Cell-Laden Hydrogels

• US 20080193536A1
• Filed: 08/14/2007
• Published: 08/14/2008
• Est. Priority Date: 08/14/2006
• Status: Abandoned Application

• Current Assignee: Alireza Khademhosseini, Jeffrey M. Karp, Omid C. Farokhzad, Robert S. Langer
• Original Assignee: Alireza Khademhosseini, Jeffrey M. Karp, Omid C. Farokhzad, Robert S. Langer
• Inventors: Farokhzad, Omid C., Langer, Robert S., Karp, Jeffrey M., Khademhosseini, Alireza
• Application Number:
  US11/838,752
• Publication Number:
  US 20080193536A1
• Time in Patent Office: Days
• US Class Current: 424/486
• CPC Class Codes: A61K 35/33 Fibroblasts
• CPC Class Codes: A61K 35/545 Embryonic stem cells; Pluripotent stem cells; Induced pluripotent stem cells; Uncharacterised stem cells
• CPC Class Codes: A61K 47/06 Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
• CPC Class Codes: A61K 9/06 Ointments; Bases therefor; Other semi-solid forms, e.g. creams, sticks, gels composition of ointments, creams or gels A61K47/00
• CPC Class Codes: A61L 27/38 containing added animal cells organs or tissue containing native cells A61L27/36
• CPC Class Codes: A61L 27/3886 comprising two or more cell types
• CPC Class Codes: A61L 27/52 Hydrogels or hydrocolloids

Abstract

The present invention provides cell-laden hydrogels and hydrogel assemblies thereof for use in tissue engineering. The invention provides microscale hydrogels (i.e. microgels) having greatest dimensions ranging between about 1 μm and 1000 μm. The present invention provides methods of producing inventive hydrogels and hydrogel assemblies and pharmaceutical compositions thereof.

Claims

1. A hydrogel comprising a crosslinked polymeric network, wherein cells are encapsulated within the hydrogel, and wherein the hydrogel has a greatest dimension ranging between about 1 μ

m and about 1000 μ

m.

2. The hydrogel of claim 1, wherein the hydrogel has a greatest dimension ranging between about 50 μ

m and about 1000 μ

m.

3. The hydrogel of claim 1, wherein the hydrogel has a greatest dimension ranging between about 100 μ

m and about 1000 μ

m.

4. The hydrogel of claim 1, wherein the hydrogel has a greatest dimension of about 50 μ

m.

5. The hydrogel of claim 1, wherein the hydrogel has a greatest dimension of about 100 μ

m.

6. The hydrogel of claim 1, wherein the hydrogel has a greatest dimension of about 250 μ

m.

7. The hydrogel of claim 1, wherein the hydrogel has a greatest dimension of about 500 μ

m.

8. The hydrogel of claim 1, wherein the hydrogel has a greatest dimension of about 750 μ

m.

9. The hydrogel of claim 1, wherein the hydrogel has a greatest dimension of about 1000 μ

m.

10. The hydrogel of claim 1, wherein the cells are distributed evenly throughout the hydrogel.

11. The hydrogel of claim 1, wherein the cells are selected from the group consisting of cardiomyocytes, myocytes, hepatocytes, keratinocytes, melanocytes, neurons, astrocytes, embryonic stem cells, adult stem cells, hematopoietic stem cells, hematopoietic cells, monocytes, neutrophils, macrophages, ameloblasts, fibroblasts, chondrocytes, osteoblasts, osteoclasts, neurons, sperm cells, egg cells, liver cells, epithelial cells from lung, epithelial cells from gut, epithelial cells from intestine, liver, epithelial cells from skin, and hybrids thereof.

12. The hydrogel of claim 1, wherein the hydrogel comprises an environment that promotes cell viability.

13. The hydrogel of claim 1, wherein the hydrogel comprises one type of cell.

14. The hydrogel of claim 1, wherein the hydrogel comprises more than one type of cell.

15. The hydrogel of claim 1, wherein the polymeric network comprises a natural polymer.

16. The hydrogel of claim 1, wherein the polymeric network comprises a carbohydrate.

17. The hydrogel of claim 1, wherein the polymeric network comprises a carbohydrate selected from the group consisting of hyaluronic acid (HA), chondroitin sulphate, dermatan sulphate, keratan sulphate, heparan sulphate, derivatives thereof, and combinations thereof.

