DE102005016443A1 - Bioabsorbable and mineralized material, useful for filling of bone defects, comprises a collagen matrix from aggregated collagen chains, where only the surface of the aggregated collagen chain is mineralized - Google Patents

Abstract

Bioabsorbable and mineralized material (I), for filling of bone defects, comprises a collagen matrix from aggregated collagen chains, where only the surface of the aggregated collagen chain is mineralized. Independent claims are also included for: (1) a molded article, for the filling of bone defects, comprising (I); and (2) a method for preparing the molded article, for the filling of bone defects with liquid medium (preferably (I)), comprising depositing a mineral substance, preferably from its dissolved ionogenic components, on a collagen matrix, precipitating the mineralized collagen matrix, separating the mineralized collagen matrix, transferring the mineralized collagen matrix into a suspension, transferring the suspension into a predefined form, freeze-drying the suspension in the predefined form. ACTIVITY : Osteopathic. MECHANISM OF ACTION : None given.

Description

The The invention primarily relates to a bioresorbable and mineralized Material for filling Bone defects.

Of the natural Bone consists of about 70% inorganic components, in particular Hydroxylapatite, and about 20% of organic components, in particular from collagen type I, which is formed by bone-forming cells, so-called osteoplasts, produced and secreted or annealed, as well as about 10% from water. To a small extent are also so-called non-collagenous Proteins, in particular osteocalcin, osteonectin and osteopontin, Cytokines, growth factors, proteoglycans and fats present.

Of the natural Bone is a constant waste and construction, a so-called "remodeling" subjected Induction of new bone formation, the so-called osteogenesis, goes as well as in fracture healing from the bone itself. The bone thus owns the fully permanent Information to form functional bones. Bone regeneration can however by the size of one Defective and diverse functional disorders, For example, metabolic diseases, natural limits set.

The Treatment of bone defects in medical care plays especially in orthopedics, Traumatology, maxillofacial surgery and hand surgery a significant role. Bone defects can have different causes for example they are caused by cysts, atrophies, tumors etc. complicated Comminuted fractures, malformations and implant loosening pose the biggest challenges Therapy. Therefore, the focus of treatment is increasingly on the Reconstruction of the defect by suitable filling materials.

to filling Bone defects are a variety of different materials to disposal, especially those that are bad or not bioresorbable are and those that are absorbed in the body become. The former consist for example of porous calcium phosphate ceramics. You have though over sufficient osteoconductive properties, d. H. as a result of the existence have a structural matrix load-bearing properties, but have the disadvantage that they in the body remain and only grow from the new bone.

at The bioabsorbable bone replacement materials are often synthetic polymers, in particular polylactide or polyglycolide. These are degraded in vivo by hydrolysis, depending on the used Polymer to a release of acidic degradation products, for example of lactic acid or glycolic acid, and thus to a local hyperacidity in the body to lead can.

Therefore, the further development of bioresorbable bone substitute materials has led to fillers consisting predominantly of natural, in particular animal, preferably equine, porcine or bovine, bone tissue. These are often demineralized bone materials in which the inorganic portion (minerals) was almost completely removed except for traces. The remaining organic matrix, which consists essentially of collagen, is very close in composition and structure to natural bone. This causes the demineralized bone material to have stabilizing or osteoconductive properties. An example of such a biomatrix is commercially available under the name ^® COLLOSS lyophilized renatured collagen extract the Applicant from bovine bone. COLLOSS ^® supports the bone growth and bone regeneration process through its structural biocompatibility. However, for larger bone defects to be treated, the load bearing capabilities of these materials may not be sufficient. In particular, it is often necessary, especially in the early stages of treatment, to additionally stabilize the affected body segment with metallic materials, for example, plates or intramedullary nails.

One promising approach to increase stability of the collagen-based Bone replacement materials provides the mineralization of collagen (M. Gelinsky, U. King, A. Sewing, W. Pompe: Porous Mineralized collagen scaffolds - a biomimetic bone substitute. Mat.-scientific. u. Werkstofftech. 2004, 35, no. 4). However, this approach will carried out under denaturing conditions. By denaturing the Kollagenmatrix, in particular as a result of acid treatment, it may be a impairment the osteoinductive properties of the bone replacement material come with regard to the formation of endogenous bone or cartilage tissue preferably should be avoided.

