[특허]PDGF fusion proteins incorporated into fibrin foams

Kuros Biosurgery AG

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PDGF fusion proteins incorporated into fibrin foams

특허상세정보
국가/구분	United States(US) Patent 등록
국제특허분류(IPC7판)	A61K-038/48
미국특허분류(USC)	424/094.64; 424/094; 530/350
출원번호	US-0342420 (2008-12-23)
등록번호	US-8226942 (2012-07-24)
발명자 / 주소	Charier, Gaëlle Müller-Maissen, Manuela Jen, Anna
출원인 / 주소	Kuros Biosurgery AG
대리인 / 주소	Pabst Patent Group LLP
인용정보	피인용 횟수 : 7 인용 특허 : 52

초록

Compositions for wound healing, use of the compositions, and kits and methods of using thereof are described herein. In a preferred aspect, the compositions are suitable for use in a method for forming a fibrin matrix or foam that can be applied or injected at the site of need. In another preferred aspect, the compositions are also suitable for use in methods for forming enhanced controlled delivery fibrin matrices that can be administered as gels or foams.

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1. A foam composition comprising: fibrinogen;thrombin, wherein the amount of thrombin is less than 0.3 IU of thrombin/mg of fibrinogen;at least one fusion protein comprising a first domain comprising a platelet-derived growth factor (PDGF) and a second domain comprising a transglutaminase substrate domain;and a biocompatible gas selected from the group consisting of CO2, N2, air, and an inert gas, in an effective amount to form the foam. 2. The composition of claim 1, further comprising a calcium source. 3. The composition of claim 1, wherein the transglutaminase substrate domain comprises a Factor XIIIa substrate domain. 4. The composition of claim 3, wherein the Factor XIIIa substrate domain comprises SEQ ID NO:1. 5. The composition of claim 1, wherein the fusion protein further comprises a degradation site between the first and the second domain. 6. The composition of claim 5, wherein the degradation site is an enzymatic or hydrolytic degradation site. 7. The composition of claim 5, wherein the degradation site is an enzymatic degradation site which is cleaved by an enzyme selected from the group consisting of plasmin and matrix metalloproteinase. 8. The composition of claim 1, wherein the fusion protein comprises the amino acid sequence of SEQ ID NO:2 or SEQ ID NO:3. 9. The composition of claim 1, wherein the concentration of the fibrinogen solution is in a range of about 10 mg/ml to about 130 mg/ml. 10. The composition of claim 1, wherein the concentration of the fibrinogen solution is about 50 mg/ml. 11. The composition of claim 1, wherein the thrombin amount is from about 0.04 to about 0.28 IU thrombin per mg of fibrinogen. 12. The composition of claim 1, wherein the thrombin amount is about 0.08 IU thrombin per mg of fibrinogen. 13. The composition of claim 1, wherein the amount of the fusion protein is from about 4 to about 12 μg fusion protein per mg of fibrinogen. 14. A kit for forming a foam comprising: (i) a first container comprising fibrinogen and at least one fusion protein comprising a first domain comprising a platelet-derived growth factor (PDGF) and a second domain comprising a substrate domain for a crosslinking enzyme; and(ii) a second container comprising thrombin, wherein the amount of thrombin is less than 0.3 IU thrombin per mg of fibrinogen, and(iii) a biocompatible gas selected from the group consisting of CO2, N2, air, and an inert gas. 15. The kit of claim 14, further comprising a calcium source. 16. The kit of claim 14, wherein the biocompatible gas is either in the first or the second container. 17. The kit of claim 14, wherein the fusion protein comprises the amino acid sequence of SEQ ID NO:2 or SEQ ID NO:3. 18. The kit of claim 14, wherein the concentration of the fibrinogen solution is in a range of about 10 mg/ml to about 130 mg/ml. 19. The kit of claim 14, wherein the thrombin amount is from about 0.04 to about 0.28 IU thrombin per mg of fibrinogen. 20. The kit of claim 14, wherein the amount of the fusion protein is from about 4 to about 12 μg fusion protein per mg of fibrinogen. 21. A method for preparing a fibrin foam comprising at least one fusion protein, the method comprising the steps of: (i) providing a fibrinogen solution;(ii) providing a thrombin solution wherein the amount of thrombin is less than 0.3 IU thrombin per mg of fibrinogen;(iii) providing at least one fusion protein comprising a first domain comprising a platelet-derived growth factor (PDGF) and a second domain comprising a transglutaminase substrate domain;(iv) providing a biocompatible gas selected from the group consisting of CO2, N2, air, and an inert gas; and(v) mixing components provided in steps (i), (ii), (iii), and (iv) to form the fibrin foam thereby covalently linking the fusion protein to fibrin through the second domain. 22. The method of claim 21, further comprising the step of providing a calcium source. 23. The method of claim 21, wherein the biocompatible gas is air. 24. The method of claim 21, wherein the volume of the fibrinogen solution is about 40 to 60% of the volume of the biocompatible gas. 25. The method of claim 21, wherein the transglutaminase substrate domain comprises a Factor XIIIa substrate domain. 26. The method of claim 21, wherein the Factor XIIIa substrate domain comprises SEQ ID NO:1. 27. The method of claim 21, wherein the fusion protein further comprises a degradation site between the first and the second domain. 28. The method of claim 27, wherein the degradation site is an enzymatic or hydrolytic degradation site. 29. The method of claim 28, wherein the degradation site is an enzymatic degradation site, which is cleavable by an enzyme selected from the group consisting of plasmin and matrix metalloproteinase. 30. The method of claim 21, wherein the fusion protein comprises SEQ ID NO:2. 31. The method of claim 21, wherein the concentration of the fibrinogen solution is in a range of about 10 mg/ml to 130 mg/ml. 32. The method of claim 21, wherein the thrombin amount is from about 0.04 to 0.28 IU thrombin per mg of fibrinogen. 33. The method of claim 21, wherein the fusion protein amount is in the range of from about 4 to 12 μg/mg of fibrinogen. 34. A fibrin foam obtained by mixing (i) a fibrinogen solution;(ii) a thrombin solution, wherein the amount of thrombin is less than 0.3 I.U. thrombin per mg of fibrinogen;(iii) at least one fusion protein comprising a first domain comprising a platelet-derived growth factor (PDGF) and a second domain comprising a transglutaminase substrate domain; and(iv) a biocompatible gas selected from the group consisting of CO2, N2, air, and an inert gas, wherein the fusion protein is covalently linked to the fibrin. 35. The fibrin foam of claim 34 wherein no more than 25% of the PDGF is released after incubation of the fibrin foam during 3 days at 37° C. in a buffer solution. 36. A method for treating a wound comprising administering the fibrin foam of claim 34 to the wound. 37. The method of claim 36, wherein the wound is an ulcer caused by diabetes. 38. The kit of claim 14, wherein following mixing of the components in the kit, a fibrin foam is formed, wherein the foam releases no more than 25% of the PDGF after incubation during 3 days at 37° C. in a buffer solution. 39. The kit of claim 16, wherein the volume of the fibrinogen solution is about 40 to 60% of the volume of the biocompatible gas.

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