Bioadhesive nanoparticles of fungal chitosan for oral DNA delivery

Chitosan is an ideal candidate for oral DNA delivery due to its mucoadhesive properties. Chitosan (CS) produced under GMP conditions from fungal source was used to encapsulate a plasmid DNA coding for a reporter gene. Nanoparticles made by complex coacervation of CS and DNA had a size around 200 nm, a positive zeta potential, a high association of DNA and protected the plasmid against nuclease degradation. Their transfection ability was assessed in differentiated intestinal Caco-2 cells. An N/P ratio of 4 and a DNA concentration of 8 microg/ml were the optimal conditions leading to a transfection efficiency similar to the one reached with polyethyleneimine (PEI)-DNA complexes without cytotoxicity. M cells in monolayers influenced DNA uptake up to 8 microg of DNA/ml when complexed with CS. Fungal trimethylchitosan was also tested but the complexes interactions were too strong to induce transfection in vitro. Confocal microscopy studies showed that CS/DNA and PEI/DNA nanoparticles were found at the apical surface of cell monolayers and DNA was co-localized within the nucleus. Quantification seemed to show that more DNA was associated with the cells when incubated with CS nanoparticles and that the presence of M cells slightly influenced DNA uptake when complexed with CS. In conclusion, we developed a new nanocarrier made of fungal CS promising for oral gene delivery and oral DNA vaccination.