Preparation of enzyme-responsive polymers

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 core of the self-immolative polymer structure is an amino-benzylal-cohol that is connected via carbamate bonds. Reaction of the amine group with phenylchloroformate converts it into a phenylcarbamate which can be polymerised in a tin-catalysed reaction under exclusion of water. After 

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. 

There are four different chemical reactions that have been used to crosslink polymers with peptide sequences (Fig. 6.8) (i) amide bond formation (ii) Michael-type addition (iii) Huisgen cycloaddition (click reaction) and (iv) radical polymerisation. The amide bond formation follows typical solid phase peptide synthesis (SPPS) protocols and does not require functionalisation of the termini of the peptide sequence. Fluorenyhnethoxycarbonyl (Fmoc) protection of the N-terminus allows attachment of the peptide sequence to an amine-bearing polymer. After removal of the Fmoc group, the amine-terminated peptide-polymer conjugate can be reacted with a second polymer bearing carboxylic acid functionalities using the same coupling chemistry (Maier et al, 2011). For Michael-type additions the peptide 

The mode in which the cross-hnker is introduced into the polymer differs and is sometimes determined by the chemical reaction used. In the case of acrylated peptides, the polymer is not prepared in advance but formed in the presence of the cross-linker. When the unmodified peptide is attached to the polymer via amide bonds, the introduction of the linker has to be carried out in two stages. First, a peptide-polymer conjugate with an amine-functionalised polymer is formed, then the conjugate is reacted with a carboxylic acid-functionalised polymer. The other two methods - Huisgen cycloaddition and Michael-type addition - allow direct cross-linking of the polymer with the peptide. 

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