Abstract:

Objective ·To develop a biomimetic membrane, glutathione (GSH)-responsive manganese-based theranostic nanoprobe, and investigate its biomembrane-targeting capability, efficacy in GSH-depletion-mediated ferroptosis induction in triple-negative breast cancer (TNBC), and real-time monitoring of MR signal activation. Methods ·A silica (SiO₂) nanoparticle core was synthesized via the sol-gel method, followed by deposition of manganese dioxide (MnO₂) onto its surface through redox co-precipitation, and then coating with 4T1 breast cancer cell membranes using an extrusion technique to yield the biomimetic nanoprobe SiO₂@MnO₂@Membrane (SMM). The morphology, particle size, and Zeta potential were characterized by transmission electron microscopy (TEM) and dynamic light scattering (DLS). Membrane protein retention on the nanoparticle surface was confirmed by Coomassie brilliant blue staining, and characteristic elements was analyzed by X-ray photoelectron spectroscopy (XPS). The GSH-responsive T₁ MR signal activation was evaluated in vitro. Reactive oxygen species (ROS) generation and GSH depletion in a cell-free system were assessed using methylene blue and 5, 5′-dithiobis (2-nitrobenzoic acid) (DTNB) assays, respectively. The cytotoxic effect of the nanoprobe on 4T1 cells was determined by the MTT assay. Intracellular ROS generation and lipid peroxide (LPO) accumulation were detected using 2′,7′-dichlorodihydrofluoresce in diacetate (DCFH-DA) and BODIPY 581/591 C11 probes. A murine subcutaneous TNBC model was established to dynamically monitor T₁-weighted MR signal activation of the nanoprobe at tumor sites. Combined with the detection of ferroptosis-related markers, including LPO and ROS, as well as Ki-67 and terminal-deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) staining in tumor tissues, the ferroptosis-mediated antitumor efficacy of SMM was evaluated. Results ·TEM revealed that the prepared SMM nanoprobe exhibited a regular spherical structure. Particle size analysis showed that after cell membrane coating, the Zeta potential shifted from (+3.2±1.0) mV to (-22.0±2.0) mV, and the hydrodynamic diameter increased from 68 nm to 78 nm, while the polydisper sity index (PDI) remained at a low level, indicating a uniform and stable dispersion. Coomassie brilliant blue staining confirmed effective biomimetic membrane coating. XPS detected characteristic signals of Mn and Si elements, and these results collectively confirmed the successful synthesis of the SMM nanoprobe. MR results showed that SMM significantly activated the T₁ signal in the presence of GSH, with relaxivity (r₁). increasing significantly from 0.591 to 6.875 (mmol/L)^-1·s^-1. At the cellular level, T₁ signal activation exhibited a time-dependent enhancement. Compared to the control groups, DCFH-DA and BODIPY 581/591 C11 staining indicated substantial intracellular accumulation of ROS and LPO in the SMM group, accompanied by a significant reduction in cell viability (28.35%±4.91% survival). In vivo experiments showed that after intravenous injection of SMM,the T₁ signal intensity in tumor regions reached a peak value of (1.23±0.14) s^-1 at 4 h, indicating that the nanoprobe could specifically accumulate in TNBC tumor tissues and activate MR T₁ signals. Tumor tissue analysis revealed significantly elevated ROS and LPO levels in the SMM group, accompanied by decreased Ki-67 expression and increased TUNEL-positive signals, further confirming its antitumor effect through ferroptosis induction. Conclusion ·A GSH-depleting 4T1 cell membrane-biomimetic nanoprobe (SMM) was successfully developed, which can specifically activate T₁ MR signals at TNBC tumor sites and synergistically induce ferroptosis through targeted accumulation, significantly enhancing antitumor efficacy.

Key words: ferroptosis, biomimetic membrane, triple-negative breast cancer, magnetic resonance imaging, manganese-based nanoparticle, glutathione depletion, reactive oxygen species