title: Localized gene delivery using antibody tethered adenovirus from polyurethane heart valve cusps and intra-aortic implants.
abstract: The present study investigated a novel approach for gene therapy of heart valve disease and vascular disorders. We formulated and characterized implantable polyurethane films that could also function as gene delivery systems through the surface attachment of replication defective adenoviruses using an anti-adenovirus antibody tethering mechanism. Our hypothesis was that we could achieve site-specific gene delivery to cells interacting with these polyurethane implants, and thereby demonstrate the potential for intravascular devices that could also function as gene delivery platforms for therapeutic vectors. Previous research by our group has demonstrated that polyurethane elastomers can be derivatized post-polymerization through a series of chemical reactions activating the hard segment amide groups with alkyl bromine residues, which can enable a wide variety of subsequent chemical modifications. Furthermore, prior research by our group investigating gene delivery intravascular stents has shown that collagen-coated balloon expandable stents can be configured with anti-adenovirus antibodies via thiol-based chemistry, and can then tether adenoviral vectors at doses that lead to high levels of localized arterial neointima expression, but with virtually no distal spread of vector. Thus, we sought to create two-device configurations for our investigations building on this previous research. (1) Polyurethane films coated with Type I collagen were thiol activated to permit covalent attachment of anti-adenovirus antibodies to enable gene delivery via vector tethering. (2) We also formulated polyurethane films with direct covalent attachment of anti-adenovirus antibodies to polyurethane hard segments derivatized with alkyl-thiol groups, thereby also enabling tethering of replication-defective adenoviruses. Both formulations demonstrated highly localized and efficient transduction in cell culture studies with rat arterial smooth muscle cells. In vivo experiments with collagen-coated polyurethane films investigated an abdominal aorta implant model in pigs using a button configuration that simulated the blood contacting environment of a vascular graft. One week explants of the collagen-coated polyurethane films demonstrated 14.3+/-2.5% of neointimal cells on the surface of the implant transduced with green fluorescent protein - adenovirus (AdGFP) vector loadings of 1 x 10(8) PFU. PCR studies demonstrated no detectable vector DNA in blood or distal organs. Similarly, polyurethane films with direct attachment of antivector antibodies to the surface were used in sheep pulmonary valve leaflet replacement studies, simulating the blood contacting environment of a prosthetic heart valve cusp. Polyurethane films with antibody tethered AdGFP vector (10(8) PFU) demonstrated 25.1+/-5.7% of attached cells transduced in these 1 week studies, with no detectable vector DNA in blood or distal organs. In vivo GFP expression was confirmed with immunohistochemistry. It is concluded that site-specific intravascular delivery of adenoviral vectors for gene therapy can be achieved with polyurethane implants utilizing the antivector antibody tethering mechanism.
authors.affiliation: The Children's Hospital of Philadelphia, Philadelphia, PA 19104-4318, USA.
authors.forename: S J
authors.initials: SJ
authors.lastname: Stachelek
authors.forename: C
authors.initials: C
authors.lastname: Song
authors.forename: I
authors.initials: I
authors.lastname: Alferiev
authors.forename: S
authors.initials: S
authors.lastname: Defelice
authors.forename: X
authors.initials: X
authors.lastname: Cui
authors.forename: J M
authors.initials: JM
authors.lastname: Connolly
authors.forename: R W
authors.initials: RW
authors.lastname: Bianco
authors.forename: R J
authors.initials: RJ
authors.lastname: Levy
mesh_terms: D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D001011:Aorta; D001705:Bioprosthesis; D001807:Blood Vessel Prosthesis; D005260:Female; D015870:Gene Expression; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000008:administration & dosage*; D049452:Green Fluorescent Proteins; D006350:Heart Valve Prosthesis; D008164:Luminescent Proteins / Q000235:genetics; D008297:Male; D011140:Polyurethanes; D019919:Prosthesis Implantation; D012756:Sheep; D013552:Swine; D001616:beta-Galactosidase / Q000235:genetics
chemical_list: D008164:Luminescent Proteins; D011140:Polyurethanes; D049452:Green Fluorescent Proteins; D001616:beta-Galactosidase