Biochemical and Biophysical Research Communications
Inhibition of vacuolar H+ ATPase enhances sensitivity to tamoxifen via up-regulation of CHOP in breast cancer cells
Introduction
Breast cancer is one of the most frequent cancers and is the leading cause of death in women worldwide [1], [2]. Hormone therapy is an adjuvant therapy for women whose breast cancer expresses hormone receptors [3], [4]. Estrogen is known to promote the growth of many breast cancers through its binding to estrogen receptor. The selective estrogen receptor antagonist tamoxifen has improved survival in breast cancer patients. Despite improvements in treatment, resistance to tamoxifen is still a major obstacle [5]. A combination therapy of tamoxifen with other drugs that cause synergistic anti-tumor effects may therefore be an attractive option in breast cancer therapy.
Vacuolar H+ ATPase (vATPase), a multi-subunit enzyme, is an ATP-driven proton pump that translocates protons from the cytoplasm into intracellular compartments and across the plasma membrane [6], [7]. vATPase consists of an ATP-hydrolyzing cytoplasmic V1 complex and a proton-translocating, membrane-bound V0 complex. By regulating cellular pH balance, vATPase is implicated in various cellular functions including endocytosis and protease activation [8], [9]. Cancer cells are forced to exist in a hypoxic and acidic environment, leading to increased glycolysis [10], [11]. The expression of vATPase is considered to be a well-designed compensatory mechanism that allows cancer cells to survive and proliferate [12]. Therefore, vATPase inhibitors could be promising cancer treatments. Bafilomycin A1 and concanamycin A, selective vATPase inhibitors targeting subunit c in V0 (ATP6V0C), have been suggested as potential anticancer agents [13], [14].
In the present study, we demonstrated that inhibition of vATPase activity by pharmacologic drugs enhanced the sensitivity to tamoxifen by up-regulating CHOP pathway in breast cancer cells. The combinatory effects of vATPase inhibitor(s) and tamoxifen were also verified in ERα/HER2-overexpressing breast cancer cells. Our findings suggest that inhibiting vATPase may provide a novel approach for the treatment of tamoxifen-insensitive breast cancer cells.
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Cell culture and reagents
MCF7 and BT474 breast cancer cell lines were obtained from the American Type Culture Collection (Manassas, VA, USA). HER2-overexpressing MCF7 breast cancer cells were a generous gift from Dr. Incheol Shin (HanYang University, Seoul, Republic of Korea). MCF7 and HER2-overexpressing MCF7 human breast cancer cells were maintained in DMEM (Invitrogen, Carlsbad, CA, USA), and BT474 cells were maintained in RPMI1640 (Invitrogen). Tamoxifen, bafilomycin A1, and concanamycin A were purchased from
vATPase inhibitors enhanced the cytotoxic effect of tamoxifen in MCF7 cells
We first investigated the effect of tamoxifen on MCF7 cell viability. MCF7 breast cancer cells were incubated for 24 h with various concentrations of tamoxifen, and cell viability was determined by MTT assay. Cell viability was reduced to less than 20% (data not shown), despite the presence of a high concentration (10 μM) of tamoxifen [18]. Although cell death in MCF7 cells undergoing combined glucose deprivation and sorafenib treatment has been reported [15], no cell death occurred in MCF7 cells
Discussion
Tamoxifen has become the most widely used drug in managing breast cancer [5], [21]. However, as with many cancer treatments, resistance to tamoxifen is a significant issue, and up to 40% of early stage breast cancer patients who receive tamoxifen as an adjuvant therapy eventually relapse with tamoxifen-resistant disease [21].
General mechanisms have been proposed to explain the development of resistance, including continued estrogen receptor signaling in the presence of ER antagonists or the
Acknowledgments
This research has been supported grants from the Radiological Translational Research Program (50451-2013), the National Nuclear R&D Program and the Basic Science Research Program (A111770) funded by the Ministry of Education, Science and Technology, and the Radiation Bio-Resource Research Program of the Korea Institute of Radiological and Medical Sciences (No. 740802) in the Republic of Korea.
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