Cell-Friendly Inverse Opal Hydrogels for Cell Encapsulation, Drug and Protein Delivery, and Functional Nanoparticle Encapsulation

• US 20140178964A1
• Filed: 04/12/2012
• Published: 06/26/2014
• Est. Priority Date: 04/27/2011
• Status: Active Grant

• Current Assignee: President and Fellows of Harvard College (Harvard Corporation)
• Original Assignee: President and Fellows of Harvard College (Harvard Corporation)
• Inventors: Mooney, David J., Kim, Jaeyun, Bencherif, Sidi A., Li, Weiwei A.
• Granted Patent: US 10,647,959 B2
• Time in Patent Office: Days
• US Class Current: 435/182
• CPC Class Codes: A61L 2300/624 Nanocapsules
• CPC Class Codes: A61L 2400/08 Methods for forming porous structures using a negative form which is filled and then removed by pyrolysis or dissolution
• CPC Class Codes: A61L 27/38 containing added animal cells organs or tissue containing native cells A61L27/36
• CPC Class Codes: A61L 27/3834 Cells able to produce different cell types, e.g. hematopoietic stem cells, mesenchymal stem cells, marrow stromal cells, embryonic stem cells
• CPC Class Codes: A61L 27/52 Hydrogels or hydrocolloids
• CPC Class Codes: A61L 27/54 Biologically active materials, e.g. therapeutic substances A61L27/227 takes precedence
• CPC Class Codes: A61L 27/56 Porous materials, e.g. foams or sponges
• CPC Class Codes: B82Y 5/00 Nanobiotechnology or nanomedicine, e.g. protein engineering or drug delivery
• CPC Class Codes: C12N 2533/74 Alginate
• CPC Class Codes: C12N 5/0012 Cell encapsulation

Abstract

The invention provides polymer scaffolds for cell-based tissue engineering.

Claims

1. An inverse opal hydrogel scaffold device comprising a polymer matrix and a sacrificial porogen, wherein said porogen comprises an ionically-crosslinked polymer, a thermosensitive polymer, a thermoresponsive polymer, a pH-sensitive polymer, or a photocleavable polymer.

2. The inverse opal hydrogel of claim 1, wherein said polymer matrix further comprises an isolated eukaryotic cell.

3. The inverse opal hydrogel of claim 1, wherein said polymer matrix is crosslinked and wherein scaffold device further comprises an isolated cell encapsulated in said crosslinked polymer matrix.

4. The inverse opal hydrogel of claim 1, wherein said polymer matrix comprises a synthetic polymer.

5. The inverse opal hydrogel of claim 1, wherein said polymer matrix comprises covalent crosslinking

6. The inverse opal hydrogel of claim 1, wherein said polymer matrix comprises a poly(lactide-coglycolide) (PLGA), poly(acrylic acid), polyethylene glycol (PEG), poly (vinyl alcohol), or polyphosphazene.

7. The inverse opal hydrogel of claim 1, wherein said sacrificial porogen comprises alginate, collagen, gelatin, fibrin, agarose, hyaluronic acid, or chitosan.

8. The inverse opal hydrogel of claim 1, wherein said ionically crosslinked polymer comprises alginate.

9. The inverse opal hydrogel of claim 1, wherein said ionically crosslinked polymer is removed by adding a metal-chelating agent selected from the group consisting of citric acid, ethylenediamine, ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaacetic acid (DTPA), and N,N-bis(carboxymethyl)glycine (NTA).

10. The inverse opal hydrogel of claim 1, wherein said thermosensitive polymer comprises agarose, gelatin, or collagen.

11. The inverse opal hydrogel of claim 1, wherein said thermoresponsive polymer is selected from the group consisting of poly(N-isopropylacrylamide), poly(N-ethylacrylamide), poly(N-cyclopropymethacrylamide), poly(N-methyl-N-ethylacrylamide), poly(N-acryloylpyrrolidine), poly(N-ethylmethacrylamide), poly(N-cyclopropylacrylamide), poly(N-cyclopropylacrylamide), poly(N,N-diethylacrylamide), poly(N-isopropylacrylamide), poly(N-vinylcaprolactam), poly(N-n-propylmethacrylamide), poly(N-methyl-N-isopropylacrylamide), poly(N-n-propylacrylamide), poly(N-methyl-N-n-propylacrylamide), and poly(N-acryloylpiperidine).

