Source: https://patents.google.com/patent/EP1374857A1/en
Timestamp: 2019-04-24 09:27:21
Document Index: 790513701

Matched Legal Cases: ['art-1', 'art-1', 'art, 5', 'art, 5', 'art, 5', 'art, 5', 'art, 5', 'art, 5', 'art, 5']

EP1374857A1 - Collagen-polysaccharide matrix for bone and cartilage repair - Google Patents
EP1374857A1
EP1374857A1 EP03078133A EP03078133A EP1374857A1 EP 1374857 A1 EP1374857 A1 EP 1374857A1 EP 03078133 A EP03078133 A EP 03078133A EP 03078133 A EP03078133 A EP 03078133A EP 1374857 A1 EP1374857 A1 EP 1374857A1
EP03078133A
1997-01-15 Priority to US783650 priority Critical
1997-01-15 Priority to US08/783,650 priority patent/US5866165A/en
1998-01-15 Application filed by DePuy Spine LLC filed Critical DePuy Spine LLC
1998-01-15 Priority to EP98902579A priority patent/EP0994694B1/en
2004-01-02 Publication of EP1374857A1 publication Critical patent/EP1374857A1/en
An injectable gel to support the repair of tissue, said gel comprising collagen covalently crosslinked to an exogenous polysaccharide, wherein said polysaccharide is crosslinked to said collagen through oxidized sugar rings on said polysaccharide which form covalent linkages to said collagen.
Biological glue comprising fibrin has a long history as a tissue adhesive medical device and is believed to be commercially available in Europe (United States patent No. 5,260,420, issued November 9, 1993). One obstacle that limits its application is the short turn over and residence time which ranges from a few days to a few weeks depending on the site of implantation. The incorporation of collagen fibers into fibrin glue has been reported (Sierra et al., 1993, Trans. Soc. Biomater., vol. 16:257 and United States Patent No. 5,290,552). However, longer coagulation times are required for the collagen/fibrin compositions compared to fibrin alone.
The type of polysaccharides which can be used include hyaluronic acid, chondroitin sulfate, dermatan 5 sulfate, keratan sulfate, heparan, heparan sulfate, dextran, dextran sulfate, alginate, and other long chain polysaccharides. In a preferred embodiment, the polysaccharide is hyaluronic acid.
The preferred polysaccharide is hyaluronic acid. The relative proportion of polysaccharide to collagen will impart various physical and biological 5 characteristics to the matrix. The proportion of polysaccharide to collagen may be characterized on a molar ratio basis or on a weight ratio basis. Typically, the ratio by weight of collagen to polysaccharide is from 99:1 to about 1:99. This represents an approximate molar ratio of 99.9:0.1 to 1:9, respectively, assuming an average molecular weight of 1,000,000 daltons for hyaluronic acid and 100,000 daltons for collagen. The molar ratio may vary depending on the actual molecular weight of the polysaccharide and collagen used. In a 5 preferred embodiment disclosed herein, the ratio by weight of collagen to polysaccharide is from 9:1 to about 1:9.
Alternatively, increasing the proportion of polysaccharide, preferably hyaluronic acid, will more closely mimic a natural cartilage matrix. In addition, a higher proportion of polysaccharide will mask some specific cell adhesive sites on collagen and will favor other cell-cell interactions and aggregation important in the development of cartilage tissue.
Growth factors which can be used with a matrix of the present invention include, but are not limited to, members of the TGF-β superfamily, including TGF-β1,2 and 3, the bone morphogenetic proteins (BMP's), the growth differentiation factors(GDF's), and ADMP-1; members of the fibroblast growth factor family, including acidic and basic fibroblast growth factor (FGF-1 and -2); members of the hedgehog family of proteins, including indian, sonic and desert hedgehog; members of the insulin- like growth factor (IGF) family, including IGF-I and -II; members of ) the platelet-derived growth factor (PDGF) family, including PDGF-AB, PDGF-BB and PDGF-AA; members of the interleukin (IL) family, including IL-1 thru -6; and members of the colony-stimulating factor (CSF) family, including CSF-1, G-CSF, and GM-CSF. Growth factor preparations are obtained either commercially or isolated and purified from tissue or from recombinant sources. Growth factors can be loaded into the collagen/HA/fibrin matrices across a wide dose range (fentogram to millgram range). Factors such as cost, safety and the desired growth factor release profile will dictate the amount of growth factor that is loaded onto the matrix.
