Gamabufotalin suppresses pancreatic cancer through redox-homeostasis disruption by G6PD downregulation

BACKGROUND: Pancreatic cancer is a leading cause of cancer-related deaths worldwide, highlighting the critical need for novel therapeutic strategies. Gamabufotalin (CS-6), a bufadienolide compound, has been shown to exert significant inhibitory effects on a variety of cancers. OBJECTIVE: This study investigates the potential therapeutic effects and underlying molecular mechanisms of CS-6 in pancreatic cancer. METHODS: The anti-cancer properties of CS-6 were assessed through both in vivo and in vitro experiments. Network pharmacology and untargeted metabolomics were employed to identify candidate molecular targets of CS-6. Molecular docking, molecular dynamics simulations, and CETSA were used to support potential target engagement between CS-6 and G6PD. G6PD expression was evaluated through public databases and immunohistochemistry. NADPH levels, ROS accumulation, apoptosis, and autophagy were measured using NADPH assays, western blot, flow cytometry, and transmission electron microscopy. Transcriptomic analysis and western blot were conducted to explore the downstream signaling pathways affected by G6PD downregulation. RESULTS: CS-6 inhibited pancreatic cancer cell growth, migration, and invasion, while promoting apoptosis. In vivo, CS-6 suppressed tumor growth and induced cell death. Elevated G6PD expression in pancreatic cancer was associated with poor prognosis. CS-6 treatment reduced NADPH levels, increased ROS accumulation, and was associated with autophagy induction, accompanied by decreased G6PD protein expression. Furthermore, CS-6 inhibited the PI3K/AKT signaling pathway, and G6PD knockdown recapitulated this effect. CONCLUSION: CS-6 treatment is associated with G6PD downregulation, redox-homeostasis disruption, and suppression of the PI3K/AKT signaling pathway, thereby contributing to autophagy-related cell death, apoptosis, and inhibition of pancreatic cancer progression. These findings highlight CS-6 as a promising therapeutic candidate for pancreatic cancer treatment, with G6PD as a potential molecular target.