Materials and methods for nerve grafting

• US 20030077258A1
• Filed: 08/13/2002
• Published: 04/24/2003
• Est. Priority Date: 08/13/2001
• Status: Abandoned Application

• Current Assignee: University of Florida Research Foundation, Incorporated (State University System of Florida)
• Original Assignee: University of Florida Research Foundation, Incorporated (State University System of Florida)
• Inventors: Muir, David F.
• Application Number:
  US10/218,315
• Publication Number:
  US 20030077258A1
• Time in Patent Office: Days
• US Class Current: 424/93.7
• CPC Class Codes: A61K 2300/00 Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00
• CPC Class Codes: A61K 35/30 Nerves; Brain; Eyes; Corneal cells; Cerebrospinal fluid; Neuronal stem cells; Neuronal precursor cells; Glial cells; Oligodendrocytes; Schwann cells; Astroglia; Astrocytes; Choroid plexus; Spinal cord tissue
• CPC Class Codes: A61K 38/51 Lyases (4)
• CPC Class Codes: A61L 2430/32 for nerve reconstruction
• CPC Class Codes: A61L 27/227 Other specific proteins or polypeptides not covered by A61L27/222, A61L27/225 or A61L27/24
• CPC Class Codes: A61L 27/3604 characterised by the human or animal origin of the biological material, e.g. hair, fascia, fish scales, silk, shellac, pericardium, pleura, renal tissue, amniotic membrane, parenchymal tissue, fetal tissue, muscle tissue, fat tissue, enamel
• CPC Class Codes: A61L 27/3675 Nerve tissue, e.g. brain, spinal cord, nerves, dura mater
• CPC Class Codes: A61L 27/3683 subjected to a specific treatment prior to implantation, e.g. decellularising, demineralising, grinding, cellular disruption/non-collagenous protein removal, anti-calcification, crosslinking, supercritical fluid extraction, enzyme treatment
• CPC Class Codes: A61L 27/3687 characterised by the use of chemical agents in the treatment, e.g. specific enzymes, detergents, capping agents, crosslinkers, anticalcification agents
• CPC Class Codes: A61L 27/383 Nerve cells, e.g. dendritic cells, Schwann cells
• CPC Class Codes: A61L 27/3878 Nerve tissue, brain, spinal cord, nerves, dura mater
• CPC Class Codes: A61L 31/047 Other specific proteins or polypeptides not covered by A61L31/044 - A61L31/046
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 25/02 for peripheral neuropathies
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00

Abstract

The subject invention pertains to compositions and methods for promoting repair of damaged nerve tissue using nerve grafts and preparation of nerve grafts. The compositions and methods of the subject invention can be employed to restore the continuity of nerve interrupted by disease, traumatic events or surgical procedures. Compositions of the subject invention comprise one or more chondroitin sulfate proteoglycan (CSPG)-degrading enzymes that promote axonal penetration into damaged nerve tissue and nerve graft. The invention also concerns methods for promoting repair of damaged nerve tissue using the present compositions and nerve tissue treated according to such methods. The invention also includes storage solutions for nerve tissue.

Claims

1. A method for preparing a nerve graft for implantation comprising applying at least one chondroitin sulfate proteoglycan-degrading enzyme to the nerve graft.

2. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme is selected from the group consisting of chondroitinase, hyalurondiase, and matrix metalloproteinase, or a combination thereof.

3. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme is selected from the group consisting of chondroitinase ABC, chondroitinase A, chondroitinase C, chondroitinase AC, hyaluronidase, matrix metalloproteinase-2, and matrix metalloproteinase-9, or a combination thereof.

4. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme is selected from the group consisting of chondroitinase ABC, chondroitinase A, chondroitinase C, and chondroitinase AC or a combination thereof.

5. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a chondroitinase.

6. The method according to claim 1, wherein the nerve graft has a first end and a second end, and wherein the chondroitin sulfate proteoglycan-degrading enzyme is applied to first end of the nerve graft, or to the second end of the nerve graft, or to both the first end and second end of the nerve graft.

7. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme is applied to the nerve graft en bloc.

8. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme is applied topically to the nerve graft.

9. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme is injected into the nerve graft.

10. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme is applied to the nerve graft by placing the nerve graft in a culture medium containing the chondroitin sulfate proteoglycan-degrading enzyme.

