Enzyme-responsive polymers

The enzyme-responsive systems exploit the high specificity of the enzyme. This ensures that the cross-links are broken only in 

The degradation properties of an enzyme-responsive polymeric formulation can be characterized upon exposure to high concentrations of substrate enzymes by monitoring changes in the elastic modulus over time. To simplify data comparison, all elastic moduli are normalized to the initial time-zero elastic modulus value for each sample. The degradation rate trends exhibited by the soluble polymers translate into tunable degradation of the formulation [71]. 

In a recent study, the potential of an artificial structural motif, azobenzene, was investigated in the preparation of enzyme sensitive polymeric nanostructures. For this purpose, an azobenzene linkage 

Xi Zhang and co-workers employed chitosan and adenosines -triphosphate (ATP) as building blocks to fabricate polymeric supra-amphiphiles based on electrostatic interactions, which can self-assemble to form spherical aggregates [73]. The spherical a egates inherit the phosphatase responsiveness of ATP. This enzyme-responsive system can be more biocompatible and block polymers are not needed in preparation, which makes it possible to fabricate the chitosan-based enz nne-responsive assemblies in a large-scale, cheap way. 

In another study, a superamphiphile was formed between a double-hydrophilic polymer (methoxy-poly(ethylene g yco )-bIock-poly[L-lysine hydrochloride]] and the natural enzyme-responsive molecule (adenosine 5 -triphosphate]. The superamphiphile self-assembled into spherical aggregates, which, upon addition of enzymes, disassemble and release loaded molecules [77]. 

A wide variety of approaches have been employed to prepare hydrogels that respond to the presence of proteases. Typically, the hydrogel is exposed to a protease enzyme, and hydrolysis of protein- or peptide-based cross-linkers in the network leads to gel degradation and subsequent release of encapsulated contents. Hubbell and coworkers formed hydrogels in the presence of cells. 

Antibody-immobilized polymer chain Antigen-immobilized polymer chain Free antigen 

---

Thornton, P.D. McConnell, G. Ulijn, R.V. Enzyme responsive polymer hydrogel beads. Chem. Commun. 2005, 47 (47), 5913-5915. 

Thornton, P. D. Mart, R. J. Ulijn, R. V. Enzyme-responsive polymer hydrogel particles for controlled release. Maiei 2007,19, 1252-1256. 

Duxbury and coworkers recently developed a new chiral enzyme-responsive polymer based on enantioselective polymer modification [151]. With the aid of two alcohol dehydrogenases that show opposite enantioselectivities in the reduction of ketones (ADH-LB and ADH-T), the two enantiomers of p-vinylpheny-lethanol were obtained in excellent yield and ee. Copolymers of these monomers with styrene using free radical polymerization afforded random block copolymers... 

Enzyme-responsive polymers properties, synthesis and applications... 

Enzyme-responsive polymers stand apart from other stimuli-responsive polymers in their ability to respond to a biological molecule to regulate the function of natural materials. Ihis is both a strength and a limitation while enzyme-responsive polymers are uniquely suited to perform tasks in their niche area (i.e., in a biological surrounding), they cannot readily be used in other applications that do not preserve the activity of enzymes. 

Within their designed areas of application, however, the versatility of enzyme-responsive polymers is unmatched by any other stimuli-responsive materials. Enzyme-responsive polymers have found applications as cell supports, injectable scaffolds and drug delivery systems and have been integrated with other stimuli-responsive polymers to obtain materials with closely tailored stimuli-responsive characteristics. While research in the development of enzyme-responsive materials (ERMs) is still in its early... 

Four classes of enzyme-responsive polymers. Source Reproduced in part from (Zelzer et al., 2013).)... 

Enzyme-responsive polymers can be classified in various ways, for example according to their structure, function or response type or mechanism. Here, we classify enzyme-responsive polymers, according to their structural elements, into polymer hydrogels, supramolecular polymers, polymer particles and self-immolative polymers. The general properties of these classes and their importance in biomaterial apphcations will be introduced. Using examples from the recent literature, we will demonstrate how enzyme responsiveness can be incorporated into these materials. 

While the previous section dealt with the structural elements that define enzyme-responsive polymers, here we will introduce different methods that can be used to integrate an enzyme-responsive functionality with a polymeric material. These methods are placed into three groups. The first method deals with the preparation of enzymatically degradable polymers, the second introduces strategies to incorporate enzyme-responsive linkers into the polymer and the third explores ways to prepare enzyme-responsive polymers enzymatically. 

The introduction of enzyme-sensitive cross-links in an otherwise nonenzyme-responsive polymer such as PEG is frequently used to prepare enzyme-responsive polymer hydrogels and polymer particles. Because of their versatility and natural predisposition as enzyme substrates, short peptide sequences are almost exclusively used as cross-linkers, although dex-tran has also been used (Klinger et al., 2012). They can be readily changed to respond to a variety of proteases such as matrix metalloproteinases,plasmin or trypsin (Lutolf et al., 2003a Yang et al, 2010). In most cases, the peptides have to be modified at the termini to introduce reactive groups that are able to react with the polymer. 

Enzyme-responsive polymer conjugates are typically block polymers of an artihdal polymer and a polypeptide. Three strategies have been explored to prepare polymer-polypeptide conjugates. Firstly, the artificial polymer (e.g., PEG/PEO) can be immobilised on a solid support and terminated with an amine group (Fig. 6.11). This makes direct SPPS of the polypeptide... 

By nature, ERMs are inherently suitable for applications in the healthcare section. Despite being a young class of materials, some exciting applications have begun to emerge. Here, three applications for enzyme-responsive polymers are highlighted cell supports, injectable scaffolds and drug delivery devices. 

Enzyme-triggered release of bioactive molecules from a polymer based material is possibly the most intensely researched application for enzyme-responsive polymers. Such delivery systems typically employ vehicles such as micelles, solid particles and capsules to contain and protect a drug and deliver it to the site of action either by injection directly into the diseased tissue or by circulation through the blood stream. At the site of action, the enzyme will degrade the polymer, cause disassembly of the micelle or swell the particle and thus cause release of the drug into the environment. 

By definition, ERMs are very well suited to bridge the gap between manmade materials and biological processes (see Table 6.1) (Ulijn, 2006 Zelzer et al., 2013). Responsiveness to enzymes provides not only a means of communication between the material and its biological environment, it also presents the possibility of designing materials that display reversible and dynamic responses to a stimulus. While reversibility is not an uncommon feature for smart polymers, a dynamic interaction wherein response of a material only persists in the presence of the stimulus or a cofactor is rare. Several reversible enzyme-responsive polymer systems have been prepared so far (Ku et al, 2011), but truly dynamic or fuelled polymer-based systems have yet to be developed. 

The predisposition of ERMs to be used as biomaterials comes with some limitations. As the material relies on enzymes as stimuli, the operating conditions must preserve the function of the enzyme. This typically restricts the use of enzyme-responsive polymers to applications that respond under these circumstances. Although several enzymes are known that operate outside these conditions (e.g., at elevated temperatures or in organic solvents), no stimuli-responsive materials have yet been developed that respond to these enzymes. Moreover, since the application of ERMs is focused on biological systems, the polymers used have to be biocompatible and be able to withstand the effects of a complex biological environment in which they have to maintain their function. 

Table 6.1 Advantages and limitations of enzyme-responsive polymers...

---