(557c) Stent-Mediated Delivery of DNA to Arterial Tissue Using Intravascular Stents Coated with Ultrathin DNA-Containing Polyelectrolyte Multilayers

Thin films and coatings that provide control over the release of DNA from the surfaces of intravascular stents will contribute significantly to the development of localized gene-based approaches to the treatment of cardiovascular disease and complications that can arise from the implantation of these devices. Methods for the localized delivery of antiproliferative small-molecule drugs from the surfaces of stents have already contributed broadly to clinical treatments for coronary artery disease, but the development of analogous gene-based approaches has advanced far more slowly. This is due, in large measure, to a lack of materials that can be used to immobilize and release macromolecular drugs such as DNA from the surfaces of interventional devices. Materials developed to promote the sustained release of small molecule drugs are generally not appropriate for the sustained and surface-mediated release of DNA.

Here, we report a layer-by-layer approach to the fabrication of ultrathin, DNA-containing multilayered polyelectrolyte films (or ?polyelectrolyte multilayers') that permit the surface-mediated and localized release of plasmid DNA from the surfaces of stainless steel intravascular stents. We demonstrated in past studies that polyelectrolyte multilayer coatings ~100 nm thick fabricated from plasmid DNA and hydrolytically degradable poly(beta-amino ester)s could be deposited on the surfaces of intravascular stents. The results of these past studies also demonstrated that these films were able to withstand mechanical challenges associated with stent deployment, and that film-coated stents could be used to sustain the release of DNA upon incubation in physiological buffer for periods of up to four days. Here, we demonstrate that intravascular stents coated with DNA-containing multilayers can be used to localize the release of DNA to arterial tissue and promote transgene expression in the arterial wall when implanted in the arteries of rabbits in vivo.

Characterization of stents coated with films fabricated using fluorescently labeled DNA demonstrated that the films were uniform and continuous after fabrication and that the films were able to withstand mechanical challenges commonly associated with stent deployment (e.g., balloon expansion) without cracking, peeling, or delaminating from the surfaces of the stents. The results of these experiments also demonstrated that forces associated with stent expansion did not influence DNA release profiles when expanded stents were incubated in physiologically relevant media (e.g., PBS, pH = 7.4, 37 °C). When stents coated with fluorescently labeled films were implanted in excised sections of porcine carotid artery (ex vivo), patterns of fluorescence corresponding to the geometries of stent struts and joints were observed after 24 hours of incubating the stented arteries in PBS at 37 °C. These results confirmed the localized transfer of DNA to the arterial wall and provide a basis for the quantitative characterization of the transport and diffusion of DNA. Finally, stents coated with films fabricated using a plasmid encoding enhanced green fluorescent protein (EGFP) were implanted in the femoral and iliac arteries of New Zealand white rabbits. Characterization of thin slices of stented arteries excised 48 hours after implantation using indirect immunohistochemical staining revealed localized expression of EGFP at the site of stent implantation. Levels of expression were highest in the medial layers of the arterial tissue, and initial characterization of these samples using hematoxylin/eosin staining did not reveal a significant inflammatory response over this initial 48-hour time period.

When combined, the results of these experiments demonstrate (i) that these DNA-containing polyelectrolyte multilayers are able to withstand the mechanical and biological challenges (e.g., balloon expansion and prolonged contact with blood and other fluids) associated with stent expansion and deployment, (ii) that DNA is released in a form that is able to transfect arterial tissue, and (iii) that DNA is transported to or disseminated over relatively large distances during the course of these in vivo experiments. Additional quantitative characterization of DNA dissemination and in vivo gene expression in a rabbit model will be presented, and materials-based approaches to tuning both the amount of DNA released and the kinetics with which it is released will be discussed.