Metal-organic nanostructures based on sono/chemo-nanodynamic synergy of TixOy/Ru reaction units: for ultrasound-induced dynamic cancer therapy

Abstract

Sonodynamic therapy (SDT) exhibits clinical potential for deep-tissue tumor treatment due to its deep tissue penetration and spatiotemporal controllability. Its core mechanism relies on ultrasound-activated sonosensitizers to generate reactive oxygen species (ROS), thereby inducing tumor cell apoptosis. However, conventional sonosensitizers face limitations in ROS yield and tumor-targeting efficiency. In this study, we innovatively designed a multifunctional metal-organic nanosheet (TiZrRu-MON) by hydrothermal coordination of [Ru(bpy)₃]²⁺ photosensitizing units with TiZr-O clusters, while incorporating Fe³⁺ to construct a cascade catalytic system. Experimental results demonstrated that: (1) Fe³⁺ lattice doping significantly enhanced charge carrier mobility and ultrasound-triggered ¹O₂ quantum yield via the formation charge transfer channels; (2) The acidic tumor microenvironment activated Fe³⁺-mediated Fenton reactions, establishing a positive feedback loop with SDT to synergistically amplify ROS generation; (3) Hyaluronic acid functionalization improved nanosheet internalization in HepG2 tumor cells through CD44 receptor-mediated endocytosis. Remarkably, ultrasound irradiation induced substantial oxidative stress and immunogenic cell death, promoting the release of damage-associated molecular patterns (DAMPs), which elevated the maturation rate of tumor-infiltrating dendritic cells (DCs) and significantly increased the proportion of CD8⁺ T cells. In a mouse subcutaneous tumor model, the system achieved effective tumor suppression with manageable systemic toxicity. This work proposes a metal-ligand coordination strategy to advance the development of high-performance sonosensitizers and immunomodulatory antitumor technologies.

Keywords: Cancer therapy; Chemodynamic therapy; Magnetic resonance imaging; Metal–organic nanostructure; Sonodynamic therapy.

Scheme 1

Schematic illustration of TiZrRuFe-MON@HA (MFH) and synergistic inhibitory mechanism of sono/chemo-dynamic therapies for HCC. Schematic diagram of the synthetic process of MFH by hydrothermal-chelation-coating method. Synergetic-enhanced ROS generation of MFH for sono/chemo-dynamic therapeutic process under US irradiation against HCC. Under US irradiation, MF is activated, leading to the separation of e^⁻/h^⁺, which react with O₂ to generate ¹O₂. Additionally, Fe²⁺ reacts with H₂O₂ to produce •OH and Fe³⁺, which is subsequently reduced back to Fe²⁺ by e⁻, forming a Fe³⁺/Fe²⁺ cycle (CDT). The synergistic effects of SDT and CDT demonstrate potent anti-tumor efficacy

Fig. 1

a TEM of TiZrRu-MON (M) (Scale bars: 200 nm), b TiZrRuFe-MON (MF) (Scale bars: 200 nm), c TiZrRuFe-MON@HA (MFH) (Scale bars: 200 nm). d Average size and e Zeta potential of M, MF and MFH. f UV–Vis absorption of the TiZr-cluster, Ru-COOH, M, MF and MFH. g, h, i XPS spectra of MF. h Ru 3p, i Fe 2p. j HAADF image and k EDS elemental mapping images of MF (Scale bars: 100 nm). l Relaxation intensity of M, Fe³⁺, and MF at different concentrations and T1-weighted MR images of MF

Scheme 2

Schematic representation of the SDT mechanism of MF under US irradiation and ROS generation detected by the probes. The reaction of •OH with TMB induces a color transition of the solution from colorless to blue, while the interaction of ¹O₂ with DPBF results in a color change from yellow to colorless

