Association and dissociation characteristics of polymer/DNA complexes used for gene delivery

The DNA association/dissociation properties of water-soluble cationic methacrylate polymers with closely related structures (poly(2-dimethylamino)ethyl methacrylate) [p(DMAEMA)], poly(2-(trimethylamino)ethyl methacrylate chloride) [p(TMAEMA)]) and the frequently used transfectant poly(L-lysine) were studied to gain a better insight into their transfection characteristics.

Association of DNA with different polymers and dissociation of the complexes, achieved by adding an excess of anionic polymers or salt, were studied by using spectroscopic techniques (fluorescence, circular dichroism (CD)), agarose gel electrophoresis and an enzymatic assay (DNase I treatment). The transfection efficiency of the polyplexes was evaluated in tissue culture with OVCAR-3 cells.

Plasmid DNA complexed with either poly(L-lysine) or p(DMAEMA) was protected against digestion by DNase I. Fluorescence and CD spectroscopy as well as gel electrophoresis revealed that p(DMAEMA) with a relatively high molecular weight and poly(L-lysine) have similar DNA association/dissociation characteristics. Therefore, differences in transfection potential of the polyplexes cannot be ascribed to differences in binding characteristics, but are probably caused by other factors. As compared with the other polymers, p(TMAEMA) has a high affinity for DNA as was concluded from the observation that poly(aspartic acid) was unable to fully dissociate complexes containing this polymer. This fact might very well explain the low transfection efficiency of these polyplexes. p(DMAEMA) with a relatively low molecular weight probably has a low affinity for DNA, which might explain both the formation of DNA aggregates (psi-DNA) and the low transfection potential obtained when using this polymer.

DNA association/dissociation studies shed light on the preferred characteristics of polymeric transfectants.