The concentration of fibrinogen used in forming the matrix is preferably 10 mg/ml or greater. The thrombin is added to the fibrinogen in a concentration of from about 0.01 NIH units to about 100 NIH units/ml and preferably from about 0.1 - 2.0 NIH units/ml. The thrombin is commercially available from a variety of sources including from Calbiochem-Novabiochem, San Diego, CA. Fibrinogen may be derived from autologous patient plasma or from commercial sources, such as Calbiochem-Novabiochem,San Diego, CA.
Retinoic acid-treated chondrocytes represent the latter stages of chondrogenesis. Retinoic acid treatment of primary is performed prior to culturing or seeding the cells on a candidate matrix (Dietz, U. et al., 1993, J. Cell Biol. 52(1):57-68).
The analysis of differentiation markers relevant to chondrogenesis and osteogenesis are evaluated at both the protein and transcriptional level. The specific markers that may be analyzed include: 1) Type II collagen and IIA, IIB isoforms; 2) Aggrecan proteoglycan; 3) Type IX, X and XI collagen; 4) Type I collagen; 5) Cartilage matrix protein (CMP); 6) Cart-1 transcription factor; 7) Fibronectin (EDA, EDB isoforms) ; 8) Decorin proteoglycan; 9) Link protein; 10) NG-2 proteoglycan; 11) Biglycan proteoglycan; 12) Alkaline phosphatase. Differentiation may be measured by Northern/PCR analysis, Western blotting or by metabolic cell labeling.
For Western blotting, solubilized protein lysates are isolated from cells cultured on composite matrices by standard techniques (Spiro R.C., et al., 1991, J. Cell. Biol., 115:1463-1473). After the lysis of cells the matrices are extracted in stronger denaturants (8 M urea, GnHCL) to remove and examine matrix-bound or incorporated proteins. Protein samples are analyzed by standard Western blotting techniques using specific polyclonal or monoclonal antibodies.
Affinity coelectrophoresis is used to analyze proteoglycan binding to a matrix of the present invention. 35SO4-labeled or iodinated proteoglycan (aggrecan) isolated from bovine and rat (or other sources) is loaded into ACE gels (Lee, M.K. et al., 1991, 88:2768-2772) containing composite matrices or collagen scaffolds alone. The binding affinity of aggrecan for collagen scaffolds plus and minus hyaluronic acid or dextran sulfate are taken as a measure of the ability of composite matrices to organize a cartilage matrix.
Alternatively, the shift in expression from Type I to Type II collagen and the splicing of the Type II collagen transcript from the Type IIA to the Type IIB isoform (Sandell, L.J. et al., 1991, J. Cell Biol. 114:1307-1319) are measured by means known to those of skill in the art to determine differentiation down a chondrogenic pathway. Also, the expression of the cartilage-associated proteoglycan, aggrecan (Schmid, T.M., et al., 1985, J. Cell Biol. 100:598-605 and Kuettner K.E. 1992, Clin. Biochem. 25:155-163) and a cartilage homeoprotein transcription factor (Cart-1) appear to be markers for cells committed to the chrondrocytic lineage.
The matrices of the present invention may be used for the treatment of bone and/or cartilage defects 5 associated with surgical resection, such as spinal fusions; trauma; disease; infection; cancer or genetic defects. The matrices according to the present invention may be administered through implantation, direct application or injection depending on the intended application of the matrix, the physical properties of the matrix and the ratio by weight of collagen to polysaccharide in the matrix.
The matrix in this case was based bn the reaction of protein amine residues on the collagen with the active aldehyde groups generated on the sugar rings of the polysaccharides. Matrices with various surface properties and biological activity are synthesized by controlling the ratios of the collagen to the polysaccharides, the type of collagen, the types of polysaccharides, as well as the density of the aldehyde groups generated on the polysaccharides.
Semed F collagen (8.1 parts) and Semed S collagen (0.9 part) were dispersed in a hyaluronate/polyaldehyde solution (1 part, 5% of the repeat units were oxidized: pH 3-3.5) containing lOmM sodium cyanoborohydride (NaCNBH3) in a heavy duty blender at low speed for 10 seconds followed by high speed for another 5 seconds. The slurry (solids concentration: 28 mg/ml) was poured into a mold, incubated at ambient temperature for 24 hours and lyophilized. This formed a sponge which was washed several times in distilled water to completely remove the NaCNBH3. The washed sponge was then lyophilized.