11. The method according to claim 10, wherein the culture medium is a defined medium.

12. The method according to claim 10, wherein the culture medium is an undefined medium.

13. The method according to claim 1, wherein said method further comprises applying a tissue adhesive to the nerve graft.

14. The method according to claim 13, wherein the tissue adhesive is a biological glue.

15. The method according to claim 13, wherein the tissue adhesive is fibrin glue.

16. The method according to claim 13, wherein the tissue adhesive is platelet gel.

17. The method according to claim 1, wherein said method further comprises applying a biologically active agent to the nerve graft.

18. The method according to claim 17, wherein the biologically active agent is a growth factor.

19. The method according to claim 18, wherein the growth factor is selected from the group consisting of nerve growth factor, fibroblast growth factor-1, fibroblast growth factor-2, epidermal growth factor, ciliary neurotrophic factor, brain-derived neurotrophic factor, neurotrophin-3, neurotrophin-4, neurotrophin-5, insulin-like growth factor-1, insulin-like growth factor-2, transforming growth factor, glial growth factor-2, vascular endothelial growth factor, granulocyte-macrophage colony stimulating factor, and lymphocyte infiltrating factor/cholinergic differentiating factor.

20. The method according to claim 1, wherein said method further comprises applying cells to the nerve graft.

21. The method according to claim 20, wherein the cells are stem cells.

22. The method according to claim 21, wherein the cells are Schwann cells.

23. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme causes lysis of chondroitin-4-sulfate, chondroitin-6-sulfate, or both chondroitin-4-sulfate and chondroitin-6-sulfate within the nerve graft.

24. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme decreases the latency of axonal ingress into the nerve graft once the nerve graft is coapted into a damaged nerve.

25. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme increases the rate of axonal ingress into the nerve graft once the nerve graft is coapted into a damaged nerve.

26. The method according to claim 1, wherein the nerve graft comprises peripheral nerve tissue.

27. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a chondroitinase, and wherein the chondroitinase is applied to the nerve graft within a concentration range from about 10 units/mL to about 1000 units/mL.

28. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a chondroitinase, and wherein the chondroitinase is applied to the nerve graft within a concentration range from about 100 units/mL to about 500 units/mL.

29. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a matrix metalloproteinase, and wherein the matrix metalloproteinase is applied to the nerve graft within a concentration range from about 0.1 μ

g/mL to about 100 μ

g/mL.

30. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a matrix metalloproteinase, and wherein the matrix metalloproteinase is applied to the nerve graft within a concentration range from about 10 μ

g/mL to about 50 μ

g/mL.

31. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a bacterial enzyme.

32. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a mammalian enzyme.

33. The method according to claim 1, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a recombinant enzyme.

34. The method according to claim 1, wherein the nerve graft is within the range of about 1 centimeter to about 10 centimeters in length.

35. The method according to claim 1, wherein the nerve graft is greater than about 10 centimeters in length.

36. The method according to claim 1, wherein the nerve graft is a living nerve graft.

37. The method according to claim 1, wherein said nerve graft is a living graft and wherein said method further comprises rendering the living nerve graft acellular.

38. The method according to claim 37, wherein the nerve graft is a living graft, and wherein the living nerve graft is rendered acellular before applying the chondroitin sulfate proteoglycan-degrading enzyme to the nerve graft.

39. The method according to claim 1, wherein the nerve graft is a living graft, and wherein said method further comprises rendering the living nerve graft acellular by freeze-killing the nerve graft.

40. The method according to claim 1, wherein the nerve graft is a living graft, and wherein said method further comprises rendering the living nerve graft acellular by chemical extraction with detergents.

41. The method according to claim 1, wherein the nerve graft is derived from a mammal.

42. The method according to claim 1, wherein the nerve graft is derived from a mammal selected from the group consisting of human, non-human primate, porcine, rodent, and bovine.

43. The method according to claim 1, wherein the nerve graft is an autograft.

44. The method according to claim 1, wherein the nerve graft is an allograft.

45. The method according to claim 1, wherein the nerve graft is a xenograft.

46. A method for promoting repair of a damaged nerve, wherein said method comprises implanting a nerve graft at the site of damage and applying at least one chondroitin sulfate proteoglycan-degrading enzyme to:

(a) the nerve graft;

or

(b) the damaged nerve;

or

(c) both the nerve graft and the damaged nerve.

47. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is selected from the group consisting of chondroitinase, hyalurondiase, and matrix metalloproteinase, or a combination thereof.

48. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is selected from the group consisting of chondroitinase ABC, chondroitinase A, chondroitinase C, chondroitinase AC, hyaluronidase, matrix metalloproteinase-2, and matrix metalloproteinase-9, or a combination thereof.

49. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is selected from the group consisting of chondroitinase ABC, chondroitinase A, chondroitinase C, chondroitinase AC, or a combination thereof.

50. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a chondroitinase.

51. The method according to claim 46, wherein said method further comprises applying a tissue adhesive to:

(a) the nerve graft;

or

(b) the damaged nerve;

or

(c) both the nerve graft and the damaged nerve.

52. The method according to claim 51, wherein the tissue adhesive is a biological glue.

53. The method according to claim 51, wherein the tissue adhesive is selected from the group consisting of fibrin glue and platelet gel.

54. The method according to claim 46, wherein said method further comprises applying a biologically active agent to:

(a) the nerve graft;

or

(b) the damaged nerve;

or

(c) both the nerve graft and the damaged nerve.

55. The method according to claim 54, wherein the biologically active agent is a growth factor.

56. The method according to claim 55, wherein the growth factor is selected from the group consisting of nerve growth factor, fibroblast growth factor-1, fibroblast growth factor-2, epidermal growth factor, ciliary neurotrophic factor, brain-derived neurotrophic factor, neurotrophin-3, neurotrophin-4, neurotrophin-5, insulin-like growth factor-1, insulin-like growth factor-2, transforming growth factor, glial growth factor-2, vascular endothelial growth factor, granulocyte-macrophage colony stimulating factor, and lymphocyte infiltrating factor/cholinergic differentiating factor.

57. The method according to claim 46, wherein said method further comprises applying cells to:

(a) the nerve graft;

or

(b) the damaged nerve;

or

(c) both the nerve graft and the damaged nerve.

58. The method according to claim 57, wherein the cells are stem cells.

59. The method according to claim 57, wherein the cells are Schwann cells.

60. The method according to claim 46, wherein the nerve damage comprises a disruption in the continuity of at least one of the nerve sheaths of the damaged nerve selected from the group consisting of basal lamina, perineurium, and epineurium.

61. The method according to claim 46, wherein the nerve damage is a neurotmesis.

62. The method according to claim 46, wherein the nerve damage is a neurotmesis, wherein said implanting comprises coapting the nerve graft into the damaged nerve, and wherein the chondroitin sulfate proteoglycan-degrading enzyme is applied to the site of coaptation.

63. The method according to claim 46, wherein the damaged nerve is partially or completely severed, having a proximal end and a distal end, wherein the nerve graft has a first end and a second end, and wherein said implanting comprises interposing the nerve graft between the proximal end and distal end of the damaged nerve, wherein the chondroitin sulfate proteoglycan-degrading enzyme is applied to at least one of the following:

(a) the proximal end of the damaged nerve;

(b) the distal end of the damaged nerve;

(c) the first end of the nerve graft; and

(d) the distal end of the nerve graft.

64. The method according to claim 63, wherein said method further comprises applying a tissue adhesive to at least one of the following:

(a) the proximal end of the damaged nerve;

(b) the distal end of the damaged nerve;

(c) the first end of the nerve graft; and

(d) the distal end of the nerve graft.

65. The method according to claim 46, wherein said implanting comprising coapting the nerve graft into the damaged nerve and suturing the nerve graft to the damaged nerve.

66. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme causes lysis of chondroitin-4-sulfate, chondroitin-6-sulfate, or both chondroitin-4-sulfate, and chondroitin-6-sulfate.

67. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme decreases the latency of axonal ingress from the damaged nerve into the nerve graft.

68. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme increases the rate of axonal ingress from the damaged nerve into the nerve graft.

69. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is applied topically.

70. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is applied by injection.

71. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is applied to the nerve graft en bloc.

72. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is applied with a pharmaceutically acceptable carrier.

73. The method according to claim 72, wherein the pharmaceutically acceptable carrier is selected from the group consisting of liquids, gels, foams, solids, and polymers.

74. The method according to claim 72, wherein the pharmaceutically acceptable carrier is selected from the group consisting of membranes, sponges, fibrous structures, powders, fleeces, and particles.

75. The method according to claim 72, wherein the pharmaceutically acceptable carrier is a controlled release system.

76. The method according to claim 72, wherein the pharmaceutically acceptable carrier is a surgical cuff, wherein the surgical cuff at least partially encircles the implanted nerve graft.

77. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a chondroitinase, and wherein the chondroitinase is applied to the damaged nerve within a concentration range from about 10 units/mL to about 1000 units/mL.

78. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a chondroitinase, and wherein the chondroitinase is applied to the damaged nerve within a concentration range from about 100 units/mL to about 500 units/mL.

79. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a matrix metalloproteinase, and wherein the matrix metalloproteinase is applied to the damaged nerve within a concentration range from about 0.1 μ

g/mL to about 100 μ

g/mL.

80. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a matrix metalloproteinase, and wherein the matrix metalloproteinase is applied to the damaged nerve within a concentration range from about 10 μ

g/mL to about 50 μ

g/mL.

81. The method according to claim 46, wherein the damaged nerve is a damaged peripheral nerve.

82. The method according to claim 46, wherein the damaged nerve is a nerve of the central nervous system and the nerve graft comprises peripheral nerve tissue.

83. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a bacterial enzyme.

84. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a mammalian enzyme.

85. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is a recombinant enzyme.

86. The method according to claim 46, wherein said application of chondroitin sulfate proteoglycan-degrading enzyme comprises administering a genetically modified host to the damaged nerve, or to the nerve graft, or to both the damaged nerve and the nerve graft, wherein the host has been genetically modified with a polynucleotide encoding the chondroitin sulfate proteoglycan-degrading enzyme.

87. The method according to claim 86, wherein the genetically modified host is a cell.

88. The method according to claim 46, wherein the nerve graft is autologous with the damaged nerve.

89. The method according to claim 46, wherein the nerve graft is allogeneic with the damaged nerve.

90. The method according to claim 46, wherein the nerve graft is xenogeneic with the damaged nerve.

91. The method according to claim 46, wherein the nerve graft is derived from a mammal.

92. The method according to claim 46, wherein the nerve graft is derived from a mammal selected from the group consisting of human, non-human primate, porcine, rodent, and bovine.

93. The method according to claim 46, wherein the damaged nerve is human and the nerve graft is derived from a mammal selected from the group consisting of non-human primate, porcine, rodent, and bovine.

94. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme is applied to the nerve graft before said implanting.

95. The method according to claim 46, wherein the chondroitin sulfate proteoglycan-degrading enzyme applied to the nerve graft after said implanting.

96. The method according to claim 46, wherein the nerve graft comprises a living graft.

97. The method according to claim 46, wherein the nerve graft comprises a living graft, and wherein said method further comprising rendering the living nerve graft acellular before said implanting of the acellular nerve graft.

98. The method according to claim 46, wherein said method further comprises resecting the damaged nerve before said implanting of the nerve graft.

99. The method according to claim 46, wherein the nerve graft is a terminal nerve graft.

100. The method according to claim 46, wherein the nerve graft is an interpositional nerve graft.

101. The method according to claim 46, wherein the nerve damage was caused by disease.

102. The method according to claim 46, wherein the nerve damage was caused by trauma.

103. The method according to claim 46, wherein the nerve damage was caused by a surgical procedure.

104. A method for promoting repair of a damaged nerve, wherein the nerve damage comprises a disruption in the continuity of at least one of the nerve sheaths of the damaged nerve selected from the group consisting of basal lamina, perineurium, and epineurium, wherein said method comprises implanting a nerve graft into the damaged nerve, and applying chondroitinase to the nerve graft, or to the damaged nerve, or to both the nerve graft and the damaged nerve.

105. The method according to claim 104, wherein said method further comprises applying a tissue adhesive to the nerve graft, or to the damaged nerve, or to both the nerve graft and the damaged nerve.

106. The method according to claim 105, wherein the chondroitinase and the tissue adhesive are applied within the same formulation.

107. The method according to claim 105, wherein the tissue adhesive is selected from the group consisting of fibrin glue and platelet gel.

108. The method according to claim 104, wherein the chondroitinase is selected from the group consisting of chondroitinase ABC, chondroitinase A, chondroitinase C, chondroitinase AC, or a combination thereof.

