Intravitreal hydrogels for sustained release of therapeutic proteins

This review highlights how hydrogel formulations can improve intravitreal protein delivery to the posterior segment of the eye in order to increase therapeutic outcome and patient compliance. Several therapeutic proteins have shown excellent clinical successes for the treatment of various intraocular diseases. However, drug delivery to the posterior segment of the eye faces significant challenges due to multiple physiological barriers preventing drugs from reaching the retina, among which intravitreal protein instability and rapid clearance from the site of injection. Hence, frequent injections are required to maintain therapeutic levels. Moreover, because the world population ages, the number of patients suffering from ocular diseases, such as age-related macular degeneration (AMD) and diabetic retinopathy (DR) is increasing and causing increased health care costs. Therefore, there is a growing need for suitable delivery systems able to tackle the current limitations in retinal protein delivery, which also may reduce costs. Hydrogels have shown to be promising delivery systems capable of sustaining release of therapeutic proteins and thus extending their local presence. Here, an extensive overview of preclinically developed intravitreal hydrogels is provided with attention to the rational design of clinically useful intravitreal systems. The currently used polymers, crosslinking mechanisms, in vitro/in vivo models and advancements are discussed together with the limitations and future perspective of these biomaterials.Peer reviewe

Photopolymerized thermosensitive poly(HPMAlactate)-PEG-based hydrogels : effect of network design on mechanical properties, degradation, and release behavior

Photopolymerized thermosensitive A-B-A triblock copolymer hydrogels composed of poly(N-(2-hydroxypropyl)-methacrylamide lactate) A-blocks, partly derivatizal with methacrylate groups to different extents (10, 20, and 30%) and hydrophilic poly(ethylene glycol) B-blocks of different molecular weights (4, 10, and 20 kDa) were synthesized. The aim of the present study was to correlate the polymer architecture with the hydrogel properties, particularly rheological, swelling, degradation properties and release behavior. It was found that an increasing methacrylation extent and a decreasing PEG molecular weight resulted in increasing gel strength and cross-link density, which tailored the degradation profiles from 25 to more than 300 days. Polymers having small PEG blocks showed a remarkable phase separation into polymer- and water-rich domains, as demonstrated by confocal microscopy. Depending on the hydrophobic domain density, the loaded protein resides in the hydrophilic pores or is partitioned into hydrophilic and hydrophobic domains, and its release from these compartments is tailored by the extent of methacrylation and by PEG length, respectively. As the mechanical properties, degradation, and release profiles can be fully controlled by polymer design and concentration, these hydrogels are suitable for controlled protein release

Convergence of printing technologies to engineer an interface between bone and cartilage

The combination of multiple three dimensional printing technologies can aid the generation of osteochondral grafts that display a strong interface between the cartilage and the bone compartment. In this study, the integration between bone biomimetic a three-dimensional (3D) printed calcium phosphate paste (PCP) and a gelatin methacryloyl (gelMA) hydrogel substrate for cartilage, was reinforced with a PCL mesh produced by melt electrospinning writing (MEW). Please download the file below for full content

Evaluation of bioink printability with quantitative methods to aid material development

During extrusion-based bioprinting, the deposited bioink filaments are subjected to deformations, such as collapse of overhanging filaments and fusion between adjacent filaments, which compromise shape fidelity of printed constructs. The degree of deformation of printed filaments could be used to quantitatively assess the printability of newly developed bioinks. This approach would be an alternative to current assessment through qualitative visual inspection after printing, which have been hampering any comparison between different bioinks. For this reason, we propose two quantitative printability tests based on the mentioned filament deformations: filament collapse of overhanging structures (Fig 1a) and filament fusion on parallel filaments (Fig 1b). Both printability tests were applied on two printable hydrogel platforms: poloxamer 407 and poly(ethylene glycol) blends (poloxamer/PEG), displaying a range of yield stress values. We also propose theoretical models for each test to predict printability from bioink yield stress. The results on poloxamer/PEG hydrogels show that as the yield stress decreases, the filament collapse is greater, decreasing the ability to maintain the shape of suspended filaments. Similarly, filament fusion occurs at bigger filament distances, decreasing resolution on the x-y plane. These results confirm that printability is largely dependent on yield stress. Our bioink printability testing is straightforward, assessible with any extrusion-based bioprinting system. The proposed method provides a quantitative evaluation based on physical deformation of printed filaments, potentially reducing long experimental trial-and-error printing with newly developed bioinks and allowing reproducible comparisons between different inks. Please click Additional Files below to see the full abstract

