Aloperine Induces Ferroptosis of Colorectal Cancer Cells via the Nrf2 Pathway.

OBJECTIVE: This study investigated the effects of ALO on apoptosis and ferroptosis in colorectal cancer HCT116 and SW480 cells and the underlying mechanisms. METHODS: Cells were treated with varying ALO concentrations. CCK-8 assay assessed proliferation. Flow cytometry detected apoptosis. Lipid peroxidation was measured by BODIPY 581/591 C11 dye oxidation and TBA method. GSH content was determined by DTNB method. ROS and intracellular iron levels were assessed using fluorescent probes and iron assays. Molecular docking analyzed ALO-Nrf2 binding. Immunofluorescence detected Nrf2 expression. Western blot quantified apoptosis-related proteins (Bax, Bcl-2) and ferroptosis-related proteins (Nrf2, GPX4, xCT, DMT1). Effects of Nrf2 overexpression on ALO-treated cells were observed. RESULTS: ALO inhibited cell viability and increased apoptosis dose-dependently. It elevated lipid peroxidation and intracellular iron while reducing GSH. Ferroptosis inhibitors DFO and Fer-1 reversed cell death and reduced apoptosis. ALO induced ROS production, upregulated Bax/Caspase3, and downregulated Bcl-2. Molecular docking suggested ALO binds to Nrf2 via hydrogen bonding. Immunofluorescence and Western blot showed ALO suppressed Nrf2, GPX4, xCT, and DMT1 expression concentration-dependently. Nrf2 overexpression significantly attenuated ALO's inhibitory effects on proliferation and its induction of ferroptosis and apoptosis. CONCLUSION: ALO suppresses colorectal cancer cell proliferation by inducing apoptosis and ferroptosis via inhibition of the Nrf2 signaling pathway.