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The development of thin films and coatings that control the release of DNA from the surfaces of materials could have a significant impact on localized approaches to gene therapy. Here, we report multilayered polyelectrolyte assemblies that sustain the release of functional plasmid DNA from the surfaces of model substrates under physiological conditions. Multilayered assemblies consisting of alternating layers of plasmid DNA encoding for enhanced green fluorescent protein (EGFP) and a synthetic degradable polyamine were deposited on planar silicon and quartz substrates using a layer-by-layer fabrication process. Film growth was monitored by ellipsometry and UV spectrophotometry and correlated linearly with the number of polymer and plasmid layers deposited. In general, the thickness of deposited layers was found to be a function of both the pH and the ionic strength of the polyelectrolyte solutions used. Films up to 100 nm thick were investigated in this study. These assemblies erode gradually upon incubation in phosphate-buffered saline at 37 degrees C, as determined by ellipsometry and UV spectrophotometry, and sustain the release of incorporated plasmid into the incubation medium for a period of up to 30 h. Characterization of the released plasmid by agarose gel electrophoresis revealed that the DNA was released in a relaxed, open circular, rather than supercoiled, topology; subsequent cell transfection experiments demonstrated that the released plasmid is transcriptionally viable and promotes the expression of EGFP in the COS-7 cell line. These layered materials could represent an approach to the controlled administration of one or more functional DNA constructs from the surfaces of biomedical materials and devices.

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Methods that permit the deposition or assembly of reactive polymer films on topologically complex substrates are useful for the patterning or chemical modification of surfaces of interest in a broad range of applications. [1][2][3][4][5] Here, we report a layer-by-layer approach to the assembly of covalently-crosslinked ultrathin films that makes use of fast and efficient 'click'-type interfacial reactions between poly(2-alkenyl azlactone)s and appropriately functionalized polyamines. In contrast to conventional, aqueous methods for the layer-by-layer fabrication of multilayered assemblies composed of polyelectrolytes, [6][7][8][9] fabrication of these materials occurs in organic solvents and is driven by rapid interfacial formation of covalent bonds during assembly. These methods permit precise, nanometer-scale control over the thicknesses and compositions of covalently crosslinked thin films. In addition, we demonstrate that it is possible to chemically tailor the properties of surfaces coated with these ultrathin films post-fabrication by exploiting the accessibility and reactivity of residual 'spring-loaded' azlactone functionality. These results suggest the basis of methods for the post-fabrication modification of curved or topologically complex surfaces coated with multilayered films and the patterning or passivation of surfaces with chemical or biological functionality of interest in the contexts of catalysis, medicine, and other areas of biotechnology. Aqueous methods for the layer-by-layer deposition of oppositely charged polyelectrolytes on surfaces are used widely for the bottom-up assembly of nanostructured polymer films. [6][7][8][9] These methods generally take advantage of multivalent weak interactions (e.g., electrostatic or hydrogen bonding interactions) between polyelectrolytes and oppositely charged surfaces and allow precise control over the thicknesses, compositions, and morphologies of thin films fabricated from a broad range of water-soluble polymers.

Materials that permit control over the release of DNA from the surfaces of topologically complex implantable devices, such as intravascular stents, could contribute to the development of new approaches to the localized delivery of DNA. We report the fabrication of ultrathin, multilayered polyelectrolyte films that permit both the immobilization and controlled release of plasmid DNA from the surfaces of stainless steel intravascular stents. Our approach makes use of an aqueous-based, layer-by-layer method for the assembly of nanostructured thin films consisting of alternating layers of plasmid DNA and a hydrolytically degradable polyamine. Characterization of coated stents using scanning electron microscopy (SEM) demonstrated that stents were coated uniformly with an ultrathin film ca. 120 nm thick that adhered conformally to the surfaces of stent struts. These ultrathin films did not crack, peel, or delaminate substantially from the surface after exposure to a range of mechanical challenges representative of those encountered during stent deployment (e.g., balloon expansion). Stents coated with eight bilayers of degradable polyamine and a plasmid encoding enhanced green fluorescent protein sustained the release of DNA into solution for up to four days when incubated in phosphate buffered saline at 37 °C, and coated stents were capable of mediating the expression of EGFP in a mammalian cell line without the aid of additional transfection agents. The approach reported here could, with further development, contribute to the development of localized gene-based approaches to the treatment of cardiovascular diseases or related conditions.