Soft hydrogels interpenetrating silicone--A polymer network for drug-releasing medical devices.

Materials for the next generation of medical devices will require not only the mechanical stability of current devices, but must also possess other properties such as sustained release of drugs in a controlled manner over a prolonged period of time. This work focuses on creating such a sophisticated material by forming an interpenetrating polymer network (IPN) material through modification of silicone elastomers with a poly(2-hydroxyethyl methacrylate) (PHEMA)-based hydrogel. IPN materials with a PHEMA content in the range of 13%-38% (w/w) were synthesized by using carbon dioxide-based solvent mixtures under high pressure. These IPNs were characterized with regard to microstructure as well as ability of the hydrogel to form a surface-connected hydrophilic carrier network inside the silicone. A critical limit for hydrogel connectivity was found both via simulation and by visualization of water uptake in approximately 25% (w/w) PHEMA, indicating that entrapment of gel occurs at low gel concentrations. The optimized IPN material was loaded with the antibiotic ciprofloxacin, and the resulting drug release was shown to inhibit bacterial growth when placed on agar, thus demonstrating the potential of this IPN material for future applications in drug-releasing medical devices.

In vitro release studies of insulin from lipid implants in solution and in a hydrogel matrix mimicking the subcutis.

Widely accepted in vitro methodologies for sustained release parenteral drug formulations remain to be established. Hydrogels have been proposed as a release matrix more closely resembling the in vivo conditions for formulations intended for subcutaneous administration. The perspective of the current work was to investigate the feasibility of developing UV imaging-based in vitro methods enabling visualization and characterization of drug release and transport of protein therapeutics intended for subcutaneous administration. Specifically, the objectives were to prepare lipid implants providing sustained release of the model protein insulin and investigate the release into 0.5% (w/v) agarose hydrogels, pH7.40, using UV imaging- and a gel sampling-based release testing method. These results were compared to insulin release into well agitated buffer solution. Irrespective of the applied in vitro release method, the insulin release from Sterotex implants with a drug load of 20% (w/w) was faster as compared to the release from implants with a load of 10% (w/w), most likely due to the higher porosity of the implants with increasing drug load. Insulin release from 10% (w/w) implants into agitated solution was faster as compared to release into agarose hydrogel. This was ascribed to the additional mass transfer resistance provided by the agarose hydrogel. Interestingly, the release profiles of insulin from implants with an initial drug load of 20% (w/w) obtained by the three in vitro methods were relatively similar. The gel-based methods, in particular UV imaging, enable monitoring local drug concentrations in the vicinity of the implant over time thereby facilitating assessment of, e.g., sink conditions. The study highlights that the selection of the in vitro release method should take into account various factors including mass transport, drug stability, data analysis and simplicity of the methodology.

Real-time UV imaging identifies the role of pH in insulin dissolution behavior in hydrogel-based subcutaneous tissue surrogate.

For parenteral biopharmaceuticals, subcutaneous diffusion and, in the case of solid implants or suspensions, dissolution may govern the clinical profile of the drug product. Insight into the dissolution and diffusion processes of biopharmaceuticals after parenteral administration is fundamental in the development of new protein drug formulations. Using insulin as a model compound, the aim of this work was to develop a UV imaging-based method to study the real-time dissolution and diffusion behavior of solid protein drugs under stagnant conditions in a hydrogel matrix mimicking the subcutaneous tissue. Dissolution of proteins and peptides is a complex phenomenon as it may be coupled to the complicated acid base properties of these substances. UV imaging allowed the real-time dissolution and diffusion processes of insulin at different pH values and of different insulins to be studied. Dissolution rates were obtained, and the quantitative performance of the developed UV imaging method was verified. It was shown that the UV imaging dissolution method was able to differentiate between the behavior of different insulins and that human insulin dissolution was highly dependent on pH. pH effects in the microenvironment of the human insulin compacts at pH 7.40 and 3.00 were observed by UV-Vis imaging, explaining the different dissolution kinetics of human insulin at pH 7.40 and 3.00 as compared to pH 5.40. In conclusion, UV-Vis imaging may be a useful tool for studying dissolution, diffusion and pH effects in the vicinity of solid protein drug in a hydrogel matrix with the aim of achieving a better understanding of in vivo dissolution processes.

Insulin diffusion and self-association characterized by real-time UV imaging and Taylor dispersion analysis.

