Mitochondrial Ca2+ removal amplifies TRAIL cytotoxicity toward apoptosis-resistant tumor cells via promotion of multiple cell death modalities

Ca2+ has emerged as a new target for cancer treatment since tumor-specific traits in Ca2+ dynamics contributes to tumorigenesis, malignant phenotypes, drug resistance, and survival in different tumor types. However, Ca2+ has a dual (pro-death and pro-survival) function in tumor cells depending on the experimental conditions. Therefore, it is necessary to minimize the onset of the pro-survival Ca2+ signals caused by the therapy. For this purpose, a better understanding of pro-survival Ca2+ pathways in cancer cells is critical. Here we report that Ca2+ protects malignant melanoma (MM) and osteosarcoma (OS) cells from tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) cytotoxicity. Simultaneous measurements using the site-specific Ca2+ probes showed that acute TRAIL treatment rapidly and dose-dependently increased the cytosolic Ca2+ concentration ([Ca2+]cyt) and mitochondrial Ca2+ concentration ([Ca2+]mit) Pharmacological analyses revealed that the [Ca2+]mit remodeling was under control of mitochondrial Ca2+ uniporter (MCU), mitochondrial permeability transition pore (MPTP), and a Ca2+ transport pathway sensitive to capsazepine and AMG9810. Ca2+ chelators and the MCU inhibitor ruthenium 360, an MPTP opener atractyloside, capsazepine, and AMG9810 all decreased [Ca2+]mit and sensitized these tumor cells to TRAIL cytotoxicity. The Ca2+ modulation enhanced both apoptotic and non-apoptotic cell death. Although the [Ca2+]mit reduction potentiated TRAIL-induced caspase-3/7 activation and cell membrane damage within 24 h, this potentiation of cell death became pronounced at 72 h, and not blocked by caspase inhibition. Our findings suggest that in MM and OS cells mitochondrial Ca2+ removal can promote apoptosis and non-apoptotic cell death induction by TRAIL. Therefore, mitochondrial Ca2+ removal can be exploited to overcome the resistance of these cancers to TRAIL.