The invention therefore has the object to provide an osteoinductive material for filling bone defects, which substantially corresponds to the natural bone composition. This material is said to have significantly improved osteoconductive properties compared to conventional collagen-based bone substitutes, ie a verbes serte have load-bearing and / or volume-stable function in the reconstruction of endogenous bones own.

These Task is solved by a material as described in claim 1. Preferred embodiments of this material are in the dependent claims 2 to 18 executed. Moldings, which are at least partially made of the material according to the invention are in the claims 19 and 20 shown. The requirements 21 to 27 relate to a suitable process for the preparation such a shaped body to the filling of bone defects. By reference, the wording of all claims to the content of the description.

at the material of the invention to the filling Bone defects are bioresorbable and mineralized material that includes a collagen matrix comprising aggregated collagen chains or molecules, wherein substantially only the surface of the assembled or the individual collagen collagen chains is, d. H. surrounded by crystallites of a mineral substance. Preferably, the collagen is mainly collagen Type I.

at the material to be mineralized may be a native collagen matrix with triple helical collagen chains (single collagen chains are too a triple helix together) act. This is special preferred because the resulting network of collagen fibers and / or fibrils are present in its natural state and not in an artificial one spatial Arrangement must be brought. This has the consequence that already in the natural one Condition left organic matrix of collagen osteoconductive or has load-carrying and / or volume-stable properties. Preferably is the native collagen matrix, especially a matrix of collagen Type I mineralized both within the matrix and at its surface. This carries to an additional Increase the osteoconductive properties of the collagen matrix.

In a particularly preferred embodiment, the material according to the invention is an originally denatured and then renatured renatured collagen matrix, the collagen chains being assembled to form a superstructure, preferably a network-like structure. Advantageously, it is in such a collagen matrix to the material COLLOSS ^® by the applicant.

It may also be according to the invention provided that a portion of the collagen chains present in a triple-helical structure, so that the renatured collagen matrix, in particular COLLOSS ^®, both areas with triple-helical structures, as well as superstructures, in particular net-like structures having,.

Preferably, the collagen matrix contains both native collagen and renatured collagen. The native collagen is preferably pure collagen, in particular type I collagen, which is produced, for example, from tendons or skin. The native collagen is used due to its aforementioned osteoconductive properties as stabilizing component for the structure of the renatured collagen matrix, preferably COLLOSS ^®.

In a particularly preferred embodiment the material of the invention has osteoinductive Properties. Preferably, the material according to the invention even by the presence of a native or renatured collagen matrix already osteoinductive properties. But it can also be preferred be, an osteoinductivity of the material by doping with corresponding osteoinductive To achieve active ingredients.

The material according to the invention preferably has at least one active substance, in particular an active substance which activates and / or stimulates the osteogenesis. This may be particularly preferred to enhance the osteoinductive properties of the mineralized material. Preferably, the active ingredient is of native origin. Such an active ingredient is to be understood as meaning a natural active ingredient which is present in its natural structure (conformation) and which is present in a substantially native or renatured collagen matrix. According to the invention, it is provided that the active ingredient can be present within the collagen matrix and / or on its surface. The material COLLOSS ^® by the applicant is a collagen matrix, which comprises at least one native active substance, preferably a plurality of the active ingredients described below.

With Advantage contains the material of the invention so-called recruiting factors, in particular chemotactic agents (chemotaxins), For example, leukotrienes, which targeted body cells, preferably mesenchymal Stem cells, cartilage progenitor cells, Fibroblasts and / or platelets, resulting in a penetration of the bioresorbable and mineralized material.

It may also be preferred that the filler according to the invention ent so-called adhesion factors for immobilization of immigrated cells on or in the mineralized material ent holds. The fiction, contemporary material is characterized advantageously characterized in that it contains in particular cytotactin, tenascin, laminin and / or fibronectin as adhesins.

In a particularly preferred embodiment the invention, the material according to the invention growth and / or Maturation factors for the proliferation and differentiation of immigrant Cells, especially cytokines on. It is preferable in the growth factors around bone growth factors, for example BMP (bovine morphogenetic protein), in particular BMP-II, BMP-VII and / or BMP-IV, as well as IGF (insuline-like growth factor), in particular in particular IGF-I, and TGF (transforming growth factor) TGF-βI. Preferably contains the material of the invention FGF (fibroblast growth factor) and PDG (platelet derived growth factor) as further growth factors.