Semed F collagen (0.9 part) Hyaluronate/polyaldehyde
Semed S collagen (0.1 part) solution (1 part, 5% of the repeat units oxidized) Solids concentration: 15mg/ml
Semed S collagen (0.1 part) solution (2 parts, 5% of the repeat units oxidized) Solids concentration: 15mg/ml
Semed S collagen (0.1 part) solution (4 parts, 5% of the repeat units oxidized) Solids concentration: 15mg/ml
Semed S collagen (0.1 part) solution (4 parts, 1% of the repeat units oxidized) Solids concentration: 15mg/ml
Collagen Type II (9 parts) Hyaluronate/polyaldehyde
solution (1 part, 5% of the repeat units oxidized) Solids concentration: 15mg/ml
Collagen Type II (1 part) Hyaluronate/polyaldehyde
Semed F collagen (7 parts) Hyaluronate/polyaldehyde
Collagen Type II (2 parts) solution (1 part, 5% of the repeat units oxidized) Solids concentration: 15mg/ml
Semed F collagen (8.1 parts) Dextran/polyaldehyde
Semed S collagen (0.9 part) solution (1 part, 5% of the repeat units oxidized) Solids concentration: 28mg/ml
Semed F collagen (8.1 parts) Dextran sulfate/polyaldehyde
Semed S collagen (0.9 part) (1 part, 5% of the repeat units oxidized) Solids concentration: 28mg/ml
Semed F collagen (8.1 parts) Chondroitin
Semed S collagen (0.9 part) sulfate/polyaldehyde (1 part, 5% of the repeat units oxidized) Solids concentration: 28mg/ml
Materials and Methods Preparation of Matrix with and without BMP
A crosslinked collagen-hyaluronate 9:1 matrix was prepared as described in Example 1 and lyophilized. For preparation of a matrix with growth factor, BMP (obtained from Intermedic Orthopedics, Denver, CO.) was dissolved and added to the lyophilized matrix to final concentration of 0.1% (50µg per 50µl of matrix). The matrix/growth factor combination was then lyophilized a second time prior to implantation into the cranial defect.
Osteoconduction in Rat Cranial Defects.
To evaluate the ability of a collagen-hyaluronic acid matrix (CN/HA scaffolds) to support the ingrowth of bony tissue, CN/HA scaffolds were implanted in defects created in the parietal bones of 6 week-old male Sprague Dawley rats by a modification of a previously described method (Mulliken, J.B. et al, 1980, reconstru. Surg. 65:553-559). Briefly, bilateral, rectangular defects of approximately 5 mm by 3 mm were made using a low speed Dremel drill fitted with an engraving bit under constant irrigation during drilling. Left defects were filled with one pre-cut dry piece of CN/HA scaffold, and right defects remained unfilled and served as untreated controls. Animals were sacrificed after 7, 14, 21 and 28 days, and calvaria were excised and fixed in 10% neutral buffered formalin.
To evaluate the ability of CN/HA scaffolds to deliver osteoinductive growth factors, implants of CN/HA containing bone morphogenetic protein (BMP) were placed in bilateral pouches created in the tibial muscles of 4 week-old male Sprague-Dawley rats. Each CN/HA matrix was loaded with 50 µg of BMP by absorption and subsequent lyophilization, and the contralateral limb received implants of CN/HA without BMP. Animals were sacrificed after 21 days, and implant materials were excised from surrounding tissue. Explanted tissues were cut into two pieces: one piece extracted and assayed for alkaline phosphatase (Lowry et al, 1954, J. Biol. Chem. 207:19 and Sampath, et al, 1981, Proc. Natl. Acad. Sci. 78:7599-7603) and the other piece fixed in 10% neutral buffered formalin, decalcified and processed for histological evaluation. Paraffin sections of 6 µm thickness were stained with hematoxylin and eosin and examined for ectopic bone formation, inflammation, and residual implant appearance. Alkaline phosphatase activity was expressed in units where 1 unit equals nmoles p-nitrophenol produced (from p-nitrophenylphosphate substrate) per minute at 37°C.
