Enzyme responsive nanocapsules for protein delivery

Abstract

The invention provides methods of making and using compositions comprising a polymer shell designed to deliver polypeptides to selected environments. In embodiments of the invention, different environmental conditions are harnessed to allow the selective degradation of the polymer shell and the consequential release of one or polypeptides encapsulated therein. In illustrative embodiments, polymer components of the shell are interconnected by peptide-containing crosslinker to moieties, linkages which maintain the integrity of the polymer shell under certain enviromental conditions, but can also be cleaved when combined with a selected protease.

1 . A composition of matter comprising:
 at least one polypeptide;   a polymeric network, wherein:   polymers in the polymeric network are coupled together by a first crosslinking agent and a second crosslinking agent so as to form a shell that encapsulates the polypeptide;   the first crosslinking agent comprises a peptide having an amino acid sequence that is cleaved by a protease; and   the second crosslinking agent does not comprise a peptide having an amino acid sequence that is cleaved by a protease;   wherein the polymers, the first crosslinking agent and the second crosslinking agent are disposed within the polymeric network in an orientation so that proteolytic cleavage of the first crosslinking agent releases the polypeptide from the shell into an external environment.   
     
     
         2 . The composition of  claim 1 , wherein relative molar amounts of the first crosslinking agent and the second crosslinking agent in the shell are at least 1:5, 1:4, 1:3, 1:2, 1:1, 2:1, 3:1, 4:1 or 5:1. 
     
     
         3 . The composition of  claim 1 , wherein the polymeric network is formed in situ on the at least one polypeptide so as to encapsulate the at least one polypeptide. 
     
     
         4 . The composition of  claim 1 , wherein shell that encapsulates the polypeptide has a diameter between 15 and 35 nanometers. 
     
     
         5 . The composition of  claim 1 , wherein the polypeptide comprises a protein that stimulates cellular growth. 
     
     
         6 . The composition of  claim 1 , wherein the first crosslinking agent comprises a peptide having D-amino acids. 
     
     
         7 . The composition of  claim 1 , wherein the composition further comprises a hydrogel in which the shell is disposed. 
     
     
         8 . The composition of  claim 7 , wherein the shell is covalently coupled to the hydrogel by a hydrogel crosslinking agent. 
     
     
         9 . The composition of  claim 8 , wherein the hydrogel crosslinking agent comprises a peptide having an amino acid sequence that is cleaved by a protease. 
     
     
         10 . The composition of  claim 9 , wherein the hydrogel crosslinking agent comprises a peptide having D-amino acids. 
     
     
         11 . The composition of  claim 1 , wherein the concentration of the first crosslinking agent in the composition relative to the concentration of the second crosslinking agent in the composition is controlled so as to modulate the rate at which the polypeptide is released from the shell into an external in vivo environment. 
     
     
         12 . The composition of  claim 11 , wherein the concentration of the first crosslinking agent is greater than the concentration of the second crosslinking agent in order to modulate the rate at which the polypeptide is released from the shell into an external in vivo environment. 
     
     
         13 . The composition of  claim 11 , wherein the concentration of the first crosslinking agent is less than the concentration of the second crosslinking agent in order to modulate the rate at which the polypeptide is released from the shell into an external in vivo environment. 
     
     
         14 . A composition of matter comprising:
 at least one polypeptide;   a polymeric network, wherein:   polymers in the polymeric network are coupled together by a first crosslinking agent and a second crosslinking agent so as to form a shell that encapsulates the polypeptide;   the first crosslinking agent comprises a peptide having an amino acid sequence that is cleaved by a protease, and a first end of the first crosslinking agent is coupled to a first polymer and a second end of first crosslinking agent is coupled to a second polymer; and   the second crosslinking agent does not comprise a peptide having an amino acid sequence that is cleaved by a protease, and a first end of the second crosslinking agent is coupled to a first polymer and a second end of second crosslinking agent is coupled to a second polymer;   wherein the polymers, the first crosslinking agent and the second crosslinking agent are disposed within the polymeric network in an orientation so that proteolytic cleavage of the first crosslinking agent releases the polypeptide from the shell into an external environment.   
     
     
         15 . The composition of  claim 14 , wherein the first crosslinking agent and the second crosslinking agent are coupled to the same polymer within the polymeric network. 
     
     
         16 . The composition of  claim 14 , wherein the first crosslinking agent and the second crosslinking agent are coupled to different polymers within the polymeric network. 
     
     
         17 . The composition of  claim 14 , wherein the first crosslinking agent comprises a peptide having D-amino acids. 
     
     
         18 . The composition of  claim 14 , wherein the concentration of the first crosslinking agent in the composition and the concentration of the second crosslinking agent in the composition are controlled so as to modulate the rate at which the polypeptide is released from the shell into an external in vivo environment. 
     
     
         19 . The composition of  claim 14 , wherein the polymeric network is formed in situ on the at least one polypeptide so as to encapsulate the at least one polypeptide. 
     
     
         20 . A composition of matter comprising:
 at least two polypeptides;   a polymeric network, wherein:   polymers in the polymeric network are coupled together by a first crosslinking agent and a second crosslinking agent so as to form a shell that encapsulates the polypeptide;   the polymeric network is formed in situ on the at least two polypeptides so as to encapsulate the polypeptides;   the first crosslinking agent comprises a peptide having an amino acid sequence that is cleaved by a protease; and   the second crosslinking agent does not comprise a peptide having an amino acid sequence that is cleaved by the protease;   wherein the polymers, the first crosslinking agent and the second crosslinking agent are disposed within the polymeric network in an orientation so that proteolytic cleavage of the first crosslinking agent releases the polypeptide from the shell into an external environment.   (c) initiating polymerization of the plurality of polymerizable monomers and the first and second crosslinking agents at interfaces between the monomers and the polypeptide;   so that the polymeric nanocapsule around the polypeptide is formed.