Claims for Patent: 6,617,161

Summary for Patent: 6,617,161

Title:	Serum-free cell growth medium
Abstract:	A chemically defined-serum free growth medium for the in vitro and ex vivo of cells and cell lines. The medium consists of about a one to one ratio (v/v) of two basal growth media containing .alpha.-ketoglutarate, insulin, transferrin, selenium, bovine serum albumin, linoleic acid, ceruloplasmin, cholesterol, phosphatidyl-ethanolamine, .alpha.-tocopherol acid succinate, reduced glutathione, taurine, triiodothyronine, hydrocortisone, parathyroid hormone, L-ascorbic acid 2-sulfate, .beta.-glycerophosphate, PDGF, EGF and FGF. Chondrocytes, when cultured in this medium in the presence of a cartilage derived morphogenetic protein or bone morphogenetic protein, retain their cartilaginous phenotype.
Inventor(s):	Luyten; Frank P. (Kraainem, BE), Erlacher; Ludwig (Vienna, AT)
Assignee:	The United States of America as represented by the Department of Health and Human Services (Washington, DC)
Application Number:	09/851,921
Patent Claims:	1. A method of maintaining a cartilaginous phenotype in chondrocytes in vitro comprising contacting a culture comprising chondrocytes, which have not lost and subsequently re-expressed a cartilaginous phenotype, with a serum-free cell growth medium comprising about a 1:1 ratio (v/v) of two basal cell culture media, said medium containing effective cell growth-promoting concentrations of .alpha.-ketoglutarate, insulin, transferrin, selenium, bovine serum albumin, linoleic acid, ceruloplasmin, cholesterol, phosphatidylethanolamine, .alpha.-tocopherol acid succinate, reduced glutathione, taurine, triiodothyronine, hydrocortisone, parathyroid hormone, L-ascorbic acid 2-sulfate, .beta.-glycerophosphate, platelet-derived growth factor (PDGF), epidermal growth factor (EGF) and basic fibroblast growth factor (bEGF) and at least one morphogenetic protein selected from the group consisting of cartilage-derived morphogenetic proteins and bone morphogenetic proteins, whereby a cartilaginous phenotype is maintained in said chondrocytes in vitro. 2. A method of repairing a joint surface defect in a mammal in need thereof comprising the steps of: removing normal cartilage in the vicinity of said surface defect; isolating chondrocytes from said cartilage; contacting a culture comprising said chondrocytes, which have not lost and subsequently re-expressed a cartilaginous phenotype, with a serum-free cell growth medium comprising about a 1:1 ratio (v/v) of two basal cell culture media, said medium containing effective cell growth-promoting concentrations of .alpha.-ketoglutarate, insulin, transferrin, selenium, bovine serum albumin, linoleic acid, ceruloplasmin, cholesterol, phosphatidylethanolamine, .alpha.-tocopherol acid succinate, reduced glutathione, taurine, triiodothyronine, hydrocortisone, parathyroid hormone, L-ascorbic acid 2-sulfate, .beta.-glycerophosphate, platelet-derived growth factor (PDGF), epidermal growth factor (EGF) and basic fibroblast growth factor (bEGF) and at least one morphogenetic protein selected from the group consisting of cartilage-derived morphogenetic proteins and bone morphogenetic proteins, whereby a cartilaginous phenotype is maintained in said chondrocytes; and implanting said chondrocytes into said surface defect. 3. The method of claim 1 or 2, wherein two basal cell culture media are selected and mixed in equal proportion and wherein said basal cell culture media are selected from the group consisting of Ham's F-12, Dulbecco's modified Eagle's medium (DMEM), Essential modified Eagle's medium (EMEM) and RPMI-1640. 4. The method of claim 1 or 2, wherein said morphogenetic protein is cartilage-derived morphogenetic protein and is selected from the group consisting of CDMP-1 and CDMP-2. 5. The method of claim 1 or 2, wherein said morphogenetic protein is bone morphogenetic protein and is selected from the group consisting of OP-1, BMP-2, BMP-3, BMP-4, BMP-5 and BMP-6. 6. The method of claim 1 or 2, wherein the concentration of .alpha.-ketoglutarate is about 1.times.10.sup.-4 M. 7. The method of claim 1 or 2, wherein the concentration of ceruloplasmin is about 0.25 U/ml. 8. The method of claim 1 or 2, wherein the concentration of cholesterol is about 5 .mu.g/ml. 9. The method of claim 1 or 2, wherein the concentration of phosphatidyl-ethanolamine is about 2 .mu.g/ml. 10. The method of claim 1 or 2, wherein the concentration of .alpha.-tocopherol acid succinate is about 9.times.10.sup.-7 M. 11. The method of claim 1 or 2, wherein the concentration of reduced glutathione is about 10 .mu.g/ml. 12. The method of claim 1 or 2, wherein the concentration of taurine is about 1.25 .mu.g/ml. 13. The method of claim 1 or 2, wherein the concentration of triiodothyronine is about 1.6.times.10.sup.-9 M. 14. The method of claim 1 or 2, wherein the concentration of hydrocortisone is about 1.times.10.sup.-9 M. 15. The method of claim 1 or 2, wherein the concentration of parathyroid hormone is about 5.times.10.sup.-10 M. 16. The method of claim 1 or 2, wherein the concentration of L-ascorbic acid 2-sulfate is about 50 .mu.g/ml. 17. The method of claim 1 or 2, wherein the concentration of PDGF is about 4 ng/ml. 18. The method of claim 1 or 2, wherein the concentration of EGF is about 10 ng/ml. 19. The method of claim 1 or 2, wherein the concentration of bFGF is about 10 ng/ml. 20. The method of claim 2, wherein said isolating chondrocytes step comprises digestion of said cartilage with collagenase.

Details for Patent 6,617,161

Applicant	Tradename	Biologic Ingredient	Dosage Form	BLA	Approval Date	Patent No.	Expiredate
Smith & Nephew, Inc.	SANTYL	collagenase	Ointment	101995	06/04/1965		2019-12-21
Nps Pharmaceuticals, Inc.	NATPARA	parathyroid hormone	For Injection	125511	01/23/2015		2019-12-21