Advantageously, the bioresorbable and mineralized material may have a combination of the active ingredients described above. This may be particularly preferred in order to achieve over-additive or synergistic effects in the reconstruction of a bone defect by the body's own material. According to the invention, it is further provided that the active substances, in particular the active substances already mentioned, partially overlap in their function and mode of action. In this way, the failure of one or more active ingredients can be compensated in an advantageous manner, without causing an impairment, in particular to a reduction in their success, preferably the Osteoinduktivität comes. With regard to the examples of native drugs described above, reference is made to the EP 0 500 556 B1 directed.

In addition, can it is preferred that the bioresorbable and mineralized material comprises an antimicrobial agent. Advantageously, the antimicrobial agent is an antibiotic, the possible Immune defense reactions in the body avoids or suppresses.

Farther Is it possible, that this Material is equipped with a cytostatic. This is special in cancerous changes in the area of the bone defect and / or the surrounding tissue advantageous. But also the use of other active ingredients, for example of antibodies, in particular for therapeutic reasons, may be useful in this context. In a further education of the invention the various active substances exemplified as well as other active ingredients be combined with each other for particularly advantageous effects, especially superadditive or synergistic effects. Regarding the compensation of a Active ingredient failure is taken to the previously mentioned reference.

In Another particularly preferred embodiment of the invention the material of the invention on the surface the collagen collagen chains with calcium phosphate and / or Calcium carbonate and / or hydroxyapatite mineralized. calcium phosphate is in the form of hydroxyapatite (calcium phosphate hydroxide) main component of mineral bone substance, 50% of bone volume accounts. Together with calcium carbonate it essentially determines the hardness of the natural Bone. Both calcium phosphate and calcium carbonate are suitable Therefore, in a preferred manner, the osteoconductive properties or to increase the load-bearing properties of a collagen matrix. By the superficial Calcium phosphate and / or calcium carbonate deposit on the pooled Collagen chains there is a subsequent reinforcement of Collagen matrix.

In In a further development of the invention, the material has pores, preferably interconnecting pores, on. The latter means that the Po renstruktur not only between the individual particles of the material according to the invention, but also within the particle is optimized. Besides that is the interconnecting porosity in particular advantageously also for Cells accessible. Furthermore, it may be preferable to use the active substances described here, but also all other active substances, on the inner and / or outer surface of the material according to the invention showcase, for these substances for the body, especially for endogenous Cells, available close.

In a preferred embodiment the pores of the material according to the invention have pore diameters of at least 100 μm, in particular pore diameters between 100 and 300 microns, preferably of about 200 microns. This is particularly beneficial because of the ingrowth of bone cells in the porous one Inventive material Pore diameters of at least 100 microns are required. Thereby is the integration of the material according to the invention in the body as well promoted its degradation or absorption.

In In a further development of the invention, the material is sterilized or is in sterilized form. For the sterilization of the material according to the invention come a variety of conventional methods into consideration. Prefers is the sterilization of bioresorbable and mineralized Materials by radioactive irradiation, preferably by gamma irradiation. Farther it is preferred that the material according to the invention is fumigated with ethylene oxide.

In Another embodiment of the invention is the material of the invention aseptically treated and / or manufactured, for example by use of sterile-filtered antibiotic solutions, preferably one sterile-filtered gentamycin solution. This may be particularly preferred because in this way the possible Problems of sterilization, such as an adverse effect the added active ingredients and / or the osteoinductive properties of the material, to be avoided.

According to the invention the material in packaged form, in particular in sterile packaged form, in front. In such a package, the material over a longer Period, in particular about be stored for a period of several months without it to an impairment the effect of bioresorbable and mineralized material, in particular to an impairment its osteoinductive properties, comes. In case of application The material can be easily removed from the packaging and be used. To the sterility of the material according to the invention It may also be preferable to use the material within sterilize the packaging, for example by irradiation or ethylene oxide gasification. Suitable as packaging materials in particular plastic materials.

