA) has been shown to have anti-tumor properties and to inhibit NF-kappa B activity and COX2. Methods: In this study, HER2-overexpressing SKBr3 breast cancer cells were treated with t10c12 CLA. Protein expression of the HER2 receptor, nuclear NF-kappa B p65, and total and phosphorylated I kappa B were examined by western blot and immunofluorescence. PGE2 levels were determined by ELISA. Proliferation was measured by metabolism of 3-(4, 5-Dimethylthiazol- 2-yl)-2, 5-diphenyltetrazolium bromide (MTT), and apoptosis was measured by FITC-conjugated Annexin V staining and flow cytometry. Results/Conclusions: We observed a significant decrease in HER2 protein expression on western blot following treatment with 40 and 80 mu M t10c12 CLA (p < 0.01 and 0.001, respectively) and loss of HER2 protein in cells using immunoflourescence that was most pronounced at 80 mM. Protein levels of nuclear NF-kappa B p65 were also significantly reduced at the 80 mM dose. This was accompanied by a significant decrease in PGE2 levels (p = 0.05). Pretreatment with t10c12 CLA significantly enhanced TNF alpha-induced apoptosis and the anti-proliferative action of trastuzumab (p = 0.05 and 0.001, respectively). These data add to previous reports of an anti-tumor effect of t10c12 CLA and suggest an effect on the HER2 oncogene that may be through CLA mediated downregulation of COX2-derived PGE(2).