Formulation of PEG–folic acid coated nanometric DNA particles from perfluoroalkylated cationic dimerizable detergents and in vitro folate-targeted intracellular delivery

Present attempts in gene delivery mediated with synthetic vectors are now oriented towards the formulation of nanometric DNA complexes reminiscent of viruses. The assembly of such multifunctional gene transfer systems was designed in order to take advantages of viral vector properties. This requires, among others, the formulation of very small- and homogeneously-sized “stealth” as well as ligand-labeled DNA nanoparticles. In this paper, we report the straightforward synthesis of an original dimerizable perfluoroalkylated thiol detergent deriving from cysteine and containing a monocationic aminotriethylene glycol and of a known analog containing a tricationic linear spermine polar head (three steps, 45–50% overall yields). These derivatives condensed DNA into a monodisperse population of negatively or positively charged, virus-sized (∼40–80 nm) [F-detergent]2/DNA nanoparticles, as confirmed by zeta potential and agarose gel electrophoresis experiments. These stable nanoparticles consisted likely of monomolecular DNA particles although particles containing a very few copies of DNA could not be excluded. These constructs were obtained following a monomolecular DNA condensation process occurring when DNA is mixed with a cationic detergent at a concentration close to its critical micellar concentration, the resulting complexes being subsequently stabilized upon oxidation of the thiol detergent into its disulfide lipidic dimer ([F-detergent]2) on the DNA matrix. DNA was also shown to be fully protected, not accessible, when condensed into cationic nanoparticles. The surface of these particles was further labeled with an amphiphilic-polyethyleneglycol–folic acid conjugate, as attested by zeta potential measurements. Size measurements and TEM analysis showed nanometric sizes (≤100 nm) for these labeled DNA nanoparticles. Noticeably with respect to their very low size, we found that the cationic, uncoated [F-detergent]2/DNA formulations were efficient “non-specific” transfection agents of KB cells and particularly more efficient than PEI polyplexes. However, though FACS analysis with (un)coated and YOYO-labeled [F-detergent]2/DNA nanoparticles showed specific uptake of the folic acid coated nanoparticles into folate-overexpressing KB cells, transfection experiments relying on luciferase expression measurements did not show any specific improvement of gene expression resulting from such a targeting.