In a further preferred embodiment of the invention, the material additionally contains a mixture of a predominantly native and / or renatured collagen matrix and hyaluronic acid, wherein the mixture is substantially spatially separate from the mineralized collagen matrix. Preferably, the mixture COLLOSS ^® contains as additional collagen component. Hyaluronic acid is a component of the so-called synovial fluid, which is responsible for the nutrient supply of the cartilage by diffusion as a "synovial fluid." Preferably, the composition is composed of mineralized collagen matrix and the mixture of a predominantly native collagen matrix and hyaluronic acid, preferably also mineralized together Treatment of chondral defects, especially osteochondral defects.

at the last-mentioned embodiments it is preferred that the spatial separated shares of this composition physically interconnected are, in particular by a chemical cross-linking. Preferably is the composition with a carbodiimide, in particular with N- (dimethylaminopropyl) -N'-ethylcarbodiimide (EDC), cross-linked. The chemical cross-linking leads to a further stabilization this material according to the invention.

The Invention Furthermore a shaped body to the filling of bone defects, said shaped body being a material according to the invention having the properties described above, preferably made of this material. Under moldings are therefore aggregates mineralized, essentially native and / or renatured To understand collagen matrix, which may be another component, For example, a mixture of predominantly native and / or renatured collagen matrix and hyaluronic acid. Preferably lie these shaped bodies as three-dimensional shaped bodies, especially as beams, rings or cylinders. It is also possible, that the moldings as a cuboid, slices or the like available. It can be shaped body create any shapes.

In a preferred embodiment The invention relates to the shaped bodies by a chemical cross-linking, in particular with a carbodiimide, preferably further stabilized with N- (dimethylaminopropyl) -N'-ethylcarbodiimide (EDC). Farther can the moldings be stabilized by the entry of a binder and / or salt. The thus obtained molded body keep their basic shape after moistening with liquids. They may preferably after compression approximate their original shape, d. H. she have elastic Properties.

• – Abscheidung einer mineralischen Substanz, vorzugsweise aus ihren gelösten ionogenen Komponenten, auf eine Kollagenmatrix,
• – Ausfällung der mineralisierten Kollagenmatrix,
• – Abtrennung der mineralisierten Kollagenmatrix,
• – Überführung der mineralisierten Kollagenmatrix in eine Suspension,
• – Überführung der Suspension in eine den Formkörper definierende Form,
• – Gefriertrocknung der Suspension in der Form.

Finally, the invention comprises a process for producing a shaped body from a liquid medium, preferably for filling bone defects, ie essentially from a material with the properties described above, comprising the steps

• Deposition of a mineral substance, preferably from its dissolved ionogenic components, onto a collagen matrix,
• Precipitation of the mineralized collagen matrix,
• Separation of the mineralized collagen matrix,
• Transfer of the mineralized collagen matrix into a suspension,
• Transfer of the suspension into a mold which defines the shaped body,
• - Freeze-drying of the suspension in the mold.

Preferably, solutions, in particular aqueous solutions, of the ionogenic components which form the mineral substance to be deposited are added to a collagen suspension. Preferably, the suspension contains native or renatured collagen, preferably COLLOSS ^®. Furthermore, it is possible that the minerals are a waste product of COLLOSS ^® -Herstellungsprozesses and are added back to a Collos ^® suspension.

In a particularly preferred manner, the Ab The mineral substance is deposited essentially only on the surface of the collagen matrix, preferably on the surface of the collagen collagen chains. The deposition of the mineral substance is carried out in a neutral pH range, in particular in a pH range between 6 and 8, preferably at a pH of 7, in order to achieve the selective superficial deposition on the collagen matrix.

Preferably becomes the precipitate mineralized collagen matrix by exceeding the solubility product the mineral substance in solution achieved. This is outwardly at the formation of a precipitate, which is essentially mineralized Contains collagen, recognizable. It must the experimental parameters, in particular the concentration of the mineral Substance and pH, adjusted so that a synchronous Assembly of collagen and mineral substance takes place.

advantageously, becomes the precipitate the collagen matrix with calcium phosphate and / or calcium carbonate and / or hydroxyapatite carried out as a mineral substance.

