Application of Strontium\nChloride Hexahydrate to Synthesize\nStrontium-Substituted Carbonate Apatite as a pH-Sensitive, Biologically\nSafe, and Highly Efficient siRNA Nanocarrier

Naked siRNAs are sensitive to enzymatic degradation,\nphagocytic\nentrapment, quick renal excretion, membrane impermeability, endosomal\nescape, and off-target effects. Designing a safe and efficient nanocarrier\nfor siRNA delivery to the target\nsite without toxicity remains a significant hurdle in gene therapy.\nCA is a unique derivative of hydroxyapatite and a highly pH-sensitive\nnanocarrier with strong particle aggregation and a high polydispersity\nindex. Strontium (Sr<sup>2+</sup>), a group two divalent metal in\nthe periodic table, has been reported for substituting calcium (Ca<sup>2+</sup>) ions from the apatite lattice and limiting particle growth/aggregation.\nThis study used strontium chloride hexahydrate (SrCl<sub>2</sub>·6H<sub>2</sub>O) salt to develop a Sr-substituted CA (Sr-CA) nanocarrier\nwith ∼30 nm size, spherical shape, less aggregation, homodispersity,\nand a fair anionic charge. Sr-CA demonstrated a large surface area-to-volume\nratio, an improved cargo loading efficiency, and enhanced cellular\nuptake in HEK-293 cells. Moreover, Sr-CA is a pH-responsive nanocarrier\nresponsible for its long physiological stability, efficient endosomal\nescape, and optimal cargo delivery within cells. These NPs have differential\neffects on MAPK1, MAP2K4, PIK3Ca, CAMK4, and p53 gene expression in\nHEK-293 cells without showing any significant cytotoxicity in cell\ngrowth properties. Gene silencing by Sr-CA-mediated siRNA delivery\nagainst MAPK1, MAP2K4, PIK3Ca, and CAMK4 genes significantly decreased\nthe level of target gene expression and cell survival, demonstrating\nsuccessful intracellular siRNA delivery in HEK-293 cells. Additionally,\nbiocompatibility testing confirmed the biological safety of the Sr-CA\nnanocarrier in mice. These findings suggest that Sr-CA nanocarriers\nare a promising siRNA delivery system, combining high efficiency with\npH-sensitive release and excellent biocompatibility, making them a\nviable option for future therapeutic applications.