Methods and compositions for regenerating connective tissue
View Patent ↗Connective tissue regenerative compositions and methods of repairing and regenerating connective tissue using such compositions are provided. The compositions generally comprise a bioactive hydrogel matrix comprising a polypeptide, such as gelatin, and a long chain carbohydrate, such as dextran. The hydrogel matrix may further include polar amino acids, as well as additional beneficial additives. Advantageously, the compositions include further components, such as osteoinductive or osteoconductive materials, medicaments, stem or progenitor cells, and three-dimensional structural frameworks. The compositions are useful for regenerating connective tissue, and can be administered to an area having injury to, or a loss of, connective tissue, such as bone, cartilage, tendon, and ligament.
1. A connective tissue regenerative composition for administration at a site in need of connective tissue regeneration, the composition comprising:
a) a bioactive hydrogel matrix that is flowable at physiological temperatures comprising:
i. a tissue-derived polypeptide derived from tissue selected from the group consisting of collagens, gelatins, keratin, decorin, aggrecan, and glycoproteins, or a tissue-derived polypeptide derived from extracts of tissue selected from the group consisting of submucosal tissues, arteries, vocal chords, pleura, trachea, bronchi, pulmonary alveolar septa, ligaments, auricular cartilage, abdominal fascia, liver, kidney, neurilemma, arachnoid, dura mater, and pia mater,
ii. a polysaccharide or a sulfated polysaccharide, and
iii. one or more components selected from the group consisting of polar amino acids, divalent cation chelators, and combinations thereof; and
b) a three-dimensional structural framework at least partially containing the bioactive hydrogel matrix and at least partially retaining the hydrogel matrix at the site in need of connective tissue regeneration, the structural framework providing support during connective tissue regeneration and simultaneously providing a space or network of spaces for cellular infiltration;
wherein the three-dimensional structural framework is a crosslinked hydrogel formed of at least two polymers crosslinked to one another and that is solid or semi-solid at physiological temperatures, the crosslinked hydrogel comprising one or more components selected from the group consisting of polar amino acids, divalent cation chelators, and combinations thereof.
2. The composition of claim 1 , wherein the tissue-derived polypeptide has a molecular mass of about 3,000 to about 3,000,000 Da.
3. The composition of claim 2 wherein the tissue-derived polypeptide has a molecular mass of about 30,000 to about 300,000 Da.
4. The composition of claim 1 , wherein the polysaccharide or sulfated polysaccharide is a polysaccharide comprising more than about 10 monosaccharide residues joined to each other by glycosidic linkages.
5. The composition of claim 1 , wherein the polysaccharide or sulfated polysaccharide is selected from the group consisting of glycosaminoglycans and glucosaminoglycans.
6. The composition of claim 1 , wherein the polysaccharide or sulfated polysaccharide is a polysaccharide selected from the group consisting of dextran, heparan, heparin, hyaluronic acid, alginate, agarose, carageenan, amylopectin, amylose, glycogen, starch, cellulose, chitin, and chitosan.
7. The composition of claim 1 , wherein the polysaccharide or sulfated polysaccharide is a sulfated polysaccharide selected from the group consisting of heparan sulfate, chondroitin sulfate, dextran sulfate, dermatan sulfate, and keratan sulfate.
8. The composition of claim 1 , wherein the polysaccharide or sulfated polysaccharide has a molecular mass of about 2,000 to about 8,000,000 Da.
9. The composition of claim 8 , wherein the polysaccharide or sulfated polysaccharide has a molecular mass of about 20,000 to about 1,000,000 Da.
10. The composition of claim 1 , wherein the tissue-derived polypeptide is gelatin and the polysaccharide or sulfated polysaccharide is dextran.
11. The composition of claim 10 , wherein the gelatin is skin-derived gelatin.
12. The composition of claim 10 , wherein the gelatin is bone-derived gelatin.
13. The composition of claim 10 , wherein the gelatin has a molecular mass of about 80,000 to about 200,000 Da.
14. The composition of claim 13 , wherein the polydispersity of the molecular mass of the gelatin is about 1 to about 3.
15. The composition of claim 14 , wherein the polydispersity of the molecular mass of the gelatin is about 1.1 to about 2.4.
