Injectable Anhydrous Poly(ethylene glycol) Polymer Liquids Form Protein Depot for Extended Controlled Release Applications Via Rapid In Situ Gelation.

Water-free preparation of protein delivery systems has the potential to overcome the limitations of hydrogel depot systems such as off-target reactions, functional group hydrolysis, and limited loading capacity. However, a major roadblock in the development and use of these systems is administration as implantation is often required. In this study, we developed a biodegradable and water-free injectable protein delivery system via inverse electron demand Diels-Alder reaction between norbornene- and tetrazine-functionalized four-armed poly(ethylene glycol) macromonomers. 1:1 mixtures of these precursors gelled rapidly in situ, taking less than 11 s to reach their gelation point. Methyl substitution of tetrazine slowed the gelation time and increased the cross-linking density, whereas oxygen incorporation into norbornene changed the mechanical properties. Introduction of hydrolytically cleavable groups enabled biodegradability. Using phenyl carbamate and phenyl carbonate ester groups, we could tune the stability. Controlled release of the protein surrogate glucose oxidase was achieved over a period of 500 days. The novel preparation method presented here is a promising step toward the development of water-free injectable protein depots for controlled drug delivery.

Investigation of the Impact of Hydrolytically Cleavable Groups on the Stability of Poly(ethylene glycol) Based Hydrogels Cross-Linked via the Inverse Electron Demand Diels-Alder (iEDDA) Reaction.

Eight-armed poly(ethylene glycol) (PEG) hydrogels cross-linked via inverse electron demand Diels-Alder reaction between norbornene and tetrazine groups are promising materials for long-term protein delivery. While a controlled release over 265 days is achieved for 15% w/v hydrogels in the previous study, the material shows high stability over 500 days despite having cleavable ester linkages between the PEG macromonomers and their functionalities. In this study, the hydrolyzable ester linkers in the PEG-norbornene precursor structure are exchanged to reduce the degradation time. To this end, 3,6-epoxy-1,2,3,6-tetrahydrophthalimide, phenyl carbamate, carbonate ester, and phenyl carbonate ester are introduced as degradable functional groups. Oscillatory shear experiments reveal that they are not affected the in situ gelation. All hydrogel types have gel points of less than 20 s even at a low polymer concentration of 5% w/v. Hydrogels with varying polymer concentrations have similar mesh sizes, all of which fell in the range of 4-12 nm. The inclusion of phenyl carbonate ester accelerates degradation considerably, with complete dissolution of 15% w/v hydrogels after 302 days of incubation in phosphate buffer (pH 7.4). Controlled release of 150 kDa fluorescein isothiocyanate-dextran over a period of at least 150 days is achieved with 15% w/v hydrogels.

Airway Delivery of Hydrogel-Encapsulated Niclosamide for the Treatment of Inflammatory Airway Disease.

Repurposing of the anthelminthic drug niclosamide was proposed as an effective treatment for inflammatory airway diseases such as asthma, cystic fibrosis, and chronic obstructive pulmonary disease. Niclosamide may also be effective for the treatment of viral respiratory infections, such as SARS-CoV-2, respiratory syncytial virus, and influenza. While systemic application of niclosamide may lead to unwanted side effects, local administration via aerosol may circumvent these problems, particularly when the drug is encapsulated into small polyethylene glycol (PEG) hydrospheres. In the present study, we examined whether PEG-encapsulated niclosamide inhibits the production of mucus and affects the pro-inflammatory mediator CLCA1 in mouse airways in vivo, while effects on mucociliary clearance were assessed in excised mouse tracheas. The potential of encapsulated niclosamide to inhibit TMEM16A whole-cell Cl- currents and intracellular Ca2+ signalling was assessed in airway epithelial cells in vitro. We achieved encapsulation of niclosamide in PEG-microspheres and PEG-nanospheres (Niclo-spheres). When applied to asthmatic mice via intratracheal instillation, Niclo-spheres strongly attenuated overproduction of mucus, inhibited secretion of the major proinflammatory mediator CLCA1, and improved mucociliary clearance in tracheas ex vivo. These effects were comparable for niclosamide encapsulated in PEG-nanospheres and PEG-microspheres. Niclo-spheres inhibited the Ca2+ activated Cl- channel TMEM16A and attenuated mucus production in CFBE and Calu-3 human airway epithelial cells. Both inhibitory effects were explained by a pronounced inhibition of intracellular Ca2+ signals. The data indicate that poorly dissolvable compounds such as niclosamide can be encapsulated in PEG-microspheres/nanospheres and deposited locally on the airway epithelium as encapsulated drugs, which may be advantageous over systemic application.

In Situ Forming iEDDA Hydrogels with Tunable Gelation Time Release High-Molecular Weight Proteins in a Controlled Manner over an Extended Time.

Off-target interactions between reactive hydrogel moieties and drug cargo as well as slow reaction kinetics and the absence of controlled protein release over an extended period of time are major drawbacks of chemically cross-linked hydrogels for biomedical applications. In this study, the inverse electron demand Diels-Alder (iEDDA) reaction between norbornene- and tetrazine-functionalized eight-armed poly(ethylene glycol) (PEG) macromonomers was used to overcome these obstacles. Oscillatory shear experiments revealed that the gel point of a 15% (w/v) eight-armed PEG hydrogel with a molecular weight of 10 kDa was less than 15 s, suggesting the potential for fast in situ gelation. However, the high-speed reaction kinetics result in a risk of premature gel formation that complicates the injection process. Therefore, we investigated the effect of polymer concentration, temperature, and chemical structure on the gelation time. The cross-linking reaction was further characterized regarding bioorthogonality. Only 11% of the model protein lysozyme was found to be PEGylated by the iEDDA reaction, whereas 51% interacted with the classical Diels-Alder reaction. After determination of the mesh size, fluorescein isothiocyanate-dextran was used to examine the release behavior of the hydrogels. When glucose oxidase was embedded into 15% (w/v) hydrogels, a controlled release over more than 250 days was achieved. Overall, the PEG-based hydrogels cross-linked via the fast iEDDA reaction represent a promising material for the long-term administration of biologics.

