Systematic studies on the formation of DNA-LDH-nanocomplexes and their application as gene carriers

Over the last decade nanometre-sized layered double hydroxides (LDH) have been successfully explored as cellular delivery agents for various drugs [1, 2] and bio-active molecules such as peptides or DNA [3-5]. Despite its intrinsic promising properties such as good biocompatibility, low cytotoxicity [6, 7], high loading of anionic/polar molecules, pH controllable release, protection of guest molecules in the interlayer [6] and controllable particle size [3, 6, 8], initial transfection experiments showed disappointing performance of LDH compared to commercially available carriers. Therefore synthesis of DNA-LDH-nanocomplexes (LDH) and transfection protocols for mammalian cell lines were optimized, resulting in high transfection efficiencies of up to 60-70 % positive cells. We found that working at fairly high temperatures for DNA-intercalation with much diluted DNA solutions respective LDH suspensions at slightly acidic pH provides the best environment for high DNA uptake. Incubation time and DNA shape (plasmid/linearized) hardly affect the amount of intercalated DNA, whereas the size of DNA fragments seems to have major impact, with smaller plasmids being intercalated at five-fold higher rates than larger ones. Transfection and expression of various GFP plasmids in mammalian cell lines (HEK 293T, NIH 3T3, COS7) gave positive cells with DNA levels as low as 0.5 μg/well (Ø 35 mm). Production of LDH nanoparticles is less costly than for liposome based commercial competitors while showing similar efficiency, providing an alternative transfection agent with superior cost-performance-ratio