Abstract C1: HDAC inhibitors trigger the autophagic switch from prosurvival to prodeath in tamoxifen-treated breast cancer cells

Abstract

Abstract Background: Hormone therapy remains the treatment of choice for patients with estrogen receptor-positive breast cancer. The majority of patients with an initial response to tamoxifen or aromatase inhibitors will develop resistance, and many patients present with tumors that are a priori resistant. One of the known survival strategies of breast cancer cells treated with hormone therapy is the induction of autophagy. Autophagy is a process by which cellular components are catabolized in autophagic lysosomes, enabling the clearance of damaged organelles and the recycling of nutrients during periods of starvation. Autophagy has been reported in response to treatment with both aromatase inhibitors and tamoxifen, in part by upregulating expression of the essential autophagy protein beclin-1. Inhibition of autophagy in breast cancer cells, increases the cytotoxicity of tamoxifen, suggesting that autophagy in these cells is oncogenic and potentially a contributor to resistance. We have shown that HDAC inhibitors potentiate the cytotoxicity of tamoxifen in breast cancer cells. These results raise the possibility that HDAC inhibitors achieve synergy by inhibiting autophagy. Methods: We sought to determine how HDAC inhibition impacts tamoxifen-induced autophagy in breast cancer cell line models. Results: Using several HDAC inhibitors, we show a synergistic increase in apoptosis and cell death with tamoxifen. The addition of an HDAC inhibitor to tamoxifen increased autophagy, in both a time and dose dependent manner. Increased autophagy required functional estrogen-mediated signaling, as depletion of ER by siRNA or treatment with fulvestrant did not result in increased autophagy as measured by LC3 levels. The inhibition of autophagy in tamoxifen-treated cells by 3-methyladenine circumvented autophagy and induced apoptosis supporting prior reports that autophagy acts as a survival mechanism in these cells. When combined with tamoxifen, HDAC inhibitors increased the expression of LC3 and Beclin1, suggesting enhanced induction of autophagy. Furthermore, LC3 and cleaved PARP were sequentially expressed in a dose and time dependent manner, with LC3 increasing first followed by PARP cleavage. In these cells, annexin-V staining further supports apoptotic rather than autophagic cell death. Together, this suggests cells transition from autophagic cell preservation to apoptotic cell death. Conclusion: An autophagic survival response in breast cancer cells has been described after nutrient starvation, tamoxifen treatment as well as following exposure to DNA damaging agents. Our data further suggests that with the synergistic induction of beclin-1 by HDAC inhibition and tamoxifen, the excess of autophagic lysosomes can no longer sustain self-preservation and triggers elimination of cells by apoptotic cell death in a fatal switch. Thus, combining tamoxifen with an HDAC inhibitor may represent a new therapeutic approach to overcome hormone therapy resistance. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):C1.