A Polyplex in a Shell: The Effect of Poly(aspartic acid)-Mediated Calcium Carbonate Mineralization on Polyplexes...

Mineralization by exposure of organic templates to supersaturated solutions is used by many living organisms to generate specialized materials to perform structural or protective functions. Similarly, it was suggested that improved robustness acquired through mineralization under natural conditions could be an important factor for virus survival outside of a host for better transfection of cells. Here, inspired by this fact, we developed a nonviral tricomponent polyplex system for gene delivery capable of undergoing mineralization. First, we fabricated anionic polyplexes carrying pDNA by self-assembly with a lipid-modified cationic polymer and coating by poly(aspartic acid). Then, we submitted the polyplexes to a two-step mineralization reaction to precipitate CaCO₃ under various supersaturations. We carried out detailed morphological studies of the mineralized polyplexes and identified which parameters of the fabrication process were influential on transfection efficiency. We found that mineralization with CaCO₃ is efficient in promoting transfection efficiency as long as a certain Ca²⁺/CO₃^2- lower limit ratio is respected. However, calcium incubation can also be used to achieve similar effects at higher concentrations depending on polyplex composition, probably due to the formation of physical cross-links by calcium binding to poly(aspartic acid). We proposed that the improved robustness and transfection efficiency provided by means of mineralization can be used to expand the possible applications of polyplexes in gene therapy.