Inhibition of NAMPT markedly enhances plasma-activated medium-induced cell death in human breast cancer MDA-MB-231 cells

Abstract

Plasma-activated medium (PAM), which is prepared by non-thermal atmospheric pressure plasma (NTP) irradiation of cell-free medium, has been shown to exhibit tumor-specific cytotoxicity. Since PAM contains reactive oxygen species (ROS) and reactive nitrogen species (RNS), its anticancer effects are considered to be responsible for oxidative stress induced by these reactive molecules. We previously reported that PAM-induced cell death is closely related to energy failure associated with a decrease in intracellular nicotinamide adenine dinucleotide (NAD+) and ATP levels. Nicotinamide phosphoribosyltransferase (NAMPT), which is a rate-limiting enzyme for NAD+ synthesis in the salvage pathway, was shown to be overexpressed in many types of cancer cells. The NAMPT inhibitor FK866 significantly depletes NAD+ and subsequently suppresses cancer cell proliferation. In this study, we examined the effects of FK866 on PAM-induced cytotoxicity using human breast cancer MDA-MB-231 cells. FK866 dose-dependently enhanced PAM-induced cell death in MDA-MB-231 cells. The combination of PAM and FK866 markedly induced intracellular NAD+ and ATP depletion. Knockdown of NAMPT by siRNA increased the cytotoxicity of PAM. The addition of NAD+ mitigated PAM-induced cell death. In addition, cotreatment with PAM and FK866 augmented ROS production and the decrease in intracellular reduced glutathione (GSH) compared to treatment with PAM alone. FK866 had little effect on PAM-induced mitochondrial dysfunction. Furthermore, the combination of PAM and FK866 decreased the level of NADPH, which is required for GSH metabolism, compared with PAM alone. Taken together, we conclude that cotreatment with NAMPT inhibitors is beneficial for anticancer therapy using PAM.