Plumbagin Induces Ferroptosis in Nonfunctioning Pituitary Adenomas via Nrf2/FTH1-Dependent Ferritinophagy

Background: Nonfunctioning pituitary adenoma (NFPA) is among the most prevalent intracranial tumors. As surgical resection remains the primary treatment, there is a need for new therapeutic strategies. Plumbagin (PLB), a bioactive naphthoquinone compound derived from the roots of Plumbaginaceae Juss, has been used in anti-tumor research in recent years, but its mechanism remains controversial.Purpose: The aim of this study was to explore the molecular mechanism of PLB-induced ferroptosis in NFPA.Methods: The cell viability of PLB against PDFS cells was first assessed by CCK-8, colony formation, and EDU assays. Network pharmacology, molecular docking, and molecular dynamics were then used to screen out targets associated with ferroptosis. Finally, the Nrf2/FTH1 pathway was validated by a 3D multicellular tumour spheroids (MCTSs) model, q-PCR, and Western blot in vitro and in vivo.Results: Compared to temozolomide (TMZ), PLB more potently inhibited the viability of PDFS cells. In vitro, PLB treatment was associated with downregulation of the Nrf2/FTH1 pathway, accompanied by increased lipid peroxidation and labile iron pool (LIP) accumulation. Furthermore, PLB treatment led to the upregulation of NCOA4 and an increase in intracellular free iron, consistent with ferritinophagy activation. These effects collectively point to the induction of ferroptosis.Conclusion: This study demonstrates that PLB may induce ferroptosis in PDFS cells. The mechanism involves suppression of the Nrf2/FTH1 pathway, subsequent upregulation of NCOA4, and ferritinophagy-driven iron overload. These results identify PLB as a potential ferroptosis inducer and warrant further investigation into its therapeutic potential for NFPA. The image shows a biochemical pathway illustrating ferritinophagy and ferroptosis. Iron in the form of Fe superscript 3 is transported by transferrin into the cell, where it is stored in ferritin. Ferritin is directed to the autolysosome, facilitated by Lamp2, where iron is released as Fe superscript 2. NCOA4 and LC3 are involved in ferritinophagy, leading to the labile iron pool. The Fenton reaction produces reactive oxygen species, contributing to ferroptosis. Nrf2 regulates FTH1, with arrows indicating promotion and inhibition. Plumbagin is shown affecting the pathway, with an illustration of a mouse receiving an injection, highlighting the interaction between NCOA4, FTH1 and Nrf2 in the brain. Arrows indicate promotion and inhibition effects within the pathway.Diagram of ferritinophagy and ferroptosis: iron regulation, NCOA4, FTH1, Nrf2 and plumbagin effects.