An injectable PEG hydrogel controlling neurotrophin-3 release by affinity peptides.

Neurotrophin-3 growth factor can improve cochlear neuron survival, and localized delivery of this protein to the round window membrane in the middle ear may be able to reverse sensorineural hearing loss. Thus, the goal of this work was to develop an injectable hydrogel delivery system that can allow localized release of neurotrophin-3 in a controlled and sustained manner. We identified a PEG hydrogel formulation that uses thiol-vinyl sulfone Michael addition for crosslinking. This injectable formulation provides elastic hydrogels with higher mechanical rigidity, better bio-adhesion and longer residence time than Poloxamer hydrogels currently being investigated clinically for hearing loss. In vivo, PEG hydrogels induce local immune responses comparable to biocompatible Poloxamer hydrogels, yet they released payloads at a ~5-fold slower rate in the subcutaneous area. Based on this injectable hydrogel formulation, we designed an affinity-based protein release system by modifying PEG hydrogels with affinity peptides specific to neurotrophin-3 proteins. We verified the sustained release of neurotrophin-3 from peptide-conjugated PEG hydrogels resulting from the reversible interaction between peptides and proteins. The rate of affinity-controlled release depends on the polymer concentrations, the affinity of peptides and the peptide-to-protein ratios. Collectively, we developed an injectable hydrogel formulation for localized delivery of neurotrophin-3, which provides affinity-controlled release and longer delivery time compared to Poloxamer hydrogels.

Localized immune tolerance from FasL-functionalized PLG scaffolds.

Intraportal allogeneic islet transplantation has been demonstrated as a potential therapy for type 1 diabetes (T1D). The placement of islets into the liver and chronic immunosuppression to control rejection are two major limitations of islet transplantation. We hypothesize that localized immunomodulation with a novel form of FasL chimeric with streptavidin, SA-FasL, can provide protection and long-term function of islets at an extrahepatic site in the absence of chronic immunosuppression. Allogeneic islets modified with biotin and engineered to transiently display SA-FasL on their surface showed sustained survival following transplantation on microporous scaffolds into the peritoneal fat in combination with a short course (15 days) of rapamycin treatment. The challenges with modifying islets for clinical translation motivated the modification of scaffolds with SA-FasL as an off-the-shelf product. Poly (lactide-co-glycolide) (PLG) was conjugated with biotin and fabricated into particles and subsequently formed into microporous scaffolds to allow for rapid and efficient conjugation with SA-FasL. Biotinylated particles and scaffolds efficiently bound SA-FasL and induced apoptosis in cells expressing Fas receptor (FasR). Scaffolds functionalized with SA-FasL were subsequently seeded with allogeneic islets and transplanted into the peritoneal fat under the short-course of rapamycin treatment. Scaffolds modified with SA-FasL had robust engraftment of the transplanted islets that restored normoglycemia for 200 days. Transplantation without rapamycin or without SA-FasL did not support long-term survival and function. This work demonstrates that scaffolds functionalized with SA-FasL support allogeneic islet engraftment and long-term survival and function in an extrahepatic site in the absence of chronic immunosuppression with significant potential for clinical translation.

Evaluation of encapsulating and microporous nondegradable hydrogel scaffold designs on islet engraftment in rodent models of diabetes.

Islet transplantation is a promising therapeutic option for type 1 diabetes mellitus, yet the current delivery into the hepatic portal vasculature is limited by poor engraftment. Biomaterials have been used as a means to promote engraftment and function at extrahepatic sites, with strategies being categorized as encapsulation or microporous scaffolds that can either isolate or integrate islets with the host tissue, respectively. Although these approaches are typically studied separately using distinct material platforms, herein, we developed nondegradable polyethylene glycol (PEG)-based hydrogels for islet encapsulation or as microporous scaffolds for islet seeding to compare the initial engraftment and function of islets in syngeneic diabetic mice. Normoglycemia was restored with transplantation of islets within either encapsulating or microporous hydrogels containing 700 islet equivalents (IEQ), with transplantation on microporous hydrogels producing lower blood glucose levels at earlier times. A glucose challenge test at 1 month after transplant indicated that encapsulated islets had a delay in glucose-stimulated insulin secretion, whereas microporous hydrogels restored normoglycemia in times consistent with native pancreata. Encapsulated islets remained isolated from the host tissue, whereas the microporous scaffolds allowed for revascularization of the islets after transplant. Finally, we compared the inflammatory response after transplantation for the two systems and noted that microporous hydrogels had a substantially increased presence of neutrophils. Collectively, these findings suggest that both encapsulation and microporous PEG scaffold designs allow for stable engraftment of syngeneic islets and the ability to restore normoglycemia, yet the architecture influences islet function and responsiveness after transplantation.

Localized lentivirus delivery via peptide interactions.

Gene delivery from biomaterial scaffolds has been employed to induce the expression of tissue inductive factors for applications in regenerative medicine. The delivery of viral vectors has been described as reflecting a balance between vector retention and release. Herein, we investigated the design of hydrogels in order to retain the vector at the material in order to enhance transgene expression. Poly(ethylene-glycol) (PEG) hydrogels were modified with poly-l-lysine (PLL) to non-covalently bind lentivirus. For cells cultured on the hydrogels, increasing the PLL molecular weight from 1 to 70 kDa led to increased transgene expression. The incubation time of the virus with the hydrogel and the PLL concentration modulated the extent of virus adsorption, and adsorbed virus had a 20% increase in the half-life at 37°C. Alternatives to high molecular weight PLL were identified through phage display technology, with peptide sequences specific for the VSV-G ectodomain, an envelope protein pseudotyped on the virus. These affinity peptides could easily be incorporated into the hydrogel, and expression was increased 20-fold relative to control peptide, and comparable to levels observed with the high molecular weight PLL. The modification of hydrogels with affinity proteins or peptides to bind lentivirus can be a powerful strategy to enhance and localized transgene expression. Biotechnol. Bioeng. 2016;113: 2033-2040. © 2016 Wiley Periodicals, Inc.

Antifouling poly(ß-peptoid)s.

A new type of polymer highly resistant to nonspecific protein adsorption is reported. Poly(N-methyl-ß-alanine) (PMeA) and poly(N-ethyl-ß-alanine) (PEtA) synthesized via cobalt-catalyzed carbonylative polymerization of N-methylaziridine and N-ethylaziridine were end-functionalized with thiol groups and grafted onto Au surfaces. Protein adsorption was studied by the surface plasmon resonance (SPR) method. The amounts of representative single proteins adsorbed onto the PMeA- and PEtA-grafted surfaces were below the detection limit of SPR at the pg/mm(2) level. After exposure to full blood plasma and serum for 10 min, protein adsorption was at the level of ∼ 100 pg/mm(2), similar to the level of protein adsorption on poly(ethylene glycol) surfaces subjected to identical conditions. These poly(ß-peptoid)s therefore provide excellent protein resistance comparable to the best antifouling materials known to date. The strong proton-accepting ability when forming hydrogen bonds is suggested to be an important attribute for these poly(ß-peptoid)s as well as other poly(tertiary amide)s as antifouling materials.