Calvarial and intramuscular implant specimens were decalcified in Formical (American Histology, Lodi, CA). Transverse gross cuts were made through the approximate center of the cranial defects prior to processing. Six µm paraffin sections at the center of the defects or explanted tissues were stained with hematoxylin and eosin. Tissues were evaluated for biocompatibility (inflammation), residual implant persistence and new bone formation. Bony healing of cranial defects were scored subjectively on a linear scale of 1 to 5, based on the width of the defect bridged with reparative bone.
Results Osteoconduction of CN/HA Matrices
Summary of Radiographic and Histological Evaluation
Radiographic Score Histological Bone Score
Time Point (% reduction in
defect area) CN/HA
(mean ± SD) Untreated
1 week 20 ± 12% 2.0 ± 1.5 1.0
2 weeks 43 ± 36% 4.0 ± 1.3 1.5 ± 0.8
3 weeks 69 ± 12% 4.0 ± 1.5 1.2 ± 0.4
4 weeks 86 ± 12% 4.8 ± 0.4 1.5 ± 0.5
In contrast, CN/HA scaffolds loaded with 50 µg BMP induced extensive ectopic bone formation after implantation in rat tibial muscle pouches for 21 days. Islands of new bone had formed throughout the CN/HA matrix within an ingrowth of fibrovascular tissue. The osseous tissue in the central region of the implant appeared less mature than the woven bone present at the periphery of the implant. The interconnecting woven bone spicules contained reversal lines, osteocytes in lacunae and many areas of presumptive hematopoietic marrow. A few scattered chondrocytes were also present within the new bone. Only traces of residual matrix material could be discerned within the bony ossicle. A thickened fibrous tissue capsule containing some chronic inflammatory cells was present peripherally. The specific alkaline phosphatase activity for 21 days explants of CN/HA scaffolds containing 50 µg BMP was 22 ± 6 units, while alkaline phosphatase was not detected (< 1 unit) in explants of CN/HA only.
CN/HA matrices implanted intramuscularly in the rat were well-tolerated and promoted the growth of fibrovascular tissue after 21 days. When CN/HA matrices contained 50 µg of BMP, extensive ectopic bone formation was induced. Although new woven bone formation was evident throughout the entire implant, the ossicle appeared to be more mature peripherally. Similar to CN/HA matrix alone implanted in cranial defects, virtually all of the matrix plus BMP had been resorbed or incorporated into the new bone. These results indicate that CN/HA matrices are suitable delivery vehicles for osteoinductive factors and support the cascade of events which occur as mesenchymal cells differentiate into bone. The efficiency of CN/HA matrices in delivery of osteoinductive factors, can be ascertained by an in vivo BMP dose-response study.
Matrices made by the process described in Example 1 comprising various ratios of collagen to polysaccharide are implanted into a full-thickness articular cartilage defect in the rabbit as described in Amiel et al., supra. Defects measuring approximately 3.7 mm in diameter and 5mm deep defect are created in the center of the medial femoral condyles of adult male New Zealand white rabbits. The defects are then either filled with matrix or left unfilled as controls. The defects are evaluated morphologically and histologically at 6 and 12 weeks.
The effect of collagen/hyaluronate gel on the coagulation time of fibrinogen was evaluated as follows: 0.5 ml of purified fibrinogen solution (30 mg/ml, PBS) was placed into a 10 ml glass tube and incubated in a water bath at 37o a 10 ml glass tube and incubated in a wafer bath at 37°C for 5 min. 0.1 ml of CaCl2 solution (0.2 M) containing 0.5 NIH U of thrombin and various concentrations of collagen and hyaluronate-polyaldehyde were added and mixed with a stir bar. The time required for the formation of gel on top of the stir bar was recorded as the coagulation time. The results are shown in Table 1.
Fibrin coagulation time in the presence of collagen and hyaluronate.
Sample Collagen
(mg/ml) HA-pALD
(mg/ml) Coagulation
sample 1 0 0 44.3 +/- 2.5
sample 2 9.0 0 55.6 +/- 1.2
sample 3 4.5 10 26 +/- 2
sample 4 3.6 8 26
* All tests were done in triplicate. Mean ± SD shown.