16. The composition of claim 10 , wherein the gelatin is present at a concentration of about 0.01 to about 40 mM.
17. The composition of claim 10 , wherein the dextran has a molecular mass of about 300,000 to about 600,000 Da.
18. The composition of claim 17 , wherein the polydispersity of the molecular mass of the dextran is 1 to about 3.
19. The composition of claim 18 , wherein the polydispersity of the molecular mass of the dextran is about 1.1 to about 2.4.
20. The composition of claim 10 , wherein the dextran is present at a concentration of about 0.01 to about 10 mM.
21. The composition of claim 1 , wherein the bioactive hydrogel matrix that is flowable at physiological temperatures further comprises at least one polar amino acid selected from the group consisting of tyrosine, cysteine, serine, threonine, asparagine, glutamine, aspartic acid, glutamic acid, arginine, lysine, histidine, and mixtures thereof.
22. The composition of claim 21 , wherein the polar amino acids are present at a concentration of about 3 to about 150 mM.
23. The composition of claim 21 , wherein the polar amino acids are present at a concentration of about 10 to about 65 mM.
24. The composition of claim 21 , wherein the polar amino acids are selected from the group consisting of L-cysteine, L-glutamic acid, L-lysine, L-arginine, and mixtures thereof.
25. The composition of claim 21 , wherein the bioactive hydrogel matrix comprises L-glutamic acid at a concentration of about 2 to about 60 mM.
26. The composition of claim 21 , wherein the bioactive hydrogel matrix comprises L-lysine at a concentration of about 0.5 to about 30 mM.
27. The composition of claim 21 , wherein the bioactive hydrogel matrix comprises L-arginine at a concentration of about 1 to about 40 mM.
28. The composition of claim 21 , wherein the bioactive hydrogel matrix comprises L-cysteine at a concentration of about 0.005 to about 5 mM.
29. The composition of claim 1 , wherein the bioactive hydrogel matrix that is flowable at physiological temperatures further comprises ethylenediaminetetraacetic acid or a salt thereof.
30. The composition of claim 29 , wherein the ethylenediaminetetraacetic acid or salt thereof is present at a concentration of about 0.01 to about 10 mM.
31. The composition of claim 1 , further comprising at least one osteoinductive or osteoconductive material.
32. The composition of claim 31 , wherein the at least one osteoinductive or osteoconductive material is selected from the group consisting of demineralized bone matrix (DBM), bone morphogenetic proteins (BMPs), transforming growth factors (TGFs), fibroblast growth factors (FGFs), insulin-like growth factors (IGFs), platelet-derived growth factors (PDGFs), epidermal growth factors (EGFs), vascular endothelial growth factors (VEGFs), and vascular permeability factors (VPFs).
33. The composition of claim 1 , wherein the crosslinked hydrogel comprises a polypeptide crosslinked to a long chain carbohydrate and wherein the long chain carbohydrate is a polysaccharide or a sulfated polysaccharide.
34. The composition of claim 33 , wherein the polypeptide of the crosslinked hydrogel is a tissue-derived polypeptide derived from tissue selected from the group consisting of collagens, gelatins, keratin, decorin, aggrecan, and glycoproteins.
35. The composition of claim 33 , wherein the polysaccharide or sulfated polysaccharide is selected from the group consisting of dextran, heparan, heparin, hyaluronic acid, alginate, agarose, carageenan, amylopectin, amylose, glycogen, starch, cellulose, chitin, chitosan, heparan sulfate, chondroitin sulfate, dextran sulfate, dermatan sulfate, and keratan sulfate.
36. The composition of claim 33 , wherein the polypeptide is gelatin and the polysaccharide is dextran or oxidized dextran.
37. The composition of claim 1 , wherein the crosslinked hydrogel is at least partially dehydrated.