Abstract
Background Protoapigenone, as a flavonoid compound with a specific nonaromatic B-ring, exhibits extraordinary antitumor activities against a broad spectrum of human cancer cells. Here we developed a novel protoapigenone analog RY10-4, which induces the apoptosis of various tumor cells, especially for breast cancer cells, but the underlying mechanism involved in the apoptotic process remains unclear. Methods MTT assay, colony-formation assay and flow cytometry were applied to evaluate the proliferation and apoptosis of breast cancer cells. Cytoplasmic calcium ([Ca2+]c) and mitochondrial calcium ([Ca2+]m) of the breast cancer cells were measured by the Fluo-2 and Rhod-2 probes, respectively. The mitochondrial reactive oxygen species (mtROS), membrane potential (ΔΨm) and permeability transition pore (mPTP) were analyzed by MitoSOX, JC-1 probes and Calcein/AM, respectively. Furthermore, Western bolt assay was adopted for the exploration of the mitochondrial apoptosis pathway. Besides, the xenograft assay was performed to investigate the role of RY10-4 in breast cancer cells in vivo. Results Obviously, RY10-4 could effectively suppress the proliferation and induce the apoptosis of breast cancer cells. Furthermore, the [Ca2+]c and [Ca2+]m of MDA-MB-231 cells were up-regulated after the treatment of RY10-4, resulting in the mtROS accumulation, ΔΨm depolarization and mPTP opening. And finally, the mitochondrial apoptosis was activated by the release of cytochrome C. Interestingly, the inhibition of mitochondrial calcium uniporter (MCU) with Ru360 attenuated the overload of [Ca2+]m and blocked the apoptosis of MDA-MB-231 cells induced by RY10-4, which was also consistent with the in vivo results. Conclusions From the results we concluded that RY10-4 could induce apoptosis of breast cancer cells by elevating [Ca2+]m through MCU. Our work contributed previously unknown insights into the mechanisms involving in the clinical efficacy of RY10-4 on breast cancer cells, which also advanced calcium homeostasis as a potential target for chemotherapeutic drugs.
The Fanconi anemia pathway sensitizes to DNA alkylating agents by inducing JNK-p53-dependent mitochondrial apoptosis in breast cancer cells