This experiment was performed with a three-way stopcock equipped with male luer slip adapter, to which two syringes were connected. Syringe A contained 5 ml of fibrinogen and hyaluronate-polyaldehyde (20 mg/ml for each, in DMEM medium), syringe B contained thrombin (2 NIH U/ml, in DMEM medium) and collagen Semed S (Kensey Nash) suspension (20 mg/ml, in DMEM medium), which had been pre-blended to a fiber diameter of about 50 mm in a heavy duty blender. The contents of syringes were admixed quickly between the two syringes and the contents were drawn into one syringe which was then angled at 60° into a phosphate buffered saline (PBS) solution (40 ml in 100 ml glass beaker) and the contents were discharged into the PBS solution. The entire discharge including mixing was completed in 10 sec. No dissolution of the gel thus formed was observed after incubation at ambient temperature or at 37°C.
This example illustrates the three-dimensional scaffold formation of a crosslinked collagen-hyaluronate matrix comprising fibrinogen and thrombin in deionized water .
The experiment was performed with a three-way stopcock equipped with male luer slip adapter, to which two syringes were connected. Syringe A contained 5 ml of fibrinogen and hyaluronate-polyaldehyde (20 mg/ml for each, 12 mM NaOH), syringe B contained thrombin (2 NIH U/ml) and purified collagen solution (Collagen corporation, 3 mg/ml, 12 mM HCl). The contents of syringes were admixed quickly between the two syringes and the contents were drawn into one syringe which was then angled at 60° into a phosphate buffered saline (PBS) solution (40 ml in 100 ml glass beaker) and the contents were discharged into the PBS solution. The entire discharge was completed in 10 sec. No dissolution of the gel thus formed was observed after incubation at ambient temperature or at 37°C.
Gels with different-compositions as described in Table 2 were prepared using a three-way stopcock equipped with male luer slip adapter, and two syringes. After mixing, the mixtures were cast in 24-well cell culture plate at the rate of 0.5 ml/well and allowed to stand at ambient temperature for 15 min. The gels thus formed were lyophilized, chopped to cubes 10 x 10 x 5 mm long, and sterilized with ethanol. These specimens were then placed in pouches made by blunt dissection in the tibial muscle of 4 to 5 week old male Sprague Dawley rats. The rats were sacrificed after 3, 7, and 14 days, and explants were evaluated histologically for biocompatibility and implant persistence.
Compositions of matrices for in vivo test
Sample Fibrinogen
(mg/ml) Collagen
(mg/ml) Total Mass
(mg/ml) Thrombin
(NIH U/ml)
FN 40 0 0 40 2
FN/HA-pALD 36 0 4 40 2
CN/HA-pALD 0 36 4 40 2
FN/CN 20 20 0 40 2
FN/CN/
HA-pALD 18 18 4 40 2
CN 0 40 0 40 2
The disclosure of this application also incudes the following clauses numbered 1 through 27.
Clause 1. A method for preparing a matrix to support the repair of tissue comprising the steps of oxidizing an exogenous polysaccharide to form a modified exogenous polysaccharide having aldehyde groups, and reacting said modified exogenous polysaccharide with collagen under conditions whereby said aldehyde groups covalently react to crosslink with collagen to form said matrix.
Clause 2. The method of clause 1 further comprising adding a growth factor to said matrix.
Clause 3. The method of clause 2 wherein said growth factor is selected from the group consisting of members of the TGF-β superfamily; members of the BMP family; the growth differentiation factors(GDF's); ADMP-1; members of the fibroblast growth factor family; members of the hedgehog family of proteins; members of the insulin- like growth factor (IGF) family; members of the platelet-derived growth factor (PDGF) family; members of the interleukin (IL) family; and members of the colony-stimulating factor (CSF) family
Clause 4. The method of clause 3 wherein the growth factor is a bone morphogenic protein (BMP).
Clause 5. The method of clause 1 wherein the polysaccharide comprises hyaluronic acid, chondroitin sulfate, dermatan sulfate, keratan sulfate, heparan, heparan sulfate, dextran, dextran sulfate, or alginate.
Clause 6. The method according to clause 5, wherein said polysaccharide comprises hyaluronic acid.
Clause 7. The method according to clause 1 wherein the collagen is selected from the group consisting of Type 1 and Type II collagen.