Assessment of release kinetics of subcutaneously administered protein therapeutics remains a complex challenge. In vitro methods capable of visualizing and characterizing drug transport properties, in the formulation as well as surrounding subcutaneous tissue environment, are desirable in drug development. Diffusion is a key process in drug release and transport. Thus, our objective was to develop a UV imaging in vitro method for direct visualization and characterization of insulin diffusivity and self-association behavior. Agarose hydrogels were used for mimicking subcutaneous tissue. Diffusivity, self-association, and apparent size of insulin were further characterized by Taylor dispersion analysis, size exclusion chromatography, and dynamic light scattering. At low insulin concentrations and pH 3.0, the hydrodynamic radius of insulin was determined by Taylor dispersion analysis to 1.5±0.1nm, corresponding to the size of insulin monomer. Increasing concentration and pH to 1mM and pH 7.4, respectively, favoring insulin hexamers, increased the insulin hydrodynamic radius to 3.0±0.1nm. The UV imaging method developed was adequately sensitive to identify and characterize, in terms of diffusion coefficients, the changes in insulin transport in hydrogel due to pH and concentration changes. In conclusion, UV imaging allowed insulin diffusion in hydrogel matrixes to be studied in real-time, and showed that insulin self-association properties were reflected in the diffusion behavior. UV imaging is a useful tool for characterization of the influence of environmental conditions on protein mass transport. Hydrogels combined with UV imaging may be of utility for in vitro testing of protein therapeutics.

Viscosity of high concentration protein formulations of monoclonal antibodies of the IgG1 and IgG4 subclass - prediction of viscosity through protein-protein interaction measurements.

The purpose of this work was to explore the relation between protein-protein interactions (PPIs) and solution viscosity at high protein concentration using three monoclonal antibodies (mAbs), two of the IgG4 subclass and one of the IgG1 subclass. A range of methods was used to quantify the PPI either at low concentration (interaction parameter (kD) obtained from dynamic light scattering, DLS) or at high concentration (solution storage modulus (G') from ultrasonic shear rheology). We also developed a novel method for the determination of PPI using the apparent radius of the protein at either low or high protein concentration determined using DLS. The PPI measurements were correlated with solution viscosity (measured by DLS using polystyrene nanospheres and ultrasonic shear rheology) as a function of pH (4-9) and ionic strength (10, 50 and 150 mM). Our measurements showed that the highest solution viscosity was observed under conditions with the most negative kD, the highest apparent radius and the lowest net charge. An increase in ionic strength resulted in a change in the nature of the PPI at low pH from repulsive to attractive. In the neutral to alkaline pH region the mAbs behaved differently with respect to increase in ionic strength. Two mAbs (A and B) showed little or no effect of increasing ionic strength, whereas mAb-C showed a remarkable decrease in attractive PPI and viscosity. Previous studies have mainly investigated mAbs of the IgG1 and IgG2 subclass. We show here, for the first time, that mAbs of the IgG4 subclass behave similar as the other subclasses. By comparison of the three tested mAbs with mAbs investigated in other studies a clear linear trend emerges between the pH of strongest attractive PPI and highest solution viscosity. The determination of PPI using either kD or apparent radius is thus a useful prediction tool in the determination of solution conditions that favors low solution viscosity at high protein concentration of therapeutically used mAb molecules. The novel methodology using apparent radius is a simple and rapid alternative to determine relative PPI directly under formulation conditions. The method can potentially serve as a high-throughput screening tool in formulation development.

Delivery of dermatan sulfate from polyelectrolyte complex-containing alginate composite microspheres for tissue regeneration.

Dermatan sulfate (DS) is a glycosaminoglycan (GAG) with a great potential as a new therapeutic agent in tissue engineering. The aim of the present study was to investigate the formation of polyelectrolyte complexes (PECs) between chitosan and dermatan sulfate (CS/DS) and delivery of DS from PEC-containing alginate/chitosan/dermatan sulfate (Alg/CS/DS) microspheres for application in tissue regeneration. The CS/DS complexes were initially formed at different conditions including varying CS/DS ratio (positive/negative charge ratio), buffer, and pH. The obtained CS/DS complexes exhibited stronger electrostatic interaction, smaller complex size, and more stable colloidal structure when chitosan was in large excess (CS/DS 3:1) and prepared at pH 3.5 as compared to pH 5 using acetate buffer. The CS/DS complexes were subsequently incorporated into an alginate matrix by spray drying to form Alg/CS/DS composite microspheres with a DS encapsulation efficiency of 90-95%. The excessive CS induced a higher level of sustained DS release into Tris buffer (pH 7.4) from the microspheres formulated at pH 3.5; however, the amount of CS did not have a significant effect on the release from the microspheres formulated at pH 5. Significant cell proliferation was stimulated by the DS released from the microspheres in vitro. The present results provide a promising drug delivery strategy using PECs for sustained release of DS from microspheres intended for site-specific drug delivery and ultimately for use in tissue engineering.