12. A method of producing an inverse opal hydrogel with open, interconnected pores comprising:

compressing a plurality of template porogen particles into a mold;

adding a composition comprising a polymer solution and a first plurality of cells to the interstitial space between template porogen particles in said mold to polymerize said template porogen particles;

removing said template porogen particles;

thereby producing an inverse opal hydrogel with open, interconnected pores.

13. The method of claim 12, wherein said template porogen particles are removed without using toxic organic solvents or lyophilization.

14. The method of claim 12, wherein said cells are encapsulated in said inverse opal hydrogel.

15. The method of claim 12, wherein said template porogen particle is an ionically crosslinked polymer, a thermosensitive polymer, a thermoresponsive polymer, a pH-responsive polymer, or a photo-cleavable polymer.

16. The method of claim 15, wherein said ionically crosslinked polymer is alginate.

17. The method of claim 15, wherein said ionically crosslinked polymer is removed by adding a metal-chelating agent selected from the group consisting of citric acid, ethylenediamine, ethylenediaminetetraacetic acid (EDTA), diethylenetriaminepentaacetic acid (DTPA), and N,N-bis(carboxymethyl)glycine (NTA).

18. The method of claim 15, wherein said thermosensitive polymer is agarose, gelatin, or collagen.

19. The method of claim 18, wherein said thermosensitive polymer is removed by increasing the temperature of said polymer, thereby altering the phase of said polymer to liquid phase.

20. The method of claim 15, wherein said thermoresponsive polymer is selected from the group consisting of poly(N-isopropylacrylamide), poly(N-ethylacrylamide), poly(N-cyclopropymethacrylamide), poly(N-methyl-N-ethylacrylamide), poly(N-acryloylpyrrolidine), poly(N-ethylmethacrylamide), poly(N-cyclopropylacrylamide), poly(N-cyclopropylacrylamide), poly(N,N-diethylacrylamide), poly(N-isopropylacrylamide), poly(N-vinylcaprolactam), poly(N-n-propylmethacrylamide), poly(N-methyl-N-isopropylacrylamide), poly(N-n-propylacrylamide), poly(N-methyl-N-n-propylacrylamide), and poly(N-acryloylpiperidine).

21. The method of claim 20, wherein said thermoresponsive polymer is removed by increasing the temperature above a lower critical solution temperature (LCST) to reduce the size of the template particles.

22. The method of claim 12, wherein said open, interconnected pores are 600 μ

m, 1000 μ

m, or 1,500 μ

m.

23. The method of claim 12, wherein said hydrogel is modified with a cell-adhering peptide, wherein said cell-adhering peptide is Arg-Gly-Asp (RGD).

24. The method of claim 12, further comprising adding a second plurality of cells to said open, interconnected pores.

25. The method of claim 12, wherein said first plurality of cells and said second plurality of cells are selected from the group consisting of mesenchymal stem cells, stromal cells, cancer cells, dendritic cells, macrophages, neutrophils, natural killer cells, or fibroblast cells.

26. The method of claim 25, wherein said first plurality of cells and said second plurality of cells are different cell types.

30. A composition comprising an inverse opal hydrogel with open, interconnected pores produced by the method of claim 12.

27. A method of producing an inverse opal hydrogel with open, interconnected pores comprising:

compressing a plurality of template porogen particles into a mold;

adding a composition comprising a polymer solution and an agent to the interstitial space between template porogen particles in said mold to polymerize said template porogen particles;

removing said template porogen particles;

thereby producing an inverse opal hydrogel with open, interconnected pores.

28. The method of claim 27, wherein said agent is selected from the group consisting of a drug, a nanoparticle, a growth factor, a cytokine, a chemokine, a hormone, a protein, a nucleic acid, or a small molecule.

29. The method of claim 28, wherein said nanoparticles are magnetic nanoparticles or gold nanoparticles.

31. A composition comprising an inverse opal hydrogel with open, interconnected pores comprising a first plurality of cells encapsulated in a hydrogel matrix and a second plurality of cells in said open, interconnected pores, wherein said first plurality of cells encapsulated in said hydrogel matrix occupy an interstitial space between said open, interconnected pores.

32. The composition of claim 31, wherein said first plurality of cells and said second plurality of cells are selected from the group consisting of mesenchymal stem cells, stromal cells, cancer cells, dendritic cells, macrophages, neutrophils, natural killer cells, or fibroblast cells.

33. The composition of claim 31, wherein said hydrogel comprises gelatin or poly(ethylene glycol) (PEG).