Clause 8 . The method according to clause 1, wherein said step of oxidizing said polysaccharide comprises treatment of said polysaccharide with periodate.
Clause 9. The method according to clause 1, wherein said collagen and said polysaccharide used to form said matrix are present in the range of 99:1 to 1:99 by weight, respectively.
Clause 10. The method according to clause 9 wherein said range is 9:1 to 1:9 by weight, respectively.
Clause 11. The method according to clause 1, wherein about 1% to 50% of the repeat units in said polysaccharide are oxidized to contain aldehyde groups.
Clause 12. The method according to clause 11, wherein about 1% to 5% of the repeat units in said polysaccharide are oxidized to contain aldehyde groups.
Clause 13. The method according to clause 1, wherein said matrix is formed by freezing and lyophilization.
Clause 14. The method according to clause 1, wherein said matrix is formed by wet laying and air drying.
Clause 15. The method of clause 1 further comprising adding fibrinogen and thrombin to form fibrin in said matrix.
Clause 16. The method of clause 1 wherein tissue is selected from the group consisting of bone, cartilage and soft tissue.
Clause 17. A matrix to support the repair of tissue, said matrix comprising collagen covalently crosslinked to an exogenous polysaccharide, wherein said polysaccharide is crosslinked to said collagen through oxidized sugar rings on said polysaccharide which form covalent linkages to said collagen.
Clause 18. The matrix of clause 15 further comprising a growth factor.
Clause 19. The matrix of clause 18 wherein said growth factor is selected from the group consisting of members of the TGF-β superfamily; members of the bone morphogenic protein family; the growth differentiation factors(GDF's); ADMP-1; members of the fibroblast growth factor family; members of the hedgehog family of proteins; members of the insulin-like growth factor (IGF) family; members of the platelet-derived growth factor (PDGF) family; members of the interleukin (IL) family; and members of the colony-stimulating factor (CSF) family.
Clause 20. The matrix of clause 18 wherein said growth factor is a bone morphogenic protein.
Clause 21. The matrix according to clause 17 wherein said polysaccharide comprises hyaluronic acid, chondroitin sulfate, dermatan sulfate, keratan sulfate, heparan, heparan sulfate, dextran, dextran sulfate or alginate.
Clause 22. The matrix according to clause 21 wherein said polysaccharide is hyaluronic acid.
Clause 23. The matrix according to clause 17 wherein said matrix comprises said collagen and said polysaccharide in a weight ratio in the range of 99:1 to 1:99.
Clause 24. The matrix of clause 17 wherein said collagen is selected from the group consisting of Type 1 collagen and Type 2 collagen..
Clause 25. The matrix of clause 17 further comprising fibrin.
Clause 26. A method of conducting the growth of bone or cartilage tissue in vivo comprising the step of administering a matrix according to clause 17 at a site of desired bone growth.
Clause 27. A method of inducing the growth of bone or cartilage tissue in vivo comprising the step of administering a matrix according to clause 18 at a site of desired bone growth.
The gel of claim 1 further comprising a growth factor.
The gel of claim 2 wherein said growth factor is selected from the group consisting of members of the TGF-β superfamily; members of the bone morphonegic protein family; the growth differentiation factors (GDF's); ADMP-1; members of the fibroblast growth factor family; members of the hedgehog family of proteins; members of the insulin-like growth (IGF) family; members of the platelet-derived growth factor (PDGF) family; members of the interleukin (IL) family; and members of the colony-stimulating factor (CSF) family.
The gel according to claim 1 wherein said polysaccharide comprises hyaluronic acid, chondroitin sulfate, dermatan sulfate, keratan sulfate, heparan, heparan sulfate, dextran, dextran sulfate or alginate.
The gel according to claim 4 wherein said polysaccharide is hyaluronic acid.
The gel of claim 1 wherein said collagen is selected from the group consisting of Type I collagen and Type II collagen.
The gel of claim 1 further comprising fibrin.
A method of manufacturing a gel for conducting the growth of bone or cartilage tissue in vivo comprising the step of forming a gel according to claim 1.
A method of manufacturing a gel for inducing the growth of bone or cartilage tissue in vivo comprising the step of forming a gel according to claim 2.
