Abstract LB-80: Role of ER calcium in Bcl-2 resistance to dosage and duration-related cytotoxic efficacy of Taxol

Abstract

Abstract Background and Objectives: Calcium is one of the key regulators in apoptosis. Among various calcium sources, our previous study shows that the endoplasmic reticulum (ER), the main intracellular calcium storage site, provides a specific target for the chemotherapy agent Taxol and thus contributes to apoptosis. Although the expression of the anti-apoptotic protein Bcl-2 has been shown to confer resistance to Taxol, the exact mechanism of how it inhibits apoptosis and the role of ER calcium in this process remains unclear. This study investigated whether ER-associated calcium changes were generated and related to resistance by Bcl-2 to Taxol-induced apoptosis. Materials and methods: Fluo4-AM and Cameleon D1ER was used to monitor cytosolic and ER calcium dynamics by advanced time-lapsed imaging. Annexin V-FITC and propidium iodide were used to detect various phases of apoptosis induced by Taxol at different dosages from 10−12 to 10−5 M and at different durations from 5 min to 12 hours. Breast cancer cell lines MDA-MB-M468 and MDA-MB-231 were used and Bcl-2 transfectant cell lines were constructed by stable transfection. Results and Significance: Our previous data shows that Taxol (above 10−6 M) induces a rapid ER calcium release (peaking at 5 minutes) that promotes apoptosis. In this study, upon the expression of Bcl-2, basal ER calcium levels are decreased and Taxol-induced ER calcium release is inhibited, playing a major role in Bcl-2 resistance to Taxol. Depending upon dose and exposure time, long-term (hours) Taxol treatment partially overcame the Bcl-2 inhibition of ER calcium release by continually stimulating the ER calcium release, ultimately inducing significant, although smaller apoptosis in the presence of Bcl-2 expression. In addition to the effect on ER calcium, our data shows that the extracellular calcium pool is involved in Bcl-2 resistance through increased capacitative calcium entry to refill the ER calcium store, preventing ER calcium depletion. Taxol does not have a significant effect on the enhanced calcium influx from the extracellular medium. Taken together, out results indicate that the ER calcium store is the focus of the Taxol attack and plays a key role in Bcl-2 resistance to cytotoxic efficacy of Taxol in breast cancer cells. This finding provides insight to better understand the complex relationship between Bcl-2, calcium and Taxol towards improving breast cancer chemotherapies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-80. doi:1538-7445.AM2012-LB-80