According to the invention, it may be preferred to add native pure collagen, in particular collagen type I, before precipitation of the mineralized collagen matrix, preferably before precipitation of the mineral substance on the collagen matrix. Advantageously, the native pure collagen is added in the form of a suspension. The added collagen is used, as already mentioned, as a stabilizing component and leads to a strengthening of the structure, in particular an increase in the osteoconductive properties of the collagen matrix, preferably COLLOSS ^®. A subsequent mineralization (deposition of the mineral substance onto the collagen matrix and precipitation of the mineralized collagen matrix) causes to be stabilized collagen matrix, preferably COLLOSS ^®, and the native pure collagen are covered together by a layer of the mineral substance.

In another preferred embodiment of the invention, the mineralization of the collagen matrix is, preferably COLLOSS ^®, and the native pure collagen, particularly of type I collagen, performed separately. The resulting suspensions of mineralized collagen matrix and mineralized native pure collagen are mixed and further processed, in particular, according to one of the following steps.

In a development of the method according to the invention is the mineralized Collagen after precipitation separated for example by centrifugation of the solution.

The from the solution Separated mineralized collagen is inventively in a preferably pourable and transferred to homogeneous suspension. These Suspension can in appropriate mold cavities, in particular in the wells of a corrugated plate, such as a 24er or 96er corrugated sheet of polystyrene, to be poured. By a Freeze drying, dependent from the shapes of the depressions, different three-dimensional porous Moldings, in particular beams, rings, cylinders, discs, cuboids or the like, getting produced. By choice of speed and temperature the freeze-drying process, the pore size in particularly advantageous Way influenced become. Preference is given to the transferred into the mold cavities of a corrugated plate suspension volumes slow at temperatures between -10 ° C and -40 ° C frozen and in an oil pump vacuum freeze-dried.

Preferably become the shaped bodies thus obtained by a chemical cross-linking, in particular with a carbodiimide, preferably crosslinked with N- (dimethylaminopropyl) -N'-ethylcarbodiimide (EDC). Advantageously, the moldings produced in this way retain their shape Moisten with liquids their basic form and also find their way Compresses almost their original form again.

In a further preferred embodiment of the invention, a pure from native collagen, particularly of type I collagen, existing molding with the collagen matrix, preferably COLLOSS ^®, be doped or modified. The collagen matrix, preferably COLLOSS ^© , can be applied to the shaped body, in particular a collagen sponge, in the form of a suspension or a gel, and preferably be connected to it by freeze-drying.

The present material for filling bone defects according to the invention is a bone substitute material which combines osteoinductive and osteoconductive properties in a particularly advantageous manner. This is preferably done by the subsequent reinforcement of the collagen matrix by the deposition of a mineral substance on the surface of the collagen fibrils. The mechanical reinforcement of the substantially native or renatured collagen matrix requires the preservation of osteoinductive properties of the material, which can be specifically enhanced by the addition of further stimulating the induction of osteogenesis substances. The material according to the invention is therefore particularly suitable for the treatment of large and complicated bones defects in which sufficient stability of the bone substitute material and rapid ingrowth of bone cells into the bone replacement material is required.

Further Features and advantages of the invention will become apparent from the following Description of a preferred embodiment by way of example in conjunction with the subclaims. in this connection can the individual features in each case for themselves realized alone or in combination with each other be.

example

1 g of collagen suspension (COLLOSS ^® Applicant) is made up to 1 l with water and then added with vigorous stirring sequentially with the following solutions: 500 ml of distilled water, 180 ml of 0.1 molar calcium chloride solution, 116 ml of 2 molar sodium chloride solution, 168 ml 0, 5 molar Tris buffer (pH = 7.52) and 22.6 ml of 0.5 molar phosphate buffer according to Sörensen (pH = 7.40). The mixture is made up to 2 liters with distilled water and allowed to stand in water for 12 hours at 35 ° C. The colorless, gelatinous precipitate which forms is centrifuged off and then stirred up again with enough mother liquor to give a pourable and homogeneous suspension. To prepare the cylindrical standard samples, about 2.5 ml each of the suspension are poured into the wells of a 24-well polystyrene cell culture plate and slowly frozen at -25 ° C. The frozen material is freeze-dried in an oil pump vacuum and then chemically crosslinked in a 1% solution of N- (dimethylaminopropyl) -N'-ethylcarbodiimide hydrochloride (EDC, Aldrich) in 80% ethanol for 1 hour. The porous scaffolds are then washed thoroughly with distilled water, then with a 1% glycine solution and finally with water again and then freeze-dried again. The scaffolds are either aseptically prepared, gamma sterilized or fumigated with ethylene oxide for sterility. In the product thus obtained, the individual collagen chains which form the net-like structure are enveloped by calcium phosphate crystallites.