Hydrogel microspheres evading alveolar macrophages for sustained pulmonary protein delivery.

Pulmonary delivery is a highly attractive alternative to injections for biologics such as therapeutic proteins. However, bioavailabilities generally suffer from the presence of phagocytic cells that clear particulate matter entering the lung. In this study, microgel particles were developed using an all-aqueous two-phase system approach and evaluated for their efficacy as an inhalable controlled release system. Norbornene- and thiol-modified four- and eight-armed poly (ethylene glycol) with an average molecular mass of 10,000 Da were prepared as macromonomers for microgel formation. Emulsions of precursor solution droplets containing macromonomers and Irgacure 2959 as photocatalyst were prepared in a dextran solution. Irradiation with UV light was used to covalently crosslink the droplets by triggering the thiol-ene reaction. The resulting microgels were processed to dry powder inhaler formulations, and respirable aerodynamic sizes were assessed in vitro. Microgels were loaded with the model proteins lysozyme and bovine serum albumin, with encapsulation efficiencies of 51.5% and 73.6%, respectively. Depending on the macromonomer type, protein-loaded microgels released their cargo over a 6-14 day period. In an MTT assay, the particles did not show significant cytotoxicity, and their recognition by alveolar macrophages was considerably lower than for polystyrene control particles. This makes the microgels a promising pulmonary delivery system for proteins and other biologics.

Fabrication of antibody-loaded microgels using microfluidics and thiol-ene photoclick chemistry.

Reducing burst effects, providing controlled release, and safeguarding biologics against degradation are a few of several highly attractive applications for microgels in the field of controlled release. However, the incorporation of proteins into microgels without impairing stability is highly challenging. In this proof of concept study, the combination of microfluidics and thiol-ene photoclick chemistry was evaluated for the fabrication of antibody-loaded microgels with narrow size distribution. Norbornene-modified eight-armed poly(ethylene glycol) with an average molecular mass of 10,000 Da, 20,000 Da, or 40,000 Da were prepared as macromonomers for microgel formation. For functionalization, either hydrolytically cleavable ester or stable amide bonds were used. A microfluidic system was employed to generate precursor solution droplets containing macromonomers, the cross-linker dithiothreitol and the initiator Eosin-Y. Irradiation with visible light was used to trigger thiol-ene reactions which covalently cross-linked the droplets. For all bond-types, molecular masses, and concentrations gelation was very rapid (<20 s) and a plateau for the complex shear modulus was reached after only 5 min. The generated microgels had a rod-like shape and did not show considerable cellular toxicity. Stress conditions during the fabrication process were simulated and it could be shown that fabrication did not impair the activity of the model proteins lysozyme and bevacizumab. It was confirmed that the average hydrogel network mesh size was similar or smaller than the hydrodynamic diameter of bevacizumab which is a crucial factor for restricting diffusion and delaying release. Finally, microgels were loaded with bevacizumab and a sustained release over a period of 30 ±â€¯4 and 47 ±â€¯7 days could be achieved in vitro.

Strain Measurement in Aluminium Alloy during the Solidification Process Using Embedded Fibre Bragg Gratings.

In recent years, the observation of the behaviour of components during the production process and over their life cycle is of increasing importance. Structural health monitoring, for example of carbon composites, is state-of-the-art research. The usage of Fibre Bragg Gratings (FBGs) in this field is of major advantage. Another possible area of application is in foundries. The internal state of melts during the solidification process is of particular interest. By using embedded FBGs, temperature and stress can be monitored during the process. In this work, FBGs were embedded in aluminium alloys in order to observe the occurring strain. Two different FBG positions were chosen in the mould in order to compare its dependence. It was shown that FBGs can withstand the solidification process, although a compression in the range of one percent was measured, which is in agreement with the literature value. Furthermore, different lengths of the gratings were applied, and it was shown that shorter gratings result in more accurate measurements. The obtained results prove that FBGs are applicable as sensors for temperatures up to 740 °C.

Polymerization of silicate on hematite surfaces and its influence on arsenic sorption.

Iron oxides and oxyhydroxides are important sorbents for arsenic in soils, sediments, and water treatment systems, but their long-term potential for arsenic retention may be diminished by the formation of polymeric silicate on their surfaces. To study these interactions, we first investigated the sorption of silicate to colloidal hematite (α-Fe(2)O(3)) in short-term (48 h) and long-term (210 days) batch experiments. The polymerization of silicate on the hematite surface was monitored by attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy. The pH dependence of silicate sorption exhibited a maximum between pH 9.0 and 9.5. The condensation of silicate on hematite surfaces adsorbed from monomeric silicate solutions steadily continued over the 210 day period, whereby surface polymerization was slower at pH 3 than at pH 6. The effect of silicate surface polymerization on arsenate and arsenite sorption was studied by use of hematite pre-equilibrated with silicate for different time periods of up to 210 days. The competitive effect of silicate on arsenate and arsenite sorption increased with increasing silicate pre-equilibration time. Only under strongly acidic conditions (pH 3), where silicate sorption was weakest and surface polymerization was slowest, was arsenate and arsenite sorption not affected by the presence of silicate. We conclude that the long-term exposure to dissolved silicate can decrease the potential of natural iron (oxyhydr)oxides for adsorbing inorganic arsenic.