CA1333050C - Biologic absorbable implant material for filling and closing soft-tissue cavities and method of its preparation - Google Patents

Abstract

There is disclosed a biologic absorbable implant material for filling and closing soft-tissue cavities and for replacing soft-tissue parts, and a method of its preparation. The implant material consists essentially of bone tissue of human or animal origin, is decalcified, and the original protein content of the osseous hard substance is unchanged. By cross-linking of these native proteins with a cross-linking agent, a high elasticity, a good stability and a marked memory-effect are attained. The method for the preparation of the implant material includes degreasing, decalcification and cross-linking of the available proteins and then drying and sterilizing of the material.

Description

`-- -The invention relates to a biologic absorbable implant material for filling and closing soft-tissue cavities and for replacing soft-tissue parts, as well as to a method of its preparation.
In the field of orthopedics, implant materials for filling bone cavities are known, which are produced by partial deproteinisation and denaturation of the residual protein from spongiosa bone tissue (German Patent No.
961,654). This material, which is known as "Kieler spongiosa", basically has the same calcium content as native spongiosa. Consequently, it is hard and totally inelastic, thus being unsuited for application in soft-tissue parts.
In bone surgery, a material produced from cortical bones, i.e., tubular bones, by decalcification is known (Journal of Surgical Research 37, 487-496 (1984)), which has an osteoinductive effect supposed to be due to its content of bone morphogenetic protein. On account of this osteoinductive property, its use outside bones is contra-indicated.
In EP-A No. 0,171,176, a composition for repairing bone defects is described, which is prepared from bone collagen by decalcification and lyophilization, forming a gel upon reconstitution. On account of the desired osteoinductive effect of this material, care is taken that no cross-link formation of the collagen occurs during preparation.
Furthermore, a collagen-based bone substitute material is known from German Auslegeschrlft No. 28 54 490. It is prepared from bones by removing the blood pigment and other ..} ~., -water-soluble proteins, degreasing and eliminating the mineral portions by means of complexing agents or ion .-exchangers and finally freeze-drying. Upon reconstitution, the material is insufficiently elastic and not suited for application in soft tissue parts.
So far, only collagen fleeces have become known as coverage and implant materials for the application at or in soft tissue parts. However, such fleeces have no dimensional stability and no elasticity in the moist state;
they are suited for firm bonding to mobile tissue, e.g., to the lung, to a limited extent only. Synthetic materials, such as acrylates, which likewisely have been proposed, still have other disadvantages, among which massive stimulus caused by a foreign body, encapsulation, insufficient healing and the possible release of toxic decomposition and side products are particularly emphasized.
Further known prior art prosthetic implant materials are produced from body tissues by treatment with protein cross-linking agents. Such materials are used in order to increase the stability of certain organs, e.g., of cardiac valves or vascular prostheses. Such implants are described, for instance, in EP-B No. 0,174,737 and in U.S. Patent No.
4,120,649.
The invention has as its object to provide a well tolerated implant material that may be used to close tissue cavities, on the one hand, and to substitute certain tissue parts, on the other hand, and which does not have the disadvantageous properties of the known products, which limit their therapeutic utilization.

1 3 ~3050 ;
-After lung resections and also after other diseases ofthe lung, bronchopleural fistulas may occur. These are septic openings or cavities forming between the bronchial tree and the pleura and filled with secretions and pus.
They are constantly passed by air and secretions during breathing and coughing, their sizes ranging from a few millimeters to several centimeters in terms of diameter.
Therefore, it is a still unsolved problem in thorax surgery how to seal such bronchial fistulas and how to enhance curing of the same, in particular, if the introduction of the implant is to be effected by way of endoscopy, which were the quickest and mildest way. Special demands are set on an implant material to be introduced and fixed endoscopically. On the one hand, mechanical spreading must be feasible, on the other hand, introduction through the bronchial tree to the fistula must be possible.
The implant must be deformable and compressible. It must be able to reassume its original shape in the presence of moisture, i.e., it must have a high memory effect. In addition, the material must offer a considerable strength, yet remain absorbable, because germs may adhere to non-absorbable materials, thus causing abscesses and new fistulas again and again. Besides, according to a further aspect of the invention, it is to be possible to obtain a gas- and liquid-tight closure immediately upon introduction of the implant. Finally, it is to be possible to incorporate into the material bactericidal and/or other pharmaceutical substances or such stimulating wound healing.
The invention by which these objects are achieved with ..~,...