Claims (27)

Bioresorbable and mineralized material for filling bone defects, characterized in that it comprises a collagen matrix of collagen chains stored together, wherein substantially only the surface of the collagen collagen chains is mineralized. Material according to claim 1, characterized that the Collagen matrix contains native and / or renatured collagen. Material according to claim 2, characterized that it the native collagen is pure collagen, especially um Type I collagen acts. Material according to one of the preceding claims, characterized characterized in that Collagen chains assembled into a triple helical structure are. Material according to claim 1 or 2, characterized that the Collagen chains to a superstructure, preferably a net-like Structure, are stored together. Material according to one of the preceding claims, characterized characterized in that it possesses osteoinductive properties. Material according to one of the preceding claims, characterized characterized in that it has at least one active ingredient. Material according to claim 7, characterized that it is an active substance of natural origin. Material according to claim 8, characterized that it a recruiting and / or adhesion and / or growth and / or Maturation factor acts. Material according to claim 8 or 9, characterized that it a cytostatic and / or an antimicrobial, in particular antibiotic, Substance acts. Material according to one of the preceding claims, characterized characterized in that surface of the collagen collagen chains mineralized, in particular with calcium phosphate and / or calcium carbonate and / or hydroxyapatite. Material according to one of the preceding claims, characterized characterized in that it Pores, preferably interconnecting pores, has. Material according to claim 12, characterized that the Pore Pore diameter of at least 100 .mu.m, in particular pore diameter between 100 and 300 μm, preferably of about 200 μm, have. Material according to one of the preceding claims, characterized in that it is sterilized, preferably by gamma irradiation and / or ethylene oxide gassing, and / or aseptically treated and / or manufactured. Material according to one of the preceding claims, characterized characterized in that it in packaged form, especially in sterile packaged form. Material according to one of the preceding claims, characterized characterized in that it in addition one Mixture of a predominantly native and / or renatured collagen matrix and hyaluronic acid is preferred mineralized together, with the mixture substantially spatially separated of the pure mineralized collagen matrix. Material according to claim 16, characterized that the Mixture and the mineralized collagen matrix interconnected are, in particular by a chemical cross-linking. Material according to claim 17, characterized that the Mixture and the mineralized collagen matrix with a carbodiimide, especially with N- (dimethylaminopropyl) -N'-ethylcarbodiimide (EDC) are. moldings to the filling of bone defects, characterized in that it comprises a material according to a of the preceding claims has, preferably consists of this material. moldings according to claim 19, characterized in that it as a three-dimensional Moldings, especially as a bar, ring or cylinder exists. Process for producing a shaped body for filling of bone defects from a liquid Medium, in particular of essentially one material after one the claims 1 to 14, comprising the steps - deposition of a mineral Substance, preferably from its dissolved ionogenic components, on a collagen matrix, Precipitation of the mineralized collagen matrix, - Separation of mineralized Collagen matrix, - transfer of the mineralized collagen matrix into a suspension, - transfer of the Suspension in a the shaped body defining form, - freeze-drying the suspension in the mold. Method according to claim 21, characterized that the Deposition of the mineral substance essentially only on the surface the collagen matrix is made. Method according to claim 21 or 22, characterized that the precipitation mineralized collagen matrix by exceeding the solubility product the mineral substance is initiated. Method according to one of claims 21 to 23, characterized that the precipitation the collagen matrix with calcium phosphate and / or calcium carbonate and / or hydroxyapatite is carried out as a mineral substance. Method according to one of Claims 21 to 24, characterized that before precipitation the mineralized collagen matrix, especially before deposition mineral substance on the collagen matrix, native virgin Collagen, preferably in the form of a suspension is added. Method according to one of claims 21 to 25, characterized that one Suspension of the mineralized collagen matrix and a suspension mineralized native pure collagen mixed and processed become. Method according to one of claims 21 to 26, characterized the existence from native pure collagen, in particular type I collagen, existing molding is modified with the collagen matrix.