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Abstract
Background
Small interfering RNA (siRNA)-mediated gene therapy is a promising strategy to temporarily inhibit the expression of genes involved in development of breast cancer. The lack of a safe and efficient gene delivery system has become a major hurdle for siRNA-mediated gene therapy in breast cancer. Our previous studies have demonstrated that inorganic amorphous calcium carbonate (ACC) hybrid nanospheres functionalized with CaIP6 (ACC/CaIP6) nanoparticles are an efficient nucleic acid delivery tool. The present study aimed to evaluate the safety and efficiency of ACC/CaIP6 in delivering siRNA targeting AKT1 (siAKT1) for the treatment of breast cancer.
Methods
The cytotoxicity of the ACC/CaIP6 nanoparticles was evaluated using a tetrazolium assay. The transfection efficiency and intracellular distribution of ACC/siAKT1 were analyzed by flow cytometry and confocal laser scanning microscopy, respectively. A series of in vitro and in vivo assays was performed to evaluate the effects of ACC/CaIP6/siAKT1 on growth of breast cancer cells.
Results
ACC/CaIP6 nanoparticles effectively transfected cells with little or no toxicity. AKT1 knockdown by ACC/CaIP6/siAKT1 inhibited cell cycle progression and promoted apoptosis of MCF-7 cells. Intratumoral injection of ACC/CaIP6/siAKT1 significantly suppressed the growth of breast cancer in mice.
Conclusion
ACC/CaIP6 nanoparticles are a safe and efficient method of delivering siRNA for gene therapy in breast cancer.
Supplementary materials
Figure S1 Intracellular distribution of ACC/CaIP6/FAM-siAKT1 in MCF-7 cells was analyzed by confocal laser scanning microscopy after incubation for 8 hours. Lysosomes were stained with Lyso Tracker Red, a percentage of the fluorescence of FAM-siAKT (green) did not overlap with the fluorescence of lysosomes (red), indicating successful escape of the FAM-siRNA from the lysosomes to the cytoplasm.
Abbreviations: ACC/CaIP6, amorphous calcium carbonate hybrid nanospheres functionalized with a Ca(II)-inositol hexakisphosphate compound; FAM-siAKT1, fluorescein-labeled AKT1-specific small interfering RNA.
Figure S2 Real-time quantitative polymerase chain reaction analysis of the relative expression of AKT1 mRNA in liver samples intratumorally injected with ACC, siAKT1, and ACC/CaIP6/siAKT1 nanoparticles.
Abbreviations: ACC/CaIP6, amorphous calcium carbonate hybrid nanospheres functionalized with a Ca(II)-inositol hexakisphosphate compound; siAKT1, small interfering AKT1.
Acknowledgments
This work was supported by the National Basic Research Program of China (2010CB934700) and the National Natural Science Foundation of China (30900539, 81172429, 81372821).
Disclosure
The authors report no conflicts of interest in this work.