iRGD-functionalized amorphous iron phosphate nanoclusters loaded with sulfasalazine activate ferroptosis and reverse mesenchymal transition in glioblastoma

Glioblastoma (GBM) remains a lethal brain malignancy because of blood–brain barrier (BBB)-restricted drug delivery, glioma stem cell (GSC)-driven recurrence, and therapy-induced cellular plasticity. Although ferroptosis induction represents a promising strategy to eliminate treatment-resistant GSCs, the therapeutic potential of sulfasalazine (SAS), a classical system Xc⁻ inhibitor, is limited by poor BBB penetration and inadequate brain tumor accumulation. Here, we engineered an iRGD-functionalized, polydopamine-assisted amorphous iron phosphate nanocluster loaded with SAS, termed iRGD-PDA-AFP-SAS, to integrate BBB-penetrating delivery, iron supply, system Xc⁻ blockade, and GSC-targeted ferroptosis therapy. The nanoclusters displayed stable morphology, homogeneous elemental distribution, amorphous structure, favorable aqueous dispersibility, and high specific surface area. Functionally, iRGD-PDA-AFP-SAS potently suppressed GSC viability, proliferation, neurosphere formation, self-renewal, and stemness marker expression. In vitro BBB models and orthotopic glioma models demonstrated efficient BBB penetration, reduced intracranial tumor burden, decreased Ki67-positive proliferation, and prolonged survival, whereas free SAS showed limited therapeutic efficacy. Mechanistically, iRGD-PDA-AFP-SAS activated ferroptosis in GSCs by increasing Fe²⁺, ROS, and MDA accumulation, depleting GSH, upregulating ACSL4, suppressing SLC7A11/GPX4, and inducing mitochondrial membrane potential collapse and ultrastructural mitochondrial damage; these effects were partially reversed by Ferrostatin-1. Transcriptomic profiling and rescue experiments identified NR4A2 downregulation as a key upstream event driving ferroptosis activation. Importantly, iRGD-PDA-AFP-SAS also reversed proneural-to-mesenchymal transition (PMT), as shown by reduced CD44/YKL-40 expression, increased SOX2/OLIG2 expression, and impaired invasion, while Ferrostatin-1 or NR4A2 overexpression partially restored mesenchymal phenotypes. Moreover, iRGD-PDA-AFP-SAS enhanced temozolomide-based therapy in a patient-derived GSC13 orthotopic glioma model. Together, this study establishes iRGD-PDA-AFP-SAS as a BBB-penetrating ferroptosis nanotherapeutic platform and reveals an NR4A2 downregulation–ferroptosis activation–PMT reversal axis that suppresses GBM progression and improves therapeutic response.