Specific N-terminal attachment of TMTHSI linkers to native peptides and proteins for strain-promoted azide alkyne cycloaddition

The site specific attachment of the reactive TMTHSI-click handle to the N-terminus of peptides and proteins is described. The resulting molecular constructs can be used in strain-promoted azide alkyne cycloaddition (SPAAC) for reaction with azide containing proteins e.g., antibodies, peptides, nanoparticles, fluorescent dyes, chelators for radioactive isotopes and SPR-chips etc

Local changes in microtubule network mobility instruct neuronal polarization and axon specification

The polarization of neurons into axons and dendrites depends on extracellular cues, intracellular signaling, cytoskeletal rearrangements, and polarized transport, but the interplay between these processes during polarization remains unresolved. Here, we show that axon specification is determined by differences in microtubule network mobility between neurites, regulated by Rho guanosine triphosphatases (GTPases) and extracellular cues. In developing neurons, retrograde microtubule flow prevents the entry of the axon-selective motor protein Kinesin-1 into most neurites. Using inducible assays to control microtubule network flow, we demonstrate that local inhibition of microtubule mobility is sufficient to guide Kinesin-1 into a specific neurite, whereas long-term global inhibition induces the formation of multiple axons. We furthermore show that extracellular mechanical cues and intracellular Rho GTPase signaling control the local differences in microtubule network flow. These results reveal a novel cytoskeletal mechanism for neuronal polarization

In situ forming IPN hydrogels of calcium alginate and dextran-HEMA for biomedical applications

AbstractIn situ forming hydrogels, which allow for the modulation of physico-chemical properties, and in which cell response can be tailored, are providing new opportunities for biomedical applications. Here, we describe interpenetrating polymer networks (IPNs) based on a physical network of calcium alginate (Alg-Ca), interpenetrated with a chemical one based on hydroxyethyl-methacrylate-derivatized dextran (dex-HEMA). IPNs with different concentration and degree of substitution of dex-HEMA were characterized and evaluated for protein release as well as for the behavior of embedded cells. The results demonstrated that the properties of the semi-IPNs, which are obtained by dissolution of dex-HEMA chains into the Alg-Ca hydrogels, would allow for injection of these hydrogels. Degradation times of the IPNs after photocross-linking could be tailored from 15 to 180days by the concentration and the degree of substitution of dex-HEMA. Further, after an initial burst release, bovine serum albumin was gradually released from the IPNs over approximately 15days. Encapsulation of expanded chondrocytes in the IPNs revealed that cells remained viable and, depending on the composition, were able to redifferentiate, as was demonstrated by the deposition of collagen type II. These results demonstrate that these IPNs are attractive materials for pharmaceutical and biomedical applications due to their tailorable mechanical and degradation characteristics, their release kinetics and biocompatibility

Creating a Functional Biomimetic Cartilage Implant Using Hydrogels Based on Methacrylated Chondroitin Sulfate and Hyaluronic Acid

