Affinity-based Delivery Systems Research Articles

Mathematical model accurately predicts protein release from an affinity-based delivery system

Affinity-based controlled release modulates the delivery of protein or small molecule therapeutics through transient dissociation/association. To understand which parameters can be used to tune release, we used a mathematical model based on simple binding kinetics. A comprehensive asymptotic analysis revealed three characteristic regimes for therapeutic release from affinity-based systems. These regimes can be controlled by diffusion or unbinding kinetics, and can exhibit release over either a single stage or two stages. This analysis fundamentally changes the way we think of controlling release from affinity-based systems and thereby explains some of the discrepancies in the literature on which parameters influence affinity-based release. The rate of protein release from affinity-based systems is determined by the balance of diffusion of the therapeutic agent through the hydrogel and the dissociation kinetics of the affinity pair. Equations for tuning protein release rate by altering the strength (KD) of the affinity interaction, the concentration of binding ligand in the system, the rate of dissociation (koff) of the complex, and the hydrogel size and geometry, are provided. We validated our model by collapsing the model simulations and the experimental data from a recently described affinity release system, to a single master curve. Importantly, this mathematical analysis can be applied to any single species affinity-based system to determine the parameters required for a desired release profile.

Minimizing the passive release of heparin-binding growth factors from an affinity-based delivery system

We consider a mathematical model that describes the leakage of heparin-binding growth factors from an affinity-based delivery system. In the delivery system, heparin binds to a peptide which has been covalently cross-linked to a fibrin matrix. Growth factor in turn binds to the heparin, and growth factor release is governed by both binding and diffusion mechanisms, the purpose of the binding being to slow growth factor release. The governing mathematical model, which in its original formulation consists of six partial differential equations, is reduced to a system of just two equations. It is usually desirable that there be no passive release of growth factor from a device, with all of the growth factor being held in place via binding until such time as it is actively released by invading cells. However, there will inevitably be some passive release, and so it is of interest to identify conditions that will make this release as slow as possible. In this paper, we identify a parameter regime that ensures that at least a fraction of the growth factor will release slowly. It is found that slow release is assured if the matrix is prepared with the concentration of cross-linked peptide greatly exceeding the dissociation constant of heparin from the peptide, and with the concentration of heparin greatly exceeding the dissociation constant of the growth factor from heparin. Also, for the first time, in vitro experimental release data are directly compared with the theoretical release profiles generated by the model. We propose that the two stage release behaviour frequently seen in experiments is due to an initial rapid out-diffusion of free growth factor over a diffusion time scale (typically days), followed by a much slower release of the bound fraction over a time scale depending on both diffusion and binding parameters (frequently months).

Fibrin matrices with affinity‐based delivery systems and neurotrophic factors promote functional nerve regeneration

Glial-derived neurotrophic factor (GDNF) and nerve growth factor (NGF) have both been shown to enhance peripheral nerve regeneration following injury and target different neuronal populations. The delivery of either growth factor at the site of injury may, therefore, result in quantitative differences in motor nerve regeneration and functional recovery. In this study we evaluated the effect of affinity-based delivery of GDNF or NGF from fibrin-filled nerve guidance conduits (NGCs) on motor nerve regeneration and functional recovery in a 13 mm rat sciatic nerve defect. Seven experimental groups were evaluated consisting of GDNF or NGF and the affinity-based delivery system (DS) within NGCs, control groups excluding the DS and/or growth factor, and nerve isografts. Groups with growth factor in the conduit demonstrated equivalent or superior performance in behavioral tests and relative muscle mass measurements compared to isografts at 12 weeks. Additionally, groups with GDNF demonstrated greater specific twitch and tetanic force production in extensor digitorum longus (EDL) muscle than the isograft control, while groups with NGF produced demonstrated similar force production compared to the isograft control. Assessment of motor axon regeneration by retrograde labeling further revealed that the number of ventral horn neurons regenerating across NGCs containing GDNF and NGF DS was similar to the isograft group and these counts were greater than the groups without growth factor. Overall, the GDNF DS group demonstrated superior functional recovery and equivalent motor nerve regeneration compared to the isograft control, suggesting it has potential as a treatment for motor nerve injury.