~ - 1 333050 242g2-444 -a biologic absorbable implant material for filling and closing soft-tissue cavities and for replacing soft-tissue parts consists in that - the implant material consists essentially of spongiosa bone tissue of human or animal origin molded into shaped products, - which is decalcified, - vhose native proteins are cross-linked with a protein cross-linking agent in order to avoid osteoinductive activity, - the shaped products exhibiting a high elasticity at a lov hysteresis between loaded and unloaded conditions.
Thus, according to the invention, the initial protein content of the decalcified osseous hard substance remains almost unchanged and the high elasticity and stability are brought about by the cross-link formation of the native proteins.
The shaped products formed of bone tissue may, for instance, have block, conical or spherical shapes.
To fill and close tissue cavities, the implant material may have spongy consistency, the pore volume of the material belng 55 to 95%. The material is compressible in the dry state; however, it reassumes its original shape in the presence of moi~ture, which means that it has a marked memory effect.
Decalcification is continued until the residual ~_ 1 3 3 3 0 5 0 content of calcium amounts to 80 mMol/g wet weight at the most.
~ According to a preferred embodiment, the implant material may contain a tissue adhesive on fibrinogen or collagen base.
The invention, furthermore, relates to a method of preparing the implant material, which is characterized in that native protein-containing bone material is decalcified by a decalcifying agent, is washed free from the decalcification agent, then is treated with a protein cross-linking agent, is repeatedly washed to remove the protein cross-linking agent, if desired, is combined with pharmaceutical solutions, such as tissue adhesive solutions on fibrinogen or collagen base, and finally is dried and sterilized.
As the cross-linking agents, aldehydes, in particular glutaraldehyde, polyepoxide compounds, diisocyanate compounds or carbodiimides may be used.
The bone material, suitably, may be degreased by means of an organic solvent, such as chloroform-methanol, before or after decalcification.
There may also be provided to treat the bone material with agents that reduce possibly present contents of undesired antigens, e.g., detergents or enzymes, before or after protein cross-linking.
The implant material according to the invention and the method of its preparation may be modified in various aspects to impart additional pharmaceutical effects and properties. Thus, when using a detergent, e.g., octylphenoxy-polyethoxyethano] Triton* X-100 in a one-*Trade-mark - 5 -1 3330~0 '`
percent concentration, in the washing solutions, it is not only possible to reduce the content of undesired antigens, but the danger of a subsequent calcification of the implant may be lowered, too. Moreover, a low content (about 0.5 M) of ethylenediaminetetraacid sodium may be provided in the washing solutions, whereby decalcification is improved. i, The intermediate treatment of the implant material with a buffer solution containing enzymatically effective substances, for instance, ficin and/or neuraiminidase, may be provided, which likewisely reduce the antigen content in the finished product.
In order to make sure that the finished product will not contain any traces of glutaraldehyde, the material may be rinsed with a glycine or lysine solution (content about 5 mg/ml) after the cross-linking treatment.
If the implant material according to the invention has been produced from spongiosa, it is suitable if the implant material is compressed, approximately to half its volume, prior to being filled into final containers.
20A special field of application of the material according to the invention is the substitution for tubular tissue parts, such as trachea parts. As the starting material for such an implant, femur bones are used, which are tubularly reduced to a wall thickness of about 2 mm by mechanical surface abrasion on a lathe, before being subjected to the combined treatment by decalcification and protein cross-linking according to the invention.
The end product according to the invention, if destined to fill tissue cavities, suitably is brought into block shape.