109. The method according to claim 104, wherein the nerve graft comprises peripheral nerve tissue.

110. The method according to claim 104, wherein the nerve graft is autologous with the damaged nerve.

111. The method according to claim 104, wherein the nerve graft is allogeneic with the damaged nerve.

112. The method according to claim 104, wherein the nerve graft is xenogeneic with the damaged nerve.

113. The method according to claim 104, wherein the nerve graft is derived from a mammal.

114. The method according to claim 104, wherein the nerve graft is derived from a mammal selected from the group consisting of human, non-human primate, porcine, rodent, and bovine.

115. The method according to claim 104, wherein the damaged nerve is human and the nerve graft is derived from a mammal selected from the group consisting of non-human primate, porcine, rodent, and bovine.

116. The method according to claim 104, wherein the nerve graft is a terminal nerve graft.

117. The method according to claim 104, wherein the nerve graft is an interpositional nerve graft.

118. A nerve graft comprising nerve tissue prepared by a method comprising applying at least one chondroitin sulfate proteoglycan-degrading enzyme to said nerve tissue.

119. The nerve graft of claim 118, wherein said chondroitin sulfate proteoglycan-degrading enzyme is selected from the group consisting of chondroitinase, hyalurondiase, and matrix metalloproteinase, or a combination thereof.

120. The nerve graft of claim 118, wherein said chondroitin sulfate proteoglycan-degrading enzyme is selected from the group consisting of chondroitinase ABC, chondroitinase A, chondroitinase C, chondroitinase AC, hyaluronidase, matrix metalloproteinase-2, and matrix metalloproteinase-9, or a combination thereof.

121. The nerve graft of claim 118, wherein said chondroitin sulfate proteoglycan-degrading enzyme is selected from the group consisting of chondroitinase ABC, chondroitinase A, chondroitinase C, chondroitinase AC, or a combination thereof.

122. The nerve graft of claim 118, wherein said chondroitin sulfate proteoglycan-degrading enzyme is a chondroitinase.

123. The nerve graft of claim 118, wherein said method further comprising applying a tissue adhesive to said nerve tissue.

124. The nerve graft of claim 118, wherein said tissue is selected from the group consisting of fibrin glue and platelet gel.

125. The nerve graft of claim 118, wherein said nerve tissue comprises living tissue, and wherein said method further comprises rendering said living nerve tissue acellular.

126. The nerve graft of claim 125, wherein said rendering said nerve tissue acellular comprises freeze-killing said nerve tissue.

127. The nerve graft of claim 118, wherein said nerve tissue is peripheral nerve tissue.

128. The nerve graft of claim 118, wherein the nerve graft is a terminal nerve graft.

129. The nerve graft of claim 118, wherein the nerve graft is an interpositional nerve graft.

130. The nerve graft of claim 118, wherein said nerve tissue is mammalian tissue.

131. The nerve graft of claim 118, wherein the nerve tissue is selected from the group consisting of human tissue, non-human primate tissue, porcine tissue, rodent tissue, and bovine tissue.

132. The nerve graft of claim 130, wherein said nerve tissue is human tissue.

133. The nerve graft of claim 118, wherein the nerve graft is an autograft.

134. The nerve graft of claim 118, wherein the nerve graft is an allograft.

135. The nerve graft of claim 118, wherein the nerve graft is a xenograft.

136. A storage solution for a nerve graft comprising at least one chondroitin sulfate proteoglycan-degrading enzyme and a culture medium.

137. The storage solution of claim 136, wherein said chondroitin sulfate proteoglycan-degrading enzyme is selected from the group consisting of chondroitinase, hyalurondiase, and matrix metalloproteinase, or a combination thereof.

138. The storage solution of claim 136, wherein said chondroitin sulfate proteoglycan-degrading enzyme is selected from the group consisting of chondroitinase ABC, chondroitinase A, chondroitinase C, chondroitinase AC, hyaluronidase, matrix metalloproteinase-2, and matrix metalloproteinase-9, or a combination thereof.

139. The storage solution of claim 136, wherein said chondroitin sulfate proteoglycan-degrading enzyme is selected from the group consisting of chondroitinase ABC, chondroitinase A, chondroitinase C, chondroitinase AC, or a combination or subcombination thereof, or a biologically active fragment or variant thereof.

140. The storage solution of claim 136, wherein said chondroitin sulfate proteoglycan-degrading enzyme is a chondroitinase.

141. The storage solution of claim 136, wherein said culture medium comprises defined medium.

142. The storage solution of claim 136, wherein said culture medium comprises defined medium.

143. The storage solution of claim 136, wherein said culture medium comprises undefined medium.