Object: Gold nanoparticles (GNPs) are being studied as potential radiosensitizers for cancer treatment because of their high atomic number. However, their accumulation in tumors can be limited by nonspecific uptake of the reticuloendothelial system (RES) and high tumor interstitial fluid pressure. Macrophages present in the RES are ideal carriers for GNP delivery due to their multiple capabilities, including phagocytosis, tumor-homing, plasticity, and hypoxia tropism. The current study investigates the potential of utilizing macrophages as tumor-specific carriers for GNP delivery and their enhancement of therapeutic efficacy and related mechanism after radiotherapy (RT) in a murine oral cancer model in cell culture and tumor-bearing mice.

Methods: The murine oral cancer cell line MCTQ1 and murine macrophage cell line Raw 264.7 were utilized in this study. Gold nanoparticles (GNPs) with diameters of 20 nm and 40 nm were created using the Turkevich method. The impact of GNP-mediated radiosensitization was evaluated through cell viability and colony formation assays. ¹³¹I-labeled GNPs were produced via the chloramine-T addition reaction at room temperature and applied to Raw 264.7 cells to investigate their cellular uptake of GNPs, determining the optimal incubation time for generating GNP-loaded Raw 264.7 cells (GNP@Rs). The migration ability of Raw 264.7 and GNP@Rs was examined using the transwell assay in vitro, while the tumor tropism of Raw 264.7 cells was investigated by monitoring the movement of fluorescence-labeled Raw264.7 cells with bioluminescence imaging in mice bearing subcutaneous MTCQ1 tumors. Subsequently, tumor-bearing mice were divided into Control, RT, GNP+RT, Raw+RT, and GNP@R+RT. GNPs, Raw 264.7 cells, and GNP@Rs were administered via intravenous injection on day 1 followed by an 8-Gy RT treatment on day 5. Tumor volumes were measured every three days to create tumor growth curves.

Results: 40 nm GNPs showed a stronger radiosensitization effect compared to 20 nm GNPs, as demonstrated by the cell viability assay. The labeling efficiency and purity of ¹³¹I-GNPs were both above 95%, with stable radio-stability observed during incubation in FBS and normal saline. The cellular uptake test indicated that the optimal conditions for producing GNP@Rs involved incubating Raw 264.7 cells with 40 nm GNPs at a concentration of 100 ppm for 6-8 hours. In vitro migration studies confirmed that Raw264.7 cells maintained their migratory ability after being loaded with GNPs in a regular medium and tumor-conditioned medium. Furthermore, in vivo imaging tracking revealed that the tumor-homing behavior of Raw 264.7 cells remained intact following GNP incubation, as evidenced by fluorescence signals persisting at tumor sites for up to 96 hours post-injection. Tumor growth monitoring results demonstrated that combining GNP@Rs with radiotherapy led to the most significant tumor, suggesting the potential of using macrophage-mediated GNP delivery for cancer radiotherapy purposes.

Conclusion: Our research found that 40 nm GNPs have a better radiosensitization effect compared to their 20 nm counterparts. We also developed a macrophage-based GNP delivery system, which proved to have potent radiosensitization effects on mice with MTCQ1 tumors. These findings suggest a promising strategy for cancer treatment.

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