Multilayered Thin Films from Boronic Acid-Functional Poly(amido amine)s as Drug-Releasing Surfaces

Purpose To evaluate the potential of poly(amido amine)-based multilayered thin films in surface mediated drug release. Methods Multilayered thin films were prepared from copolymers of phenylboronic acid-functional poly(amido amine)s and chondroitin sulfate (ChS) in the presence of Alizarin Red S (ARS) as a reporter molecule. Multilayer buildup and ARS incorporation were evaluated with UV–vis spectroscopy. Glucose responsiveness of the multilayers was investigated. Finally, cellular uptake of ARS by COS-7 cells grown on the films was assessed. Results Multilayers based on alcohol containing polymers (ABOL-BA-PAA#ChS + ARS) displayed higher ARS incorporation than multilayers based on amine-containing polymers (DAB-BA-PAA#ChS + ARS). At physiological pH, a swift initial release of up to ~40% of the ARS content was observed during the first 12 h of incubation, followed by a much slower, gradual release of ARS. The multilayers were further evaluated by culturing COS-7 cells on top of multilayer-coated well plates. Cellular uptake of the fluorescent ARS-boronate ester was quantified through flow cytometry, and a maximum uptake of up to 30% was observed. Confocal microscopy confirmed the presence of ARS-boronate ester-containing particles in the nuclei of cells. Conclusions The investigated multilayered thin films are effective in surface-mediated delivery of the model compound ARS. These multilayered surfaces are promising as drug-releasing delivery surface for coating stents, prostheses, and other implants

Multilayered Thin Films from Boronic Acid-Functional Poly(amido amine)s

Purpose To investigate the properties of phenylboronic acid-functional poly(amido amine) polymers (BA-PAA) in forming multilayered thin films with poly(vinyl alcohol) (PVA) and chondroitin sulfate (ChS), and to evaluate their compatibility with COS-7 cells. Methods Copolymers of phenylboronic acid-functional poly(amido amine)s, differing in the content of primary amine (DAB-BA-PAA) or alcohol (ABOL-BA-PAA) side groups, were synthesized and applied in the formation of multilayers with PVA and ChS. Biocompatibility of the resulting films was evaluated through cell culture experiments with COS-7 cells grown on the films. Results PVA-based multilayers were thin, reaching ~100 nm at 10 bilayers, whereas ChS-based multilayers were thick, reaching ~600 nm at the same number of bilayers. All of the multilayers are stable under physiological conditions in vitro and are responsive to reducing agents, owing to the presence of disulfide bonds in the polymers. PVA-based films were demonstrated to be responsive to glucose at physiological pH at the investigated glucose concentrations (10–100 mM). The multilayered films displayed biocompatibility in cell culture experiments, promoting attachment and proliferation of COS-7 cells. Conclusions Responsive thin films based on boronic acid functional poly(amido amine)s are promising biocompatible materials for biomedical applications, such as drug releasing surfaces on stents or implants

Enhancing Cellular Internalization of Single-Chain Polymer Nanoparticles via Polyplex Formation

Intracellular delivery of nanoparticles is crucial in nanomedicine to reach optimal delivery of therapeutics and imaging agents. Single-chain polymer nanoparticles (SCNPs) are an interesting class of nanoparticles due to their unique site range of 5–20 nm. The intracellular delivery of SCNPs can be enhanced by using delivery agents. Here, a positive polymer is used to form polyplexes with SCNPs, similar to the strategy of protein and gene delivery. The size and surface charge of the polyplexes were evaluated. The cellular uptake showed rapid uptake of SCNPs via polyplex formation, and the cytosolic delivery of the SCNPs was presented by confocal microscopy. The ability of SCNPs to act as nanocarriers was further explored by conjugation of doxorubicin

Multilayered thin films from poly(amido amine)s for controlled delivery

Layer-by-layer (LbL) assembly is a versatile and economical method to coat existing biomaterials with multilayered thin films for added functionality. The technique requires at least two complementary macromolecules to be deposited alternately onto a surface, forming thin multiple layers with properties that are customizable through the deposition conditions such as pH, temperature, and ionic strength. As a class of synthetic peptidomimetic polymers that have shown significant promise as therapeutic delivery agent through the ability to interact with DNA, proteins, carbohydrates, and small drug molecules, poly(amido amine)s (PAA) are promising multilayer components. This thesis describes a series of reports on the use of PAA as a component for LbL assembly. The reports describe the study on structure-function relationship of the various PAAs to the build-up profiles, and properties and functionality of the resulting multilayers. The report starts with the pairing of PAAs with DNA to yield multilayer-coated surfaces that can facilitate cell transfection. The optimization of such surface-mediated cell transfection and its successful demonstration on 2D and 3D tissue engineering scaffolds from several common biomaterials are described in the last Chapter. The PAA-multilayers were also studied as drug-eluting surfaces. For this purpose, boronic ester formation was utilized to provide multi-responsive interactions with small drug compounds. In one Chapter, boronic acid-functionalized PAAs were paired with chondroitin sulfate, a natural biopolymer, to entrap alizarin red S, a model for catechol drugs. The multilayer-coated surfaces successfully promote intracellular uptake of polymer-complexed alizarin red S in vitro. In another Chapter, PAA was paired with poly(vinyl alcohol), an FDA-approved biomaterial, to entrap bortezomib, a drug for multiple myeloma. Depending on how the multilayers were fabricated, the therapeutic efficacy could be tuned to differently affect target cells depending on their spatial location. The results in this thesis indicate great promise for poly(amido amine)-based multilayered thin films