Heparin-Binding-Affinity-Based Delivery Systems Releasing Nerve Growth Factor Enhance Sciatic Nerve Regeneration

The controlled delivery of nerve growth factor (NGF) to the peripheral nervous system has been shown to enhance nerve regeneration following injury, although the effect of release rate has not been previously studied with an affinity-based delivery system (DS). The goal of this research was to determine if the binding site affinity of the DS affected nerve regeneration in vivo using nerve guidance conduits (NGCs) in a 13-mm rat sciatic nerve defect. These DSs consisted of bi-domain peptides that varied in heparin-binding affinity, heparin and NGF, which binds to heparin with moderate affinity. Eight experimental groups were evaluated consisting of NGF with DS, control groups excluding one or more components of the DS within silicone conduits and nerve isografts. Nerves were harvested 6 weeks after treatment for analysis by histomorphometry. These DSs with NGF resulted in a higher frequency of nerve regeneration compared to control groups and were similar to the nerve isograft group in measures of nerve fiber density and percent neural tissue, but not in total nerve fiber count. In addition, these DSs with NGF contained a significantly greater percentage of larger diameter nerve fibers, suggesting more mature regenerating nerve content. While there were no differences in nerve regeneration due to varying peptide affinity with these DSs, their use with NGF enhanced peripheral nerve regeneration through a NGC across a critical nerve gap.

Affinity-based release of glial-derived neurotrophic factor from fibrin matrices enhances sciatic nerve regeneration

Glial-derived neurotrophic factor (GDNF) promotes both sensory and motor neuron survival. The delivery of GDNF to the peripheral nervous system has been shown to enhance regeneration following injury. In this study, we evaluated the effect of affinity-based delivery of GDNF from a fibrin matrix in a nerve guidance conduit on nerve regeneration in a 13 mm rat sciatic nerve defect. Seven experimental groups were evaluated which received GDNF or nerve growth factor (NGF) with the delivery system within the conduit, control groups excluding one or more components of the delivery system, and nerve isografts. Nerves were harvested 6 weeks after treatment for analysis by histomorphometry and electron microscopy. The use of the delivery system (DS) with either GDNF or NGF resulted in a higher frequency of nerve regeneration vs. control groups, as evidenced by a neural structure spanning the 13 mm gap. The GDNF DS and NGF DS groups were also similar to the nerve isograft group in measures of nerve fiber density, percent neural tissue and myelinated area measurements, but not in terms of total fiber counts. In addition, both groups contained a significantly greater percentage of larger diameter fibers, with GDNF DS having the largest in comparison to all groups, suggesting more mature neural content. The delivery of GDNF via the affinity-based delivery system can enhance peripheral nerve regeneration through a silicone conduit across a critical nerve gap and offers insight into potential future alternatives to the treatment of peripheral nerve injuries.

The effect of controlled growth factor delivery on embryonic stem cell differentiation inside fibrin scaffolds

The goal of this project was to develop 3-D biomaterial scaffolds that present cues to direct the differentiation of embryonic stem (ES) cell-derived neural progenitor cells, seeded inside the scaffolds, into mature neural phenotypes, specifically neurons and oligodendrocytes. Release studies were performed to determine the appropriate conditions for retention of neurotrophin-3 (NT-3), sonic hedgehog, and platelet-derived growth factor (PDGF) by an affinity-based delivery system incorporated into fibrin scaffolds. Embryoid bodies containing neural progenitors were formed from mouse ES cells, using a 4−/4+ retinoic acid treatment protocol, and then seeded inside fibrin scaffolds containing the drug delivery system. This delivery system was used to deliver various growth factor doses and combinations to the cells seeded inside the scaffolds. Controlled delivery of NT-3 and PDGF simultaneously increased the fraction of neural progenitors, neurons, and oligodendrocytes while decreasing the fraction of astrocytes obtained compared to control cultures seeded inside unmodified fibrin scaffolds with no growth factors present in the medium. These results demonstrate that such a strategy can be used to generate an engineered tissue for the potential treatment of spinal cord injury and could be extended to the study of differentiation in other tissues.

