Bisphosphonate stabilized calcium phosphate nanoparticles for effective delivery of plasmid DNA to macrophages

Calcium phosphate (CP) nanoparticles (NPs) have been used in various applications for delivery of nucleic acid (NA) cargos with ideal biocompatibility and safety. However, some critical issues such as poor stability and aggregation in water solution hinder the industrial application of CP NPs. To further utilize CP NPs for NA delivery, this study specifically focused on the modification of CP NPs to achieve a rapid synthesis and improvement on dispersibility and colloidal stability by using a bisphosphonate (BP) and BSA (named as BCP NPs). Compared with CP NPs, BCP NPs show better stability and dispersity in the cell culture medium, higher efficiency in cellular uptake, and faster dissolution in acidic environments, which are essential requirements for NA vaccine delivery. The cell viability (MTT) assay indicates that BCP NPs have a similar or lower cytotoxicity than free alendronate and Lipofectamine 2000 reagent (L2K) to macrophages (MΦs), a type of typical antigen-presenting cells (APCs). Furthermore, BCP NPs exhibited 85% plasmid DNA (pDNA) loading efficiency and a good endosome escape property. Using a plasmid expressing enhanced green fluorescent protein (pEGFP) as a model system, we showed that BCP NP transfection resulted in a high-level EGFP expression in MΦs, which was even greater than the commercial L2K and electroporation method. This is the first application of a low amount of BP and BSA to modify CP-based NPs with low MΦ cytotoxicity and favorable dispersity, and our data suggest these BCP NPs are an excellent delivery platform for pDNA to MΦs