US 2008.0026.032A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2008/0026032 A1 ZUBERY et al. (43) Pub. Date: Jan. 31, 2008 (54) COMPOSITE IMPLANTS FOR PROMOTING Publication Classification BONE REGENERATION AND (51) Int. Cl. AUGMENTATION AND METHODS FOR A6F 2/00 (2006.01) THER PREPARATION AND USE A638/00 (2006.01) A6IP 9/00 (2006.01) (76) Inventors: Yuval ZUBERY, Cochav Yair (52) U.S. Cl. ............................................ 424/423: 514/2 (IL); Arie Goldlust, Ness Ziona (57) ABSTRACT (IL); Thomas Bayer, Tel-Aviv (IL); Eran Nir, Rehovot (IL) Collagen based matrices cross-linked by a reducing Sugar(s) are used for preparing composite matrices, implants and scaffolds. The composite matrices may have at least two Correspondence Address: layers including reducing Sugar cross-linked collagen matri DANEL, SWIRSKY ces of different densities. The composite matrices may be 55 REUVEN ST. used in bone regeneration and/or augmentation applications. BET SHEMESH 99.544 Scaffolds including glycated and/or reducing Sugar cross linked collagen exhibit improved support for cell prolifera (21) Appl. No.: 11/829,111 tion and/or growth and/or differentiation. The denser col lagen matrix of the composite matrices may have a dual effect initially functioning as a cell barrier and later func (22) Filed: Jul. 27, 2007 tioning as an ossification Supporting layer. The composite matrices, implants and scaffolds may be prepared using different collagen types and collagen mixtures and by cross Related U.S. Application Data linking the collagen(s) using a reducing Sugar or a mixture (60) Provisional application No. 60/833,476, filed on Jul. of reducing Sugars. The composite matrices, implants and 27, 2006. scaffolds may include additives and/or living cells. Patent Application Publication Jan. 31, 2008 Sheet 1 of 4 US 2008/0026.032 A1 : ?? ¿? Patent Application Publication Jan. 31, 2008 Sheet 2 of 4 US 2008/0026.032 A1 [×××××××C) FIG.2 Patent Application Publication Jan. 31, 2008 Sheet 4 of 4 US 2008/0026.032 A1 G(5)I,H. [uu]H10NETHONOdS STTO O AWN US 2008/0026032 A1 Jan. 31, 2008 COMPOSITE IMPLANTS FOR PROMOTING 0010) 3. The graft should be safe from risk of disease BONE REGENERATION AND transmission. AUGMENTATION AND METHODS FOR 0011. 4. The graft material should preferably serve as THER PREPARATION AND USE a scaffold that encourages cells to migrate and populate the secluded space of the bone defect. CROSS-REFERENCE TO RELATED US 0012 5. The graft should preferably undergo complete APPLICATIONS degradation within 6-12 months. 0013 6. The graft should preferably mimic bone 0001. This application claims priority from and the ben matrix proteins and should be capable of undergoing efit of U.S. Provisional Patent Application Ser. No. 60/833, ossification. 476 filed on Jul. 27, 2006 entitled “COMPOSITE 0.014 7. Preferably the graft should serve as a carrier IMPLANTS FOR PROMOTING BONE REGENERA for suitable growth factors. TION AND AUGMENTATION AND METHODS FOR 0.015 8. The graft should be easy to handle even by THEIR PREPARATION AND USE' incorporated herein by inexperienced clinicians requiring minimal skills for its reference in its entirety. preparation and implantation to save time and reduce possible complications. FIELD OF THE INVENTION 0016. It would therefore be advantageous to have a bone 0002 The present invention relates generally to implant graft or implant combining as many as possible of the above able devices for promoting regeneration and augmentation properties. of bone and more specifically of composite reducing Sugar cross-linked collagen based matrices, methods for their use SUMMARY OF THE INVENTION and methods for their preparation. 0017. There is therefore provided, in accordance with an embodiment of a method of the present application a method BACKGROUND OF THE INVENTION for preparing a composite multi-density cross-linked col 0003 Alveolar bone loss is secondary to early tooth loss lagen implantable device. The method includes the steps of and periodontal disease, leading to severe functional and compressing a suspension including fibrillated collagen par esthetic problems. In the last three decades the replacement ticles in a first Suspending solution to form a first matrix of missing or hopeless teeth is possible via the use of dental having a first density, applying to the first matrix a suspen implants. These, however require Sufficient bony housing to sion including fibrillated collagen particles in a second accommodate an implant of appropriate length and diameter Suspending solution to form a second matrix attached to the to be able to withstand the oclussal load on the future first matrix the second matrix having a second density lower prosthetic device, and to provide optimal esthetic results. than the first density, drying the first matrix and the second Thus, in many cases, alveolar bone augmentation is man matrix to form a dry multi-density composite matrix, and datory for functional and esthetic long term success of dental reacting the multi-density composite matrix with a reducing implants. Sugar to form the