Fig. 2

a ESR signal of ¹O₂ captured by TEMP. (US power density: 1.0 W/cm², duty cycle: 50%, 5 min) b The SDT performance of DI water, M, and MF through changes in the UV–Vis absorption spectrum of DPBF. (US power density: 1.0 W/cm², duty cycle: 50%, 5 min) c The SDT performance of MF under different US irradiation times using the UV–Vis absorption spectrum of DPBF. (US power density: 1.0 W/cm², duty cycle: 50%, 5 min) d ESR signal of •OH captured by DMPO. (H₂O₂, 100 μM) e The CDT performance of DI water, Fe³⁺, M, and MF via the UV–Vis absorption spectrum of TMB. (H₂O₂, 100 μM) f The CDT performance of MF at different concentrations using the UV–Vis absorption spectrum of TMB. H₂O₂ (H₂O₂, 100 μM) g, h Evaluation of •OH generation by MF (Zr concentration, 100 μM) under no g or with h US conditions via TMB UV–Vis absorption spectroscopy. (H₂O₂, 100 μM) i Measurement of ROS levels in cells from different treatment groups. a Control, b SDT, c CDT, d SDT + CDT. (H₂O₂, 100 μM; US power density: 1.0 W/cm², duty cycle: 50%, 5 min) *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001; ns = not significant

Fig. 3

a TEM image of HepG2 cells co-cultured with MFH. b HepG2 cells were analyzed by 1.5 T MR scanning after uptake of MF or MFH. c Cellular uptake of rhodamine B-labeled MF and MFH were assessed via fluorescence microscopy after 1, 4, 8, and 12 h (Scale bars: 20 μm). d Zr concentration analysis after cellular uptake of MF or MFH after 1, 4, 8, and 12 h. e CLSM images of HepG2 cells after different treatments to detect the production of ROS labeled by DCFH-DA. (Scale bars: 100 μm). (US power density: 1.0 W/cm², duty cycle: 50%, 5 min). (F) Flow cytometric profiles of ROS generation. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001; ns = not significant

Fig. 4

a The viability of HepG2 cells from different treatment groups. a SDT, b CDT, c SDT + CDT. b Colony formation assays of different treatments. a Control, b SDT, c CDT, d SDT + CDT. c Apoptosis analysis of HepG2 cells post-treatment via flow cytometry using Annexin V-FITC/PI dual staining. d Live/dead fluorescence imaging of HepG2 cells after different treatments, stained with calcein-AM (green, live) and PI (red, dead) (Scale bars: 100 μm). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001; ns = not significant

Fig. 5

MFH + US elicited enhanced ICD and promoted DC maturation in vitro. a Representative CLSM images showing the remanent of HMGB1 from Hepa1-6 cells. Scale bar = 20 μm. b Extracellular ATP levels in Hepa1-6 cells post-treatments (n = 3). c Representative CLSM images showing the CRT surface exposure on Hepa1-6 cells. Scale bar = 20 μm. d FCM analysis of CRT expression on Hepa1-6 cell surfaces. e, f Semi-quantitative assessment of BMDCs maturation following co-culture with differentially pretreated Hepa1-6 cells, analyzed by FCM (n = 3). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001; ns = not significant

Fig. 6

Biodistribution and therapeutic efficacy of MFH + US in HepG2 tumor-bearing nude mice. a Schematic of tumor modeling and therapeutic protocol (US: 1.0 W/cm², 50% duty cycle, 5 min). b, c In vivo MR imaging of subcutaneous tumors following i.v. injection of MF/MFH at indicated timepoints (0–24 h). d Body weight changes and e Tumor growth curves post-treatments. f Excised tumor photographs. g Histological analysis: H&E, Ki-67 and TUNEL immunohistochemistry (Scale bar = 100 μm). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001; ns = not significant

Fig. 7

MFH + US enhanced sonodynamic immunotherapy in Hepa1-6 subcutaneous tumors. a Schematic of tumor modeling and therapeutic protocol (US: 1.0 W/cm², 50% duty cycle, 5 min). b Tumor growth curves and c weights across groups. d Body weight monitoring. e, f FCM analysis of tumor-infiltrating mature DCs (CD80⁺CD86⁺): e representative plots and f quantification. (n = 3) g, h FCM analysis of tumor-infiltrating CD8⁺ T-cell (CD3⁺CD8⁺): g representative plots and h quantification (n = 3). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001; ns = not significant