18. The hydrogel of claim 1, wherein the polymeric network comprises a carbohydrate selected from the group consisting of alginate, chitosan, agarose, heparin, dextran, derivatives thereof, and combinations thereof.

19. The hydrogel of claim 1, wherein the polymeric network comprises a protein or peptide.

20. The hydrogel of claim 1, wherein the polymeric network comprises a protein selected from the group consisting of collagen, elastin, fibrin, derivatives thereof, and combinations thereof.

21. The hydrogel of claim 1, wherein the polymeric network comprises a synthetic polymer.

22. The hydrogel of claim 1, wherein the polymeric network comprises a polymer selected from the group consisting of poly(ethylene glycol) (PEG), poly(hydroxyethyl methacrylate) (PHEMA), poly(vinyl alcohol) (PVA), derivatives thereof, and combinations thereof.

23. The hydrogel of claim 1, wherein the polymeric network comprises one type of polymer.

24. The hydrogel of claim 1, wherein the polymeric network comprises more than one type of polymer.

25. The hydrogel of claim 1, wherein the polymeric network is crosslinked using a chemical crosslinking agent.

26. The hydrogel of claim 25, wherein the polymeric network is crosslinked using a chemical crosslinking agent selected from the group consisting of electrophiles and nucleophiles.

27. The hydrogel of claim 25, wherein the polymeric network is crosslinked using a chemical crosslinking agent selected from the group consisting of glutaraldehyde, acetaldehyde, formaldehyde, derivatives thereof, and combinations thereof.

28. The hydrogel of claim 1, wherein the polymeric network is crosslinked by altering pH such that crosslinking occurs.

29. The hydrogel of claim 1, wherein the polymeric network is crosslinked by an ionic crosslinking mechanism.

30. The hydrogel of claim 29, wherein the polymeric network is crosslinked due to formation of ionic bridges between divalent cations and the polymer.

31. The hydrogel of claim 29, wherein the polymeric network is crosslinked due to interactions between cations and negatively charged functional groups.

32. The hydrogel of claim 1, wherein the polymeric network is crosslinked by a physical crosslinking mechanism.

33. The hydrogel of claim 32, wherein the polymeric network is crosslinked by performing repeated cycles of freezing and thawing.

34. The hydrogel of claim 1, wherein the polymeric network is crosslinked using irradiative crosslinking mechanisms.

35. The hydrogel of claim 34, wherein the polymeric network is crosslinked using electron beam irradiation, gamma irradiation, or combinations thereof.

36. The hydrogel of claim 1, wherein the polymeric network is crosslinked using photocrosslinking mechanisms.

37. The hydrogel of claim 36, wherein the polymeric network is crosslinked by a photoinitiator and ultraviolet (UV) light.

38. The hydrogel of claim 37, wherein the photoinitiator is 2-hydroxy-1-(4-(hydroxyethoxy)phenyl)-2-methyl-1-propanone.

39. The hydrogel of claim 1, wherein the polymeric network is crosslinked using thermal crosslinking mechanisms.

40. The hydrogel of claim 1, wherein the polymeric network is crosslinked by a thermal initiator and heat.

41. The hydrogel of claim 40, wherein the thermal initiator is selected from the group consisting of peroxides, peracids, peracetates, and persulfates.

42. The hydrogel of claim 1, wherein a therapeutic agent to be delivered is encapsulated within the hydrogel.

43. The hydrogel of claim 1, wherein the hydrogel is biocompatible, biodegradable, or both.

44. The hydrogel of claim 1, wherein the hydrogel is characterized by a cuboid, rectangular, spherical, conical, pyramid-like, cylindrical, tubular, ring-shaped, tetrahedral, hexagonal, or octagonal shape.

45. A hydrogel assembly comprising a plurality of hydrogels, wherein hydrogel comprise a crosslinked polymeric network, wherein cells are encapsulated within hydrogels, and wherein hydrogels has a greatest dimension ranging between about 1 μ

m and about 1000 μ

m.

46. A method comprising:

providing a suspension of cells;

providing a polymer capable of forming hydrogels;

mixing the suspension of cells and the polymer to form a precursor solution;

pouring the precursor solution into stamps, wherein the stamps comprise a plurality of wells and ridges;

crosslinking the precursor solution inside the molds such that hydrogels are formed, wherein the hydrogels have a greatest dimension ranging between about 1 μ

m and about 1000 μ

m; and

harvesting the hydrogels.

