Cu2+-Citrate-Chitosan Complex Nanoparticles for the Chemodynamic Therapy of Lung Cancer

Abstract

Lung cancer poses a significant threat to human health. Surgical intervention is the preferred treatment modality for lung cancer, but a large number of patients are deprived of the opportunity for surgery for various reasons and are compelled to undergo radiotherapy and chemotherapy, which entail systemic adverse reactions. In recent years, with the advancement of nanomedicine, chemodynamic therapy (CDT) based on free radicals has been extensively investigated. In this study, we fabricated copper-citrate-chitosan composite nanoparticles (CuCC NPs) by encapsulating copper-citrate complexes with natural chitosan polymers, resulting in a substantial reduction in the biotoxicity of copper ions. The CuCC NPs selectively accumulated in tumor tissues through the enhanced permeability and retention effect (EPR) and gradually degraded within the acidic and glutathione (GSH)-rich microenvironment of the tumor, thereby releasing the loaded copper ions. Through CDT, the copper ions converted the overexpressed hydrogen peroxide (H₂O₂) in the tumor tissue into hydroxyl radicals (•OH), leading to the eradication of tumor cells. In animal experiments, CuCC NPs exhibited remarkable efficacy in CDT. Further histopathological and hematological analyses demonstrated that CuCC NPs could induce substantial apoptosis in tumor tissues while maintaining an extremely high level of safety.

Figure 1

(a) TEM image of CuCC NPs. (b) XRD spectrum of CuCC NPs. (c) FTIR spectrum of CuCC NPs, chitosan, citrate, and CuCl₂. (d) ESR spectra of CuCC NPs with different treatments.

Figure 2

Cytotoxicity of CuCC NPs on (a) BEAS-2B and (b) A549 cells. Coincubation of (c) BEAS-2B and (d) A549 cells with CuCC NPs, followed by CLSM observation of intracellular •OH generation; the scale bar is 50 μm.

Figure 3

(a) CLSM detection of •OH generation in A549 cells; the scale bar is 50 μm. (b) Flow cytometry analysis of ROS-positive A549 cells after coincubation with CuCC NPs at different concentrations.

Figure 4

A549 cells were coincubated with CuCC NPs at different concentrations, followed by assessment of cell status using (a) calcein/PI cell viability/cytotoxicity assay kit and (b) annexin-V FITC/PI cell apoptosis assay kit. The scale bar is 50 μm.

Figure 5

CDT treatment in vivo: (a) Tumor volume curves for each group. (b) Average body weight curves for each group. (c–e) A549 tumor-bearing mice before treatment (on the left) and after treatment (on the right) and photographs of tumors in different groups after the treatment (below).

Figure 6

(a) Histological sections of tumor tissues from each group stained with H and E. The scale bar is 100 μm. (b) Tunel-stained sections of tumor tissues from each group. The scale bar of is 50 μm. Immunofluorescence detection of apoptosis-related proteins (c) Bax and (d) Bcl-2 expression. The scale bar of is 50 μm.