Multiple and time-scheduled in situ DNA delivery mediated by β-cyclodextrin embedded in a polyelectrolyte multilayer

Abstract

The basic premise of gene therapy is that genes can be used to produce in situ therapeutic proteins. The controlled delivery of DNA complexes from biomaterials offers the potential to enhance gene transfer by maintaining an elevated concentration of DNA within the cellular microenvironment. Immobilization of the DNA to the substrate to which cells adhere maintains the DNA in the cell microenvironment for subsequent cellular internalization. Here, layer-by-layer (LBL) films made from poly(L-glutamic acid) (PLGA) and poly(L-lysine) (PLL) containing DNA were built in the presence of charged cyclodextrins. The biological activities of these polyelectrolyte films were tested by means of induced production of a specific protein in the nucleus or in the cytoplasm by cells in contact with the films. This type of coating offers the possibility for either simultaneous or sequential interfacial delivery of different DNA molecules aimed at cell transfection. These results open the route to numerous potential applications in patch vaccination, for example.gene delivery ͉ layer-by-layer films ͉ transfection T he basic premise of somatic gene therapy is that genes can be used to cause in vivo production of therapeutic proteins. Controlled and efficient gene delivery has implications in many fields ranging from basic science to clinical medicine. Current strategies to enhance gene delivery involve the complexation of DNA with cationic polymers or lipids delivery. Cationic polymers or lipids can self-assemble with DNA to form particles that are capable of being endocytosed by cells (1). These complexes reduce effective size and cellular degradation of DNA (2) and are often delivered as a bolus, added to culture wells in vitro. Bolus delivery of these complexes can be hindered by mass transport limitations or deactivation processes, such as degradation or aggregation. For example, in vitro studies have estimated that bolus addition of complexes to the culture media results in internalization of only 20% of the DNA added (3). These limitations motivated the development of alternative delivery strategies.The controlled delivery of DNA complexes from biomaterials offers the potential to enhance gene transfer by maintaining an elevated concentration of DNA within the cellular microenvironment (4). DNA delivery systems from biomaterials are designed to maintain elevated concentrations locally by supplying DNA to balance the loss by degradation. The continued presence of the DNA during cell division seems to facilitate entry into the nucleus (5). In recent years, considerable effort has been devoted to the design and the controlled fabrication of structured materials with functional properties (6). The layer-by-layer (LBL) buildup of polyelectrolyte films from oppositely charged polyelectrolytes (7) offers opportunities for the preparation of functionalized biomaterial coatings. This technique allows the preparation of supramolecular nano-architectures (8-13) exhibiting specific properties in terms of control of cell activation (8-11) ...

Multilayered Films Fabricated from Combinations of Degradable Polyamines: Tunable Erosion and Release of Anionic Polyelectrolytes

“…Ongoing investigations in our laboratory are focused on controlling the architectures of these films with a view toward fabricating assemblies that permit the staged release of multiple film components. 33,38 We demonstrate here, however, that the incorporation of multiple different layers of polyamines 1-3 into multilayered films can also provide a straightforward and highly tunable approach to exerting control over film erosion and the release of SPS. Figure 4 shows a plot of ellipsometric thickness versus the number of bilayers deposited during the fabrication of a (1/SPS) 4 (2/SPS) 4 film and a (2/SPS) 4 (1/SPS) 4 film (fabricated using the general approach described above).…”

Layer-by-Layer-Assembled Multilayer Films for Transcutaneous Drug and Vaccine Delivery

“…[18][19][20][21] Drug-loaded degradable multilayers have been explored for the sustained release of small-molecule antibiotics, protein therapeutics, or plasmid DNA. 3,7,18,19,[21][22][23][24] The mild aqueous conditions for encapsulating molecules into multilayer films preserves the bioactivity of fragile biomolecules such as proteins and nucleic acids. 21,22,25 By employing degradable polyelectrolytes as building blocks, the ability to tune the degradation kinetics of multilayer assemblies has been demonstrated and used to control the release kinetics of compounds embedded in these films.…”

Active multilayered capsules for in vivo bone formation

“…Therapeutics and biomolecules including peptides, proteins, and nucleic acid have been embedded in PEM films, which offer new opportunities for the preparation of functionalized bioactive coatings (19)(20)(21). These supramolecular nanoarchitectures can be designed to exhibit specific properties, including control of cell activation, inflammation (22,23) and localized drug, growth factor or nucleic acid delivery (24,25). The embedded biomolecules, which are either chemically bound to polyelectrolytes or physically adsorbed, have been shown to retain their biological activity in many studies (22)(23)(24)(25)(26).…”

“…These supramolecular nanoarchitectures can be designed to exhibit specific properties, including control of cell activation, inflammation (22,23) and localized drug, growth factor or nucleic acid delivery (24,25). The embedded biomolecules, which are either chemically bound to polyelectrolytes or physically adsorbed, have been shown to retain their biological activity in many studies (22)(23)(24)(25)(26). Bioactive proteins can be directly integrated in the architecture without any covalent bonding with a polyelectrolyte and keep a secondary structure close to their native form.…”