Silk Fibroin Hydrogels and Uses Thereof
The present specification provides for methods for purifying fibroins, purified fibroins, methods of conjugating biological and synthetic molecules to fibroins, fibroins conjugated to such molecules, methods of making fibroin hydrogels, fibroin hydrogels and fibroin hydrogel formulations useful for a variety of medical uses, including, without limitation uses as bulking agents, tissue space fillers, templates for tissue reconstruction or regeneration, cell culture scaffolds for tissue engineering and for disease models, surface coating to improve medical device function, or drug delivery devices.
1 . A hydrogel formulation comprising:
a) a gel phase, the gel phase including hydrogel particles comprising a substantially sericin-depleted silk fibroin and an amphiphilic peptide; and
b) a carrier phase.
2 . The hydrogel formulation of claim 1 , wherein the hydrogel particles comprises about 1% (w/v) to about 10% (w/v) of silk fibroin.
3 . The hydrogel formulation of claim 1 , wherein the amphiphilic peptide comprising a RGD motif.
4 . The hydrogel formulation of claim 3 , wherein the amphiphilic peptide is 23 RGD.
5 . The hydrogel formulation of claim 1 , wherein the amphiphilic peptide comprises of a tail region, followed by a spacer region and finally a RGD motif.
6 . The hydrogel formulation of claim 1 , wherein the hydrogel particles comprises a molar ratio of 1:10 to 10:1 moles of the amphiphilic peptide per mole of the silk fibroin.
7 . The hydrogel formulation of claim 6 , wherein the hydrogel particles comprises a molar ratio of 3:1 moles of the amphiphilic peptide per mole of the silk fibroin.
8 . The hydrogel formulation of claim 1 , wherein the hydrogel particles comprises a protein structure having a β-sheet conformation of at least 80%.
9 . The hydrogel formulation of claim 1 , wherein the hydrogel particles comprises a protein structure having a β-sheet conformation of at least 50%.
10 . The hydrogel formulation of claim 1 , wherein the hydrogel particles comprises a protein structure having a β-sheet conformation of at least 20%.
11 . The hydrogel formulation of claim 1 , wherein the hydrogel particles comprises a protein structure having an α-helical and random coil conformation of at most 20%.
12 . The hydrogel formulation of claim 1 , wherein the hydrogel particles have a cross-sectional area from about 5 μm 2 to about 1000 μm 2 .
13 . The hydrogel formulation of claim 1 , wherein the hydrogel particles have a cross-sectional area from about 20 μm 2 to about 50 μm 2 .
14 . The hydrogel formulation of claim 1 , wherein the carrier phase comprises saline.
15 . The hydrogel formulation of claim 1 , wherein the carrier phase comprises a surfactant solution.
16 . The hydrogel formulation of claim 1 wherein the gel phase is 50% to 99% of the total formulation volume, the remainder being a carrier solution.
17 . The hydrogel formulation of claim 16 , wherein the gel phase is 75% of the total formulation volume, the remainder being a carrier solution.
18 . The hydrogel formulation of claim 1 , wherein the hydrogel formulation further comprising lidocaine.
19 . The hydrogel formulation of claim 1 , wherein a synthetic molecule having the formula: (molecule X) n -(spacer peptide) 0-300 -(five-amino-acid peptide tail) is conjugated to the silk fibroin.
20 . The hydrogel formulation of claim 1 , wherein, upon injection, the hydrogel particles remains substantially at the injection site for one month to eighteen months.
21 . The hydrogel formulation of claim 4 , wherein the said formulation, upon injection, remains substantially at the injection site for five weeks to six weeks and consists of about 1% (w/v) to about 3% (w/v) silk fibroin, about 20% (v/v) to about 50% (v/v) saline, an average particle size range of about 20 μm 2 to about 50 μm 2 .
22 . The hydrogel formulation of claim 4 , wherein the said formulation, upon injection, remains substantially at the injection site for two months to three months and consists of about 3% (w/v) to about 5% (w/v) silk fibroin, about 20% (v/v) to about 50% (v/v) saline, an average particle size range of about 20 μm 2 to about 40 μm 2 .
23 . The hydrogel formulation of claim 4 , wherein the said formulation, upon injection, remains substantially at the injection site for five months to six months and consists of about 4% (w/v) to about 6% (w/v) silk fibroin, about 20% (v/v) to about 50% (v/v) saline, an average particle size range of about 20 μm 2 to about 40 μm 2 .
24 . The hydrogel formulation of claim 4 , wherein the said formulation, upon injection, remains substantially at the injection site for eight months to ten months and consists of about 6% (w/v) to about 8% (w/v) silk fibroin, about 20% (v/v) to about 40% (v/v) saline, an average particle size range of about 20 μm 2 to about 40 μm 2 .
25 . The hydrogel formulation of claim 4 , wherein the said formulation, upon injection, remains substantially at the injection site for eight months to ten months and consists of about 6% (w/v) to about 8% (w/v) silk fibroin, about 20% (v/v) to about 40% (v/v) saline, an average particle size range of about 20 μm 2 to about 40 μm 2 .
26 . The hydrogel formulation of claim 4 , wherein the said formulation, upon injection, remains substantially at the injection site for twelve months to eighteen months and consists of about 8% (w/v) to about 10% (w/v) silk fibroin, about 0% (v/v) to about 40% (v/v) saline, an average particle size range of about 20 μm 2 to about 1000 μm 2 .
27 . A method of making a hydrogel formulation, the method comprising the steps of:
a) obtaining a solution comprising a substantially sericin-free, depolymerized silk fibroin; and
b) adding an enhancer solution comprising an alcohol and an amphiphilic peptide to the silk fibroin solution;
c) rinsing the hydrogel in a solvent to remove residual enhancer solution
d) homogenizing hydrogel in a physiological-acceptable buffer to form hydrogel particles;
28 . The method of claim 27 , wherein the method further comprises step (e) milling the hydrogel particles to a desired particle size.