EP03078133A 1997-01-15 1998-01-15 Collagen-polysaccharide matrix for bone and cartilage repair Withdrawn EP1374857A1 (en)
US783650 1997-01-15
US08/783,650 US5866165A (en) 1997-01-15 1997-01-15 Collagen-polysaccharide matrix for bone and cartilage repair
EP98902579A EP0994694B1 (en) 1997-01-15 1998-01-15 Collagen-polysaccharide matrix for bone and cartilage repair
EP98902579A Division EP0994694B1 (en) 1997-01-15 1998-01-15 Collagen-polysaccharide matrix for bone and cartilage repair
EP1374857A1 true EP1374857A1 (en) 2004-01-02
EP03078133A Withdrawn EP1374857A1 (en) 1997-01-15 1998-01-15 Collagen-polysaccharide matrix for bone and cartilage repair
EP98902579A Expired - Lifetime EP0994694B1 (en) 1997-01-15 1998-01-15 Collagen-polysaccharide matrix for bone and cartilage repair
AT (1) AT252886T (en)
AU (1) AU727430B2 (en)
NZ (1) NZ336480A (en)
EP1912661A1 (en) * 2005-07-20 2008-04-23 Sewon Cellontech Co., Ltd. Simple method of transplanting injectable chondrocyte for autologous chondrocyte transplantation
KR101955549B1 (en) * 2010-11-23 2019-03-07 엘라스타겐 피티와이 리미티드 Preparation and/or formulation of proteins cross-linked with polysaccharides
SG11201509238WA (en) * 2013-05-09 2015-12-30 Agency Science Tech & Res Modified collagen molecules
CN103342824B (en) * 2013-06-28 2015-07-01 华南理工大学 Application method of cyclodextrin-aldehyde cross-linking agent
KR101863532B1 (en) * 2017-06-15 2018-06-01 세원셀론텍(주) Manufacturing and use method of cartilage tissue repair composition
1997-01-15 US US08/783,650 patent/US5866165A/en not_active Expired - Lifetime
1998-01-15 WO PCT/US1998/000838 patent/WO1998031345A1/en active IP Right Grant
1998-01-15 DE DE69819329T patent/DE69819329T2/en not_active Expired - Lifetime
1998-01-15 AU AU59203/98A patent/AU727430B2/en not_active Ceased
1998-01-15 AT AT98902579T patent/AT252886T/en unknown
1998-01-15 DK DK98902579T patent/DK0994694T3/en active
1998-01-15 CA CA002277110A patent/CA2277110C/en not_active Expired - Fee Related
1998-01-15 JP JP53455198A patent/JP3348861B2/en not_active Expired - Fee Related
1998-01-15 ES ES98902579T patent/ES2209107T3/en not_active Expired - Lifetime
1998-01-15 US US09/007,731 patent/US5972385A/en not_active Expired - Lifetime
1998-01-15 EP EP03078133A patent/EP1374857A1/en not_active Withdrawn
1998-01-15 EP EP98902579A patent/EP0994694B1/en not_active Expired - Lifetime
1998-01-15 NZ NZ336480A patent/NZ336480A/en not_active IP Right Cessation
EP1912661A4 (en) * 2005-07-20 2009-09-09 Sewon Cellontech Co Ltd Simple method of transplanting injectable chondrocyte for autologous chondrocyte transplantation
US5866165A (en) 1999-02-02
CA2277110A1 (en) 1998-07-23
DE69819329D1 (en) 2003-12-04
AU727430B2 (en) 2000-12-14
ES2209107T3 (en) 2004-06-16
AT252886T (en) 2003-11-15
JP3348861B2 (en) 2002-11-20
WO1998031345A1 (en) 1998-07-23
DK0994694T3 (en) 2004-02-16
NZ336480A (en) 2001-03-30
JP2000514698A (en) 2000-11-07
CA2277110C (en) 2003-04-22
EP0994694B1 (en) 2003-10-29
EP0994694A4 (en) 2002-07-10
US5972385A (en) 1999-10-26
EP0994694A1 (en) 2000-04-26
DK994694T3 (en)
DE69819329T2 (en) 2004-05-13
AU5920398A (en) 1998-08-07
AU662024B2 (en) 1995-08-17 The use of biomaterials for tissue repair
CA1259914A (en) 1989-09-26 Methods of bone repair using collagen
US20050158358A1 (en) 2005-07-21 Tissue engineering scaffolds promoting matrix protein production
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