composite multi-density cross-linked 0004. The most common techniques for bone augmenta collagen implantable device. tion procedures involve the use of bone grafts under a barrier 0018. Furthermore, in accordance with an embodiment of that prevents Soft tissue invasion, and allows a selective cell the method of the present application, the step of reacting line with osteogenic capabilities to populate the defect. includes incubating the composite multi-density implantable These are used to facilitate migration and differentiation of device with a reducing Sugar in an incubation Solution mesenchymal cells to form osteoblasts and lay down bone including ethanol. within the defect. In addition, Such devices may serve as a 0019. Furthermore, in accordance with an embodiment of scaffold that Supports cell migration. The grafts may be the method of the present application, the incubation solu derived from natural Sources (human and other animals), or tion includes 70% ethanol. from various synthetic materials, as is known in the art. 0020. Furthermore, in accordance with an embodiment of Bone grafts are normally used as a powder with particle size the method of the present application, the reducing Sugar is ranging from 0.25-2 mm mixed with patient’s blood as a selected from D(-) ribose and DL glyceraldehyde. coagulum or mixed with sterile saline. In some cases, gel or 0021. Furthermore, in accordance with an embodiment of putty like consistency of the implant provide improved the method of the present application, at least one additional handling of the material. Substance is added to at least one of the first Suspending 0005. A major shortcoming of such bone grafts is the Solution, said second Suspension solution, said first matrix, long term resorption and replacement of the graft that may and said second matrix. compromise the mechanical properties of the resulting aug 0022. Furthermore, in accordance with an embodiment of mented bone. the method of the present application, the method also 0006 Similar problems may also be encountered in the includes the step of adding living cells to the composite treatment of various bone defects such as orthopaedic bone implantable device. The cells are selected from cultured deficiencies. These devices (matrices) may be used for cells, stem cells, human cells, animal cells, fibroblasts, augmentation and treatment of bone fractures, and the like. pluripotent bone marrow cells, pluripotent stem cells, bone 0007 Materials for supporting bone augmentation should building cells, osteoblasts, mesenchymal cells, mammalian ideally have the following properties: cells, primary cells, genetically modified cells, nerve cells 0008 1. The ability to mechanically support a barrier. and any combinations thereof. 0009 2. The graft material should be biocompatible 0023 There is also provided, in accordance with an with minimal allergic or immunogenic reactions. embodiment of the implantable device of the present appli US 2008/0026032 A1 Jan. 31, 2008 cation, a composite multi-density cross-linked collagen 0035. There is also provided, in accordance with an implantable device prepared by any of the above methods. embodiment of the methods of the present application, a 0024. There is also provided, in accordance with an method for using a reducing Sugar cross-linked collagen embodiment of the implants of the present application, a matrix as an improved scaffold for cell proliferation and cell composite multi-density cross-linked collagen based differentiation. The method includes the steps of providing implant. The implant includes a first reducing Sugar cross a scaffold comprising a collagen matrix cross-linked with a linked collagen based matrix having a first density and at reducing Sugar, and incubating the scaffold with living cells least a second reducing Sugar cross-linked collagen based to induce improved growth and/or proliferation and/or dif matrix attached to the first reducing Sugar cross-linked ferentiation of the cells. collagen based matrix. The second collagen based matrix 0036 Furthermore, in accordance with an embodiment of has a second density lower than the first density. the methods of the present application, the cells are selected 0025. Furthermore, in accordance with an embodiment of from cultured cells, stem cells, human cells, animal cells, the implants of the present application, the first and the fibroblasts, pluripotent bone marrow cells, pluripotent stem second reducing Sugar cross-linked collagen based matrices cells, bone building cells, osteoblasts, mesenchymal cells, are obtained by cross-linking collagen with a reducing Sugar mammalian cells, primary cells, genetically modified cells, in an incubation Solution including ethanol. nerve cells and any combinations thereof. 0026. Furthermore, in accordance with an embodiment of 0037. Furthermore, in accordance with an embodiment of the implants of the present application, the incubation solu the methods of the present application, the scaffold is tion comprises 70% ethanol. obtained by incubating a collagen based matrix with a 0027.