Abstract P4-01-07: Endoplasmic Reticulum-Associated Calcium Regulates Resistance of the Anti-Apoptotic Protein Bcl-2 to Taxol-Induced Apoptosis in Breast Cancer Cells

Abstract

Abstract Background: The unique anti-apoptotic function of Bcl-2 results in cellular resistance to many chemotherapeutic agents used in solid tumor treatments. However, there is a controversy regarding the relationship between Bcl-2 resistance and the mechanism of action for Taxol, an anticancer agent commonly used in breast cancer treatment. Although previous studies showed both Bcl-2 and Taxol may have effects on calcium homeostasis independently, no studies have addressed whether calcium is the keystone in the relationship between Bcl-2 resistance and Taxol action. Objective: Since most internal calcium is stored in the endoplasmic reticulum (ER), this study focused on the role of ER calcium changes in Bcl-2-activated resistance to Taxol-induced apoptosis. The ER calcium channel IP3R and its relationship with Bcl-2 and Taxol were investigated in order to clarify the mechanism underlying calcium regulation. Methods and Materials: For this study, MDA-MB-468, a Bcl-2 negative breast carcinoma cell line, its Bcl-2 stable transfectant and MCF-7, a Bcl-2 positive breast carcinoma cell line were used. The dynamic calcium changes induced by Taxol in these cells were determined by measuring free cytosolic calcium fluctuations using Fluo4-AM and ER calcium changes by the calcium cameleon, D1ER. Taxol-induced apoptosis was measured using the Annexin V-FITC assay. The calcium interfering agents BAPTA-AM and 2-APB were applied to evaluate whether Taxol-induced calcium changes were related to this apoptotic event. Western blot was used to detect changes induced by Taxol treatment in Bcl-2 and IP3R protein levels. Results: Taxol induced a rapid ER calcium release followed by a gradual ER calcium depletion in both Bcl-2 positive and negative cell lines, indicating that ER calcium store provides a direct target for Taxol, independently of Bcl-2 expression. Interfering with these calcium changes inhibited the Taxol-induced apoptosis, suggesting ER calcium release promotes Taxol-induced apoptosis. Our results also showed that Bcl-2 inhibited the basal ER calcium release. Taxol stimulated ER calcium release and thus attenuated Bcl-2-initiated resistance to apoptosis. Relevance: This research addressed the question of whether ER calcium changes play a critical or marginal role in mediating resistance of Bcl-2 to Taxol-induced apoptosis. Elucidating the role of calcium in this process will not only help to clarify the mechanism of drug resistance associated with Bcl-2, but also aid in more effective application of Taxol in breast cancer treatment. Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P4-01-07.