Abstract

Cationic liposome - DNA (CL-DNA) complexes (or lipoplexes) are among the most promising materials for delivery of genes into mammalian cells for gene therapy applications. These complexes are found to remain stable in monovalent salt solutions until ca. 500 mM. Long term circulation in blood requires a repulsive polymer coat to prevent removal by the immune system. A viable way of obtaining coated CL-DNA particles is incorporation of polyethylene glycol lipids (PEG-lipids) before mixing with DNA (Ewert et al. Topics Curr. Chem. 2010, 296, 191-226). Here we show that pegylated complex stability in salt depends on the preparation method of the particles. CL-DNA particles with 5 mol% of PEG 2000 form stable complexes with well defined lamellar peaks in water. The structure remains stable after addition of a 150 mM NaCl or DMEM medium (both with ionic strength near physiological conditions). Conversely, if complexes are initially formed in salt or DMEM medium, the structure is less stable with small-angle X-ray scattering (SAXS) showing the coexistence of narrow and broad lamellar peaks. If the PEG amount in the bilayers is increased to 10%, the particles formed in water continue to be well defined, but the ones formed in salt and DMEM exhibit small domain sizes as revealed in SAXS experiments. Cryo-TEM images also show onion-like structures with very few layers. Since pegylated CL-DNAs are widely used in gene therapy applications, these results are of significance since it is reasonable to expect that the degree of compactness of these particles (which depends on the preparation method) will strongly affect complex stability and transfection efficiency.Funded by: NIH GM59288. BFBS acknowledges funding from the European Community under a Marie Curie IOF grant (n252701).

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