Controlled release of glial‐derived neurotrophic factor from fibrin matrices containing an affinity‐based delivery system

This research evaluated the controlled release of glial-derived neurotrophic factor (GDNF) from an affinity-based delivery system (ABDS) as potential treatment for peripheral nerve injury. The ABDS consisted of a bidomain peptide containing a transglutaminase substrate, allowing crosslinking into fibrin matrices, and a heparin-binding domain based on the antithrombin-III heparin-binding domain, heparin, and GDNF, which was sequestered based on its heparin-binding affinity. The objective of this research was to determine the release rate and biological activity of GDNF released from the ABDS in vitro. The ratio of peptide to heparin was found to modulate the rate of GDNF release. The biological activity of GDNF released from the ABDS was assayed using chick dorsal root ganglia (DRGs) neurite extension. Neurite extension was equivalent for fibrin matrices containing the ABDS for all concentrations of GDNF tested versus DRGs grown with GDNF in the media. Furthermore, neurite extension was enhanced in fibrin matrices containing 100 ng/mL of GDNF with the ABDS versus matrices with GDNF at a simliar dose but no ABDS. These results suggest that GDNF can be retained and released in a biologically activity form from the ABDS, and thus this approach may prove useful for the treatment of peripheral nerve injury.

Effect of controlled delivery of neurotrophin-3 from fibrin on spinal cord injury in a long term model

The goal of this work was to assess the effect of the controlled delivery of neurotrophin-3 (NT-3) from an affinity-based delivery system in fibrin scaffolds on regeneration following spinal cord injury (SCI). A heparin-based delivery system (HBDS) was used to immobilize NT-3 within fibrin scaffolds via non-covalent interactions. The fibrin scaffolds were implanted in lesions immediately after injury in an adult rat model of SCI (complete ablation of a 2 mm segment of the cord at T9). Delivery of NT-3 was controlled by an affinity-based delivery system that limits drug loss by diffusion and releases the drug via cell-mediated processes. Twelve weeks after injury and treatment, animals treated with fibrin scaffolds and NT-3, with or without the delivery system, did not show functional improvement over saline controls. Substantial cavitation at edges of the lesion was present, and while neuronal fibers were present inside the lesion, traced corticospinal and dorsal sensory tracts did not regenerate into the lesion. Therefore, while previous studies indicate that the controlled delivery of NT-3 from fibrin scaffolds may increase the short term regenerative response, the continued degeneration of the cord, indicative of the severity of the injury, limits the long term regeneration stimulated by this treatment. Chronic or repeated treatments or a less severe injury model may prove useful in assessing the utility of controlled delivery systems for the treatment of spinal cord injury.

Controlled release of neurotrophin-3 from fibrin gels for spinal cord injury

The goal of this work was to assess the feasibility of using affinity-based delivery systems to release neurotrophin-3 (NT-3) in a controlled manner from fibrin gels as a therapy for spinal cord injury. A heparin-based delivery system (HBDS) was used to immobilize NT-3 within fibrin gels via non-covalent interactions to slow diffusion-based release of NT-3, thus allowing cell-activated degradation of fibrin to mediate release. The HBDS consists of three components: immobilized linker peptide, heparin and NT-3. The linker peptide contained a Factor XIIIa substrate and was covalently cross-linked to fibrin during polymerization. This immobilized linker peptide sequesters heparin within fibrin gels, and sequestered heparin binds NT-3, preventing its diffusion. Mathematical modeling was performed to examine the effect of heparin concentration on the fraction of NT-3 initially bound to fibrin. In vitro release studies confirmed that heparin concentration modulates diffusion-based release of NT-3. Fibrin gels containing the HBDS and NT-3 stimulated neural outgrowth from chick dorsal root ganglia by up to 54% versus unmodified fibrin, demonstrating that the NT-3 released is biologically active. In a preliminary in vivo study, fibrin gels containing the HBDS and NT-3 showed increased neural fiber density in spinal cord lesions versus unmodified fibrin at 9 days.