Elastic biomaterials have been developed by using both natural and synthetic polymers for a wide range of biomedical applications where elasticity plays a critical role. For example, elastic hydrogels are ideal for tissue engineering applications that require stretchable biomaterials (Shin S R, et al., ACS Nano 7, 2369-2380 (2013); Kharaziha M, et al., Biomaterials 35, 7346-7354 (2014); Paul A, et al., ACS Nano 8, 8050-8062 (2014); Annabi N, et al., Adv Funct Mater 23, 4950-4959 (2013)) such as engineering soft and elastic tissues, like skin and blood vessels (Böttcher-Haberzeth S, et al., Burns 36, 450-460 (2010); Kim T G, et al., Adv Funct Mater 22, 2446-2468 (2012)). Other biomedical applications including tissue adhesives (Annabi N, et al., Nano Today 9, 574-589 (2014); Elvin C M, et al., Biomaterials 31, 8323-8331 (2010)), smart hydrogels (Xia L-W, et al., Nat Commun 4, (2013)), and flexible electronics (Rogers J A, et al., Science 327, 1603-1607 (2010)) also demand materials with high elasticity and rapid response to applied mechanical forces. Synthetic elastomers (Wang Y, et al., Nat Biotech 20, 602-606 (2002)), interpenetrating networks (Omidian H, et al., Macromol Biosci 6, 703-710 (2006)), and nanocomposite hydrogels (Li Y, et al., Macromolecules 42, 2587-2593 (2009)) have been investigated for generating elastic substrates but properties such as cell adhesion, degradability, and overall biocompatibility must be artificially incorporated into these polymeric systems (Zhu J, et al., Expert review of medical devices 8, 607-626 (2011)).
Alternatively, recombinant protein-based polymers such as elastin-like polypeptides (ELPs) are biocompatible (MacEwan S R, et al., Biopolymers 94, 60-77 (2010); Nettles D L, et al., Adv Drug Delivery Rev 62, 1479-1485 (2010)) and have been widely investigated for biomedical applications (Nettles D L, et al., Adv Drug Delivery Rev 62, 1479-1485 (2010); Nettles D L, et al., Tissue engineering Part A 14, 1133-1140 (2008); McHale M K, et al., Tissue Eng 11, 1768-1779 (2005)). They recapitulate the extensibility of natural elastin using a pentapeptide repeat, VPGXG, where X is any amino acid besides proline (SEQ ID NO: 1) (MacEwan S R, et al., Biopolymers 94, 60-77 (2010); Baldock C, et al., Proc Natl Acad Sci 108, 4322-4327 (2011)). The unique properties of ELPs include their reversible thermoresponsive nature (Meyer D E, et al., Nat Biotech 17, 1112-1115 (1999)), modular design (Raphel J, et al., JMater Chem 22, 19429-19437 (2012); Betre H, et al., Biomacromolecules 3, 910-916 (2002)), and mechanical properties (McHale M K, et al., Tissue Eng 11, 1768-1779 (2005)), making them suitable candidates for various applications such as thermoresponsive drug carriers, thermal purification components, self-assembly building blocks, and hydrogels for tissue regeneration (Nettles D L, et al., Adv Drug Delivery Rev 62, 1479-1485 (2010); McHale M K, et al., Tissue Eng 11, 1768-1779 (2005); Xia X X, et al., Biomacromolecules 12, 3844-3850 (2011)). However, many ELP-based biomaterials have lower elasticity and mechanical properties compared to synthetic polymeric scaffolds (Trabbic-Carlson K, et al., Biomacromolecules 4, 572-580 (2003)). They also require chemical modification and long crosslinking time to form three dimensional (3D) scaffolds, limiting their capability for 3D cell encapsulation (Raphel J, et al., JMater Chem 22, 19429-19437 (2012)).
To alter the mechanical properties of ELPs, various crosslinking approaches such as physical (through temperature changes), chemical, and enzymatic crosslinking have been applied (Chou C, et al., Chemical Science 2, 480-483 (2011)). Above an ELP's lower critical solution temperature (LCST), ELP agglomeration occurs, forming coacervates (viscous liquids), which can be used as a cell carrier or injectable delivery system (MacEwan S R, et al., Biopolymers 94, 60-77 (2010)). However, these viscous liquids are not capable of providing the mechanics necessary for many tissue engineering applications. For the formation of more stable gels, chemical crosslinking of ELPs has been performed. ELPs can be crosslinked through chemical functionalization of the protein sequence (e.g. addition of vinyl groups) using N-hydroxysuccinimide (NHS) reactions, glutaraldehyde (Ifkovits J L, et al., Tissue Eng 10, 2369-2385 (2007); Wissink M J B, et al., Biomaterials 22, 151-163 (2001)), or through reacting amines or carboxyl groups in the biopolymer (Nettles D L, et al., Adv Drug Delivery Rev 62, 1479-1485 (2010); Nettles D L, et al., Tissue engineering Part A 14, 1133-1140 (2008); Raphel J, et al., J Mater Chem 22, 19429-19437 (2012); Chou C, et al., Chemical Science 2, 480-483 (2011)). Chemical modifications can introduce well-defined concentrations of crosslinkers per molecule, providing control over both the location and amount of added crosslinkable groups. However, long reaction times and generation of toxic byproducts in chemical crosslinking methods may limit the applications of resulting ELP biomaterials in situations where rapid gelation in biological conditions is required. For example, NETS-based coupling reactions commonly require minutes to hours to react generating toxic byproducts (Huang L, et al., Macromolecules 33, 2989-2997 (2000); Zhang K, et al., J Am Chem Soc 127, 10136-10137 (2005)), which is not ideal for clinical applications. Enzymatic crosslinking of proteins also suffers from similar restrictions (McHale M K, et al., Tissue Eng 11, 1768-1779 (2005)).
To address these limitations, photocrosslinkable ELP-based scaffolds have been developed. For example, non-canonical amino acids have been recombinantly incorporated to provide photocrosslinkable sites within the protein but the protein yield using this technology was very low, limiting the scalability and widespread applications of this approach (Nagapudi K, et al., Macromolecules 35, 1730-1737 (2002)). Photocrosslinking of ELPs has also been performed by functionalizing lysine groups with acrylate moieties (K. Nagapudi, W. T. Brinkman, J. E. Leisen, L. Huang, R. A. McMillan, R. P. Apkarian, V. P. Conticello, E. L. Chaikof, Macromolecules 2002, 35, 1730) or NETS ester-diazirine crosslinker (Raphel J, et al., J Mater Chem 22, 19429-19437 (2012)) to produce stable biomaterials. These approaches have been utilized to design dried films or fibers with high modulus (40-60 MPa) and low fracture strain (2%) but rarely for the formation of bulk hydrogels, which are necessary for any 3D applications, including cell encapsulation (Bertassoni L E, et al., Lab on a Chip 14, 2202-2211 (2014)), micropatterning (Annabi N, et al., Biomaterials 34, 5496-5505 (2013)), or molding (Nettles D L, et al., Adv Drug Delivery Rev 62, 1479-1485 (2010); Foo CTWP, et al., Proc Natl Acad Sci 106, 22067-22072 (2009); Tang S, et al., Macromolecules 47, 791-799 (2014)) by soft lithography. The functionalized ELP polymers also required long UV exposure times (1-2 h) to be crosslinked (Raphel J, et al., J Mater Chem 22, 19429-19437 (2012); Nichol J W, et al., Biomaterials 31, 5536-5544 (2010)), limiting their capability for 3D cell encapsulation and fast polymerizable materials for surgical application or injectable fillers.
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