47. The method of claim 46, wherein the concentration of cells within the precursor solution ranges between about 0.1% and about 80%.

48. The method of claim 46, wherein the concentration of cells within the precursor solution ranges between about 1.0% and about 50%.

49. The method of claim 46, wherein the concentration of cells within the precursor solution ranges between about 1.0% and about 20%.

50. The method of claim 46, wherein the concentration of cells within the precursor solution ranges between about 1.0% and about 10%.

51. The method of claim 46, wherein the concentration of cells within the precursor solution is about 5%.

52. The method of claim 46, wherein the concentration of polymer within the precursor solution ranges between about 1% and about 40%.

53. The method of claim 46, wherein the concentration of polymer within the precursor solution ranges between about 1% and about 20%.

54. The method of claim 46, wherein the concentration of polymer within the precursor solution ranges between about 1% and about 10%.

55. The method of claim 46, wherein the concentration of polymer within the precursor solution is about 5%.

56. The method of claim 46, wherein the precursor solution further comprises a chemical crosslinking agent, photoinitiator, thermal initiator, or combinations thereof.

57. The method of claim 46, wherein the stamps comprise poly(dimethylsiloxane) (PDMS).

58. The method of claim 57, wherein the surfaces of the PDMS stamps are made to be hydrophilic by plasma cleaning.

59. The method of claim 46, wherein the hydrogels are harvested by rinsing the stamps with an aqueous solution, water-miscible solvent, or combination thereof.

60. The method of claim 46, further comprising steps of:

assembling the hydrogels such that a hydrogel assembly is formed; and

performing a second crosslinking step, wherein the second crosslinking step is performed in order to crosslink individual hydrogels to one another.

61. A method, comprising:

providing a plurality of microgels;

assembling the plurality of microgels such that a hydrogel assembly is formed.

62. The method of claim 61, wherein all of the microgels of the plurality of microgels are identical to one another.

63. The method of claim 61, wherein all of the microgels of the plurality of microgels are not identical to one another.

64. The method of claim 61, wherein the hydrogel assembly is formed by a self-assembly mechanism.

65. The method of claim 61, wherein microgel surfaces are modified in a way that promotes self-assembly.

66. The method of claim 65, wherein modified microgel surfaces can interact with one another via hydrophobic forces, electrostatic forces, polymer chain entanglement, or affinity interactions.

67. The method of claim 65, wherein microgel surfaces are modified with one or more functional groups.

68. The method of claim 67, wherein the functional group is selected from the group consisting of carboxyl groups, amino groups, and hydroxyl groups.

69. The method of claim 68, wherein microgel surfaces are modified with two or more substances that specifically bind to one another with high affinity.

70. The method of claim 69, wherein microgel surfaces are modified with proteins, nucleic acids, lipids, carbohydrates, metals, small molecules, or drugs which are capable of participating in affinity interactions, such that individual microgels bind to one another via affinity interactions.

71. The method of claim 65, wherein microgel shape is controlled in a way that promotes self-assembly via a geometric lock-and-key mechanism.

72. The method of claim 65, wherein hydrogel assemblies are formed utilizing surface tension mechanisms.

73. The method of claim 72, wherein surface tension mechanisms involve placing individual microgels within a hydrophobic bath such that a two-phase system is developed, wherein each microgel is separated from the oil phase by a surface tension, and wherein surface tension is minimized when the microgels self-assemble.

74. The method of claim 61, wherein the hydrogel assembly is formed manually.

75. The method of claim 74, wherein the hydrogel assembly is formed manually using a micromanipulator.

76. The method of claim 61, further comprising an additional crosslinking step performed after hydrogel assembly has occurred, wherein the additional crosslinking step is performed in order to crosslink individual microgels to one another.

77. A method comprising:

providing a subject in need of tissue regeneration; and

administering one or more cell-laden hydrogels or cell-laden hydrogel assemblies to the subject in order to promote tissue regeneration.

78. A method comprising:

providing a subject having a location in need of tissue regeneration; and

providing a precursor solution comprising cells and at least one polymer capable of forming a hydrogel;

administering the precursor solution in situ to the subject at or near the location in need of tissue regeneration; and

crosslinking the precursor solution in situ such that a hydrogel is formed at or near the location in need of tissue regeneration.