Poly(ethylene glycol) hydrogels formed by thiol-ene photopolymerization for enzyme-responsive protein delivery

Abstract

Degradable hydrogels have been extensively used in biomedical applications such as drug delivery, and recent interest has grown in hydrogels that degrade in recognition of a cellular response. This contribution describes a poly(ethylene glycol) (PEG) hydrogel platform with human neutrophil elastase (HNE) sensitive peptide cross-links formed using thiol-ene photopolymerization rendering the gel degradable at sites of inflammation. Further, protein therapeutics can be physically entrapped within the network and selectively released upon exposure to HNE. HNE-responsive hydrogels exhibited surface erosion where the degradation kinetics was influenced by changes in peptide k(cat), concentration of HNE, and concentration of peptide within the gel. Using this platform, we were able to achieve controlled, zero-order release of bovine serum albumin (BSA) in the presence of HNE, and release was arrested in the absence of HNE. To further exploit the advantages of surface eroding delivery systems, a smaller protein (carbonic anhydrase) was delivered at the same rate as BSA and only dependent on gel formulation and environmental conditions. Also, protein release was predicted from a 3-layered hydrogel device using mass loss data. Lastly, the bioactivity of lysozyme was maintained above 90% following the exposure to thiol-ene photopolymerization conditions.

Fig. 1

Schematic illustrating the design and formation of enzyme-responsive PEG hydrogels fabricated via thiol-ene photopolymerization (reaction mechanism included) for the controlled release of protein therapeutics. Enzyme cleavage occurs between the P1 and P1’ amino acid residues.

Fig. 2

Dynamic time sweep photorheology of thiol-ene photopolymerization hydrogel formation with PEG_5k. Gel point is approximated when the storage modulus (G’, ●) overcomes the loss modulus (G”, □). Dashed line (---) illustrates when the UV light was turned on to initiate polymerization.

Fig. 3

Gel mass loss profiles upon exposure to HNE (A-C). Influence of HNE peptide cross-link (A), concentration of HNE (B), and concentration of substrate (C) were experimentally studied. (D) Representative equilibrium swelling ratio as a function of time in the presence of HNE. All gels were made of 10 wt% monomer and exposed to 1μM HNE (unless otherwise noted).

Fig. 4

BSA release profiles in the presence of 1μM HNE. (A) Protein release from thiol-ene hydrogels fabricated with varying peptide cross-links. (B) Temporal control of protein release when HNE is dosed (□) and when it is removed (). HNE substrate: CGAAPV↓RGGGGC.

Fig. 5

Protein release from thiol-ene hydrogels as a function of fractional mass loss. Each data point represents the same time. Gel formulation: PEG_5k-norbornene + CGAAPVRGGGC + protein (1mg mL^-1). [HNE] = 1 μM.

Fig. 6

BSA release from 3-layered hydrogel formed by subsequent thiol-ene photopolymerizations. ■ = PEG_5k-norbornene + CGAAPVRGGGC + BSA (1 mg mL^-1), □ = PEG_5k-norbornene + CGAAP(Nva)GGGGC + BSA (1 mg mL^-1). Solid (—) and dashed (- - -) lines represent predicted release.

Fig. 7

Influence of thiol-ene photopolymerization on the bioactivity of lysozyme in the presence of varying concentrations of photoinitiator, I-2959. All samples were exposed to 365 nm light for 10 minutes with or without macromers present in the prepolymer solution.