Multilayered Thin Films from Poly(amido amine)s and DNA

Dip-coated multilayered thin films of poly(amido amine)s (PAAs) and DNA have been developed to provide surfaces with cell-transfecting capabilities. Three types of PAAs, differing in side chain functional groups, were synthesized and characterized for their properties in forming multilayered structures with ultrasonicated calf thymus DNA (CTDNA) as model DNA. All three polymers display a multilayer build-up in linear profiles as demonstrated by UV spectroscopy. More highly charged side chains were found to provide the lowest deposition of DNA. Surface profiles of the obtained films were investigated by atomic force microscopy (AFM) and static water contact angle measurements to reveal complete surface coverage after at least four layer pair depositions, where alternating patterns of surface profiles were observed depending on whether the cationic polymer or the anionic DNA layer was on top. The stability of the formed surfaces was investigated in vitro under physiological and reductive conditions. Owing to the presence of disulfide bonds in the PAA main chain, the films were readily degraded in the presence of 1 mM of DTT in vitro. Under non-reductive physiological conditions, two of the thicker films underwent thermodynamic rearrangement, which resulted in release of approximately half of the incorporated material within 1 h, which was caused by the physiological salt concentration. Further, this unpacking phenomenon proved useful in transfecting COS-7 cells seeded on top of these multilayers containing functional plasmid DNA encoding for green fluorescence protein (GFP). Two out of the three different multilayers facilitated good COS-7 cell attachment, proliferation, and transfection in vitro within 2 days of culture. Fluorescence staining further revealed the presence of DNA-containing released film material among cultured cells. The present work demonstrates the possibility of coating surfaces with thin films that are conveniently adjustable in thickness and amount of active agent to provide cell-transfecting functionality. In this manner transfection can be achieved by simply culturing cells on a multilayer-coated surface in their optimal culture condition (in the presence of serum) and without the need of removing the transfection agent to avoid cytotoxicity

Enhancing Cellular Internalization of Single-Chain Polymer Nanoparticles via Polyplex Formation

Intracellular delivery of nanoparticles is crucial in nanomedicine to reach optimal delivery of therapeutics and imaging agents. Single-chain polymer nanoparticles (SCNPs) are an interesting class of nanoparticles due to their unique site range of 5-20 nm. The intracellular delivery of SCNPs can be enhanced by using delivery agents. Here, a positive polymer is used to form polyplexes with SCNPs, similar to the strategy of protein and gene delivery. The size and surface charge of the polyplexes were evaluated. The cellular uptake showed rapid uptake of SCNPs via polyplex formation, and the cytosolic delivery of the SCNPs was presented by confocal microscopy. The ability of SCNPs to act as nanocarriers was further explored by conjugation of doxorubicin

Responsive Layer-by-Layer Films

Layer-by-layer (LbL) assembly is the process of building functional multilayered thin films. Owing to its highly modular and versatile nature, it has been used to coat a wide variety of different surfaces, including inorganic substrates, membranes, implants, nanoparticles and even living cells. It provides ways to induce responsiveness through both the chemically engineered macromolecular components, and the way the multilayers are built up. For example, assembly degradability can be adjusted by using degradable polymers or crosslinkers, while physical properties can be altered through the use of additives, or by the assembly method. This chapter is dedicated to LbL fabrication-specific responsiveness, and to recent developments in multilayers composed of specifically tailored polymers. It further focuses on chemically and biologically responsive LbL systems, with main applications in the biomedical field. The introduction covers general aspects of LbL assembly and physicochemical aspects of the assemblies. The second part describes physicochemical aspects in more detail with examples on how variation in deposition conditions, e.g. pH and ionic strength, as well as specific additives, induce responsiveness to the resulting multilayers. It also highlights several reports on compartmentalized multilayered coating fabrication for tunable disassembly or release of incorporated materials. The third part describes multilayers fabricated with chemically tailored biomaterials for different chemical and biological responsiveness. More specifically, multilayer disassembly can be triggered by the inherent responsiveness of one of the multilayer components, through incorporation of labile bonds that respond to specific external triggers, or through disruption of the interlayer interaction between two or more multilayer components