The load-bearing function of articular cartilage tissue contrasts with the poor load-bearing capacity of most soft hydrogels used for its regeneration. The present study explores whether a hydrogel based on the methacrylated natural polymers chondroitin sulfate (CSMA) and hyaluronic acid (HAMA), injected into warp-knitted spacer fabrics, could be used to create a biomimetic construct with cartilage-like mechanical properties. The swelling ratio of the combined CSMA/HAMA hydrogels in the first 20 days was higher for hydrogels with a higher CSMA concentration, and these hydrogels also degraded quicker, whereas those with a 1.33 wt% of HAMA were stable for more than 120 days. When confined by a polyamide 6 (PA6) spacer fabric, the volumetric swelling of the combined CSMA/HAMA gels (10 wt%, 6.5 × CSMA:HAMA ratio) was reduced by ~53%. Both the apparent peak and the equilibrium modulus significantly increased in the PA6-restricted constructs compared to the free-swelling hydrogels after 28 days of swelling, and no significant differences in the moduli and time constant compared to native bovine cartilage were observed. Moreover, the cell viability in the CSMA/HAMA PA6 constructs was comparable to that in gelatin-methacrylamide (GelMA) PA6 constructs at one day after polymerization. These results suggest that using a HydroSpacer construct with an extracellular matrix (ECM)-like biopolymer-based hydrogel is a promising approach for mimicking the load-bearing properties of native cartilage

Exploring silk fibroin aerogels via different coagulation approaches

Aerogels are highly porous nanostructured materials with a high specific surface area, a very low density and a low thermal conductivity. Bio-aerogels, prepared from biopolymers, have gained interest for biomedical applications. While most bio-aerogels are polysaccharide-based, protein-based aerogels using silk fibroin (SF) from Bombyx mori cocoons remain underexplored. Our study delved into the impact of coagulation methods on SF aerogel properties, specifically, comparing ethanol treatment (Method 1) and sodium dihydrogen phosphate (NaH2PO4) (Method 2). Aerogels obtained through Method 1 exhibited a volume shrinkage of approximately 12–22 % and a density of about 0.06–0.07 g cm−3, while those obtained via Method 2 demonstrated a more substantial volume shrinkage of approximately 60–70 % and a higher density of around 0.2 g cm−3. These differences significantly influenced the internal structure of the aerogels, manifesting distinct morphological features of the materials. Mechanical tests revealed that SF aerogels derived from Method 2 displayed superior stress resistance at 80 % strain and higher elastic recovery when compared to samples of Method 1. In conclusion, the choice in coagulation methods broadens the mechanical property and density window for this type of aerogel which offers opportunities for a wide range of biomedical applications

Self-Healing Thermosensitive Hydrogel for Sustained Release of Dexamethasone for Ocular Therapy

The aim of this study was to develop an injectable hydrogel delivery system for sustained ocular delivery of dexamethasone. To this end, a self-healing hydrogel consisting of a thermosensitive ABA triblock copolymer was designed. The drug was covalently linked to the polymer by copolymerization of methacrylated dexamethasone with N-isopropylacrylamide (NIPAM) and N-acryloxysuccinimide (NAS) through reversible addition-fragmentation chain transfer (RAFT) polymerization, using poly(ethylene glycol) (PEG) functionalized at both ends with a chain transfer agent (CTA). Hydrogel formation was achieved by mixing aqueous solutions of the formed thermosensitive polymer (with a cloud point of 23 °C) with cystamine at 37 °C, to result in covalent cross-linking due to the reaction of the N-hydroxysuccimide (NHS) functionality of the polymer and the primary amines of cystamine. Rheological analysis showed both thermogelation and covalent cross-linking at 37 °C, as well as the self-healing properties of the formed network, which was attributed to the presence of disulfide bonds in the cystamine cross-links, making the system injectable. The release of dexamethasone from the hydrogel occurred through ester hydrolysis following first-order kinetics in an aqueous medium at pH 7.4 over 430 days at 37 °C. Based on simulations, administration of 100 mg of hydrogel would be sufficient for maintaining therapeutic levels of dexamethasone in the vitreous for at least 500 days. Importantly, dexamethasone was released from the hydrogel in its native form as determined by LC-MS analysis. Cytocompatibility studies showed that at clinically relevant concentrations, both the polymer and the cross-linker were well tolerated by adult retinal pigment epithelium (ARPE-19) cells. Moreover, the hydrogel did not show any toxicity to ARPE-19 cells. The injectability of the hydrogel, together with the long-lasting release of dexamethasone and good cytocompatibility with a retinal cell line, makes this delivery system an attractive candidate for treatment of ocular inflammatory diseases