Glutathione-Responsive Disulfiram Delivery from Thiol-Functionalized N-Doped Carbon Nanodots: An in vitro Proof-of-Concept Study.

PURPOSE: To develop and evaluate a glutathione (GSH)-responsive "Off-On" drug delivery system based on thiol-functionalized N-doped carbon dots (SNCDs) for controlled intracellular release of disulfiram (DSF). MATERIALS AND METHODS: SNCDs were synthesized using a solvothermal technique and subsequently characterized using dynamic laser scattering, high-resolution electron imaging, UV-Vis spectrophotometry, and Fourier-transform infrared spectroscopy (FTIR). DSF was immobilized onto SNCDs through redox-cleavable disulfide linkages, and loading efficiency was determined by UV-Vis analysis. The in vitro drug release kinetics were examined under simulated physiological (pH 7.4) and tumor-relevant acidic (pH 5.4) conditions, evaluating the impact of glutathione (GSH) by performing experiments with and without its inclusion. Cytotoxicity and compatibility studies in MCF-7 breast tumor cells, evaluated via MTT test, were used to investigate SNCD biocompatibility and the anticancer performance of DSF-SNCDs. RESULTS: The FTIR spectra confirmed effective thiol functionalization and revealed oxygenated and nitrogenous groups on the particle surface suitable for disulfide linkage formation. DSF loading efficiency reached approximately 75%. Under non-reducing conditions, DSF-SNCDs exhibited minimal premature release (≤30% over 24 h at pH 7.4 and 5.4), demonstrating an "Off" state. In contrast, the presence of 5% (w/v) GSH at pH 5.4 triggered rapid DSF release, achieving approximately 80% cumulative release within 24 h, corresponding to the "On" state through reductive disulfide bond cleavage. Bare SNCDs showed high biocompatibility (>85% cell viability at 50 µg/mL), whereas DSF-SNCDs produced significantly enhanced cytotoxicity under GSH conditions, confirming redox-activated drug release. CONCLUSION: SNCDs provide an effective GSH-responsive nanoplatform for DSF delivery, combining high drug loading, excellent stability under non-reducing conditions, and efficient intracellular drug release under reductive environments. This strategy minimizes premature drug leakage while enhancing anticancer activity, demonstrating strong potential for cancer therapy.