Ferrocene–Cystamine Conjugate Encapsulated Hyaluronic Acid Nanomicelles with Sequential pH/GSH Responsiveness for Synergistic Chemodynamic-Immunotherapy

Abstract Metastasis and postoperative recurrence of malignant tumors remain critical obstacles that undermine clinical therapeutic efficacy. Traditional chemodynamic therapy (CDT) is plagued by three inherent, hard-to-resolve drawbacks: abundant intratumoral glutathione (GSH) quenches reactive oxygen species; robust systemic antitumor immunity fails to be activated; and unbound iron catalysts initiate off-target Fenton cascades within healthy tissues, inducing systemic oxidative injury and toxic side effects. To address these critical barriers, we rationally designed a bifunctional ferrocene–cystamine conjugate (FD) and encapsulated this payload within amphiphilic hyaluronic acid–cinnamaldehyde (HA-CA) Schiff-base nanomicelles, thereby fabricating a nanoplatform designed to target HA receptors (presumably CD44) with sequential pH/GSH responsiveness. The resultant HA-CA@FD nanomicelles attain a high FD loading efficiency of 36 ± 2% and display excellent colloidal stability under physiological environments. Once exposed to the acidic, reductive tumor microenvironment, the nanocarrier undergoes structural dissociation to liberate loaded FD. The released FD depletes intracellular GSH through two cooperative redox routes—disulfide bond cleavage and reversible Fe2+/Fe3+ cycling—while simultaneously generating massive cytotoxic hydroxyl radicals (·OH) to elicit potent immunogenic cell death. In vitro biological characterizations validate that the nanomedicine raises the proportion of mature dendritic cells to 66.7% and elevates the percentage of anti-tumor M1-type macrophages from 3.9 to 48.2%. In vivo assays using bilateral 4T1 tumor-bearing mice reveal that intratumoral injection of HA-CA@FD diminishes primary tumor weight by 67% and abscopal contralateral tumor weight by 52%. This therapeutic regimen substantially augments intratumoral infiltration of CD4+/CD8+ effector T cells and diminishes immunosuppressive regulatory T cell (Treg) populations, accompanied by elevated memory T-cell signatures that suggest potential for establishing durable antitumor immune responses. This modular, biocompatible biopolymer nanosystem surmounts the core limitations of classic CDT agents, furnishing a straightforward, translation-ready nanoengineering paradigm for precision combinatorial chemo-dynamic-immunotherapy against solid tumors.