Enhancing Osteosarcoma Killing and CT Imaging Using Ultrahigh Drug Loading and NIR-Responsive Bismuth Sulfide@Mesoporous Silica Nanoparticles

Abstract

Despite its 5-year event-free survival rate increasing to 60-65% due to surgery and chemotherapy, osteosarcoma (OS) remains one of the most threatening malignant human tumors, especially in young patients. Therefore, a new approach that combines early diagnosis with efficient tumor eradication and bioimaging is urgently needed. Here, a new type of mesoporous silica-coated bismuth sulfide nanoparticles (Bi₂ S₃ @MSN NPs) is developed. The well distributed mesoporous pores and large surface areas hold great promise for drug protection and encapsulation (doxorubicin (DOX), 99.85%). Moreover, the high photothermal efficiency of Bi₂ S₃ @MSNs (36.62%) offers great possibility for cancer synergistic treatment and highly near-infrared-triggered drug release (even at an ultralow power density of 0.3 W cm^-2 ). After covalently conjugated to arginine-glycine-aspartic acid (RGD) peptide [c(RGDyC)], the NPs exhibit a high specificity for osteosarcoma and finally accumulate in the tumor cells (tenfold more than peritumoral tissues) for computed tomography (CT) imaging and tumor ablation. Importantly, the synergistic photothermal therapy-chemotherapy of the RGD-Bi₂ S₃ @MSN/DOX significantly ablates the highly malignant OS. It is further proved that the superior combined killing effect is achieved by activating the mitochondrial apoptosis pathway. Hence, the smart RGD-Bi₂ S₃ @MSN/DOX theranostic platform is a promising candidate for future applications in CT monitoring and synergistic treatment of malignant tumors.

Keywords: Bi2S3@MSN; X-ray computed tomography; mitochondrial apoptosis pathway; osteosarcoma; photothermal therapy-chemotherapy.

Figure 1.

Characterization of the Bi₂S₃@MSN. a) XRD spectra of the Bi₂S₃ NPs and the JCPDS standard for Bi₂S₃. b) TEM image of the Bi₂S₃@MSN. Scale bar = 100 nm. c) Nitrogen adsorption/desorption isotherms. d) Pore size distribution. e) UV–vis–NIR absorption spectrum of the Bi₂S₃@MSN and RGD–Bi₂S₃@MSN, the RGD–Bi₂S₃@MSNs exhibit a new peak in 270 nm, indicating the successful conjugation of RGD. f) Quantitative temperature change of Bi₂S₃@MSN different concentrations under 1 W cm^–2 808 nm laser irradiation.

Figure 2.

NIR-triggered release of DOX from Bi₂S₃@MSN at varied power densities. a) Illustration of drug release behavior with or without NIR laser irradiation. Burst drug release occurred after applying NIR laser irradiation. b) Photothermal conversion efficiency (η) of the NPs. Changes in drug release and temperature under c) 0.3 W cm^–2, d) 0.5 W cm^–2, e) 1 W cm^–2, f) and 0 W cm^–2 NIR irradiation.

Figure 3.

In vitro active targeting effect of the RGD–Bi₂S₃@MSN. a) Fluorescence images ofUMR-106 cells cocultured with free FITC, FITC–Bi₂S₃@MSN, and FITC–RGD–Bi₂S₃@MSN. b) Flow cytometry analysis of UMR-106 cells stained by the free FITC, FITC–Bi₂S₃@MSN, and FITC–RGD–Bi₂S₃@MSN. c) Quantitative analysis of fluorescence intensity of different groups in (b). Scale bar = 50 μ.m. Each value is the mean ± standard deviation (n = 3 in each group); ^*p < 0.05, ^**p < 0.01, ^***p < 0.001 compared to the control group.

Figure 4.

Biocompatibility and tumor cells killing effect in vitro. a) Cell viability of UMR-106 cells cocultured with different Bi₂S₃@MSN concentrations for 24 h. b) Hemolysis test of human RBCs exposed to the different concentrations of Bi₂S₃@MSN. The inset figure shows the picture of the centrifuge tubes after centrifugation. (+) represents the purified water, (–) represents the PBS solution. c) Tumor cells killing effect of different groups. The RGD–Bi₂S₃@MSN/DOX+NIR exhibited the most significantly killing effect. d) Fluorescence microscopy images of live/dead dye-stained UMR-106 cells after co-cultured with different NPs and treatment with or without 808 nm (1 W cm^–2) NIR irradiation. Scale bar = 50 μ.m. e) Effects of different treatments on the express levels of Bcl-2 and caspase-3 proteins in UMR-106 cells after cocultured for 24 h using western blot. Each value is the mean ± standard deviation (n = 3 in each group); ^*p < 0.05, ^**p < 0.01, ^***p < 0.001 compared to the control group.

Figure 5.

CT imaging performance of the NPs. a) In vitro CT value (HU) of Bi₂S₃@MSN and iobitridol. Inset: CT images of the Bi₂S₃@MSN and iobitridol suspensions with different concentrations. b) In vivo CT images of UMR-106 tumor-bearing nude mice recorded at 2 and 24 h after i.v. injection of the RGD–Bi₂S₃@MSN and Bi₂S₃@MSN (dosage: 200 of Bi₂S₃@MSN (5 mg kg^–1)). The tumor site is highlighted by the red circle.

Figure 6.

In vivo therapy effect of different treatment groups. a) NIR photothermal images of UMR-106 bearing nude mice after i.v. injection of RGD–Bi₂S₃@MSN and Bi₂S₃@MSN. b) Quantitative temperature change of the tumor site from (a). c) Representative images of tumor bearing mice from different treatment groups. d) Images of tumor collected from tumor bearing mice with different treatment. e) Tumor volume growth curves of different treatment groups. f) Body weight changes in different treatment groups. Each value is the mean ± standard deviation (n = 4 in each group); ^*p < 0.05, ^**p < 0.01, ^***p < 0.001 compared to the control group.

Figure 7.

Histological study and western blot. a) H&E staining of tumors collected from different treatments. b) Effects of RGD–Bi₂S₃@MSN/DOX, RGD–Bi₂S₃@MSN+NIR, and RGD–Bi₂S₃@MSN/DOX+NIR on Bcl-2 and caspase-3 protein expression in tumor tissues using western blot. c) H&E staining of main organs in different treatment groups.

Scheme 1.

The smart RGD-Bi₂S₃@MSN/DOX nanoplatform for OS real-time X-ray CT imaging and NIR-responsive photothermal therapy-chemotherapy.