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1 33305() The material may be sterilized by gamma rays and may be stored in containers, which, for instance, contain a 70 -% ethanol solution. The blocks are rinsed with a Ringer solution prior to being applied into patients.
It is also possible to store the blocks in a dry state after sterilization with gamma rays, wherein, according to a particularly preferred embodiment of the invention, the blocks are soaked with a fibrin sealant under sterile conditions and subsequently are lyophilized in the final container.
Naturally, the product according to the invention, in addition to the block or tubular shape already described, may be made available also in disk or sheet form.
The method according to the invention and the product produced thereby will be explained in more detail in the following example.
1 kg of spongiosa bone material from the calf was obtained by cutting the head of a femur into cubes.
These spongiosa cubes were rinsed with hot water (50C) and subsequently were decalcified at room tempera-ture in 0.6N hydrochloric acid (10 1) under continuous motion and by repeatedly changing, pressing and washing the spongiosa blocks (24 to 72 hours). Subsequently, the blocks were washed free of acid and incubated with 1 % glutaralde-hyde in a 0.05 M phosphate buffer solution (1 1) at a pH of 7.4 at room temperature for one hour so as to effect cross-linking.
By at least five times of washing with a sterile washing solution and pressing of the spongiosa, the residual glutaraldehyde was removed from the spongiosa. The `, 1 3~30~0 -bloeks were freeze-dried, filled into final containers and sterilized within the same by means of 2.5 Mrad gamma rays.
.- A thus produced implant material in block form was inserted into patients as a fistula closure by way of endoscopy: The patient was intubated with a rigid bronehoseope and respiration in the open system was performed by means of jet ventilation. After inspection of the fistula, a suitable spongiosa piece was cut and eompressed so as to be able to be guided through the bronehus with the bronchoseope. The implant was placed into the fistula by biopsy forceps, where it got moistened upon eontaet with the bronehial wall, expanding very rapidly and spreading. Immediately upon setting of the implant, 1 to 2 ml of a fibrin sealant were dropped about the implant by a three-lumen spraying eatheter, the implant, thus, swelling further and bonding with the bronehial wall. An immediate air-tight elosure of the fistula was obtained.
The implant material aeeording to the invention does not exert any osteoinductive effect, which can be proved by measuring alkaline phosphatase and by histologic investigations. Consequently, the material can safely be used for application in soft tissue regions.
The implant material produced according to the invention has excellent elasticity properties. For the purpose of eomparison with known preparations, spongiosa bloeks having an edge length of 1.5 cm were produced, once without treatment with a protein cross-linking agent and once with the treatment according to the invention by eombined deealeification and protein cross-linking as described in the above embodiment. The blocks were dried 1 3~30~0 and then were moistened with a 0.9 % NaCl solution. All the blocks were checked in a loading apparatus (transducer 0 -.1000 pond at a loading speed of 1 cm/min), with load andrelief curves resulting, which are illustrated in the drawing.
The load curve and the relief curve (power - distance) of the spongiosa blocks that have not been subjected to cross-linking are denoted by 1 and 2; 1' and 2' represent the respective curves of the preparations according to the invention. It is apparent that, with the prior art preparations, which have not been cross-linked, a relatively low strength and an intense hysteresis occur, whereas the spongiosa blocks according to the invention do not develop any signs of hysteresis and exhibit high strength values.
The tissue tolerance of spongiosa blocks according to the invention, having the dimensions of 0.5 x 0.5 x 1 cm and produced according to the exemplary embodiment, was tested in rat experiments. The blocks were implanted subcutaneously into the backs of the rats, were taken at various time intervals, were fixed and processed histologically.
It appeared that noncross-linked spongiosa blocks showed more intensive inflammatory reactions, more foreign-body giant cells, more fibrosis and the tendency to absorption already after a fortnight, clear signs of dis-integration occurring after 4 weeks. In contrast, no such side effects could be observed with the samples treated according to the invention.

Claims (18)

1. A biologically absorbable implant material for filling or closing soft-tissue cavities or for replacing soft-tissue parts, which consists essentially of a shaped body molded from decalcified spongiosa bone tissue of human or animal origin whose native proteins are cross-linked by a protein cross-linking agent so as to avoid an osteoinductive effect, the shaped body having a high elasticity at a low hysteresis between loaded and unloaded states.

2. An implant material according to claim 1 which has a spongy consistency, the pore volume of the material being from 55 to 95%.

3. An implant material according to claim 2, which is compressible in a dry state, yet resumes its original shape in the presence of moisture, namely which has a marked memory effect.

4. An implant material according to claim 3, which has a residual calcium content of not more than 80 mmol/g wet weight.

5. An implant material according to any one of claims 1 through 4, which further comprises a pharmaceutically acceptable substance impregnated therein.

6. An implant material according to claim 5, wherein the pharmaceutically acceptable substance is a tissue adhesive based on fibrinogen or collagen.

7. A method of producing a shaped biologically absorbable implant material for filling or closing soft-tissue cavities or for replacing soft-tissue parts, which method comprises:
(a) decalcifying native protein-containing spongiosa bone material of human or animal origin with a decalcifying agent, (b) washing the decalcified material until the material becomes free of the decalcifying agent, (c) treating the washed material with a protein cross-linking agent, (d) repeatedly washing the material to remove the cross-linking agent, and (e) finally drying and sterilizing the material.

8. A method according to claim 7, which further comprises:
(f) impregnating the material with a solution containing a pharmaceutically acceptable substance between the repeated washing step (d) and the drying and sterilizing step (e).

9. A method according to claim 8, wherein the pharmaceutically acceptable substance is a tissue adhesive based on fibrinogen or collagen.

10. A method according to claim 7, 8 or 9, wherein the protein cross-linking agent is an aldehyde, a polyepoxide compound, a diisocyanate compound or a carbodimide.

11. A method according to claim 10, wherein the protein cross-linking agent is glutaraldehyde.

12. A method according to claim 7, 8 or 9, wherein the bone material is degreased with an organic solvent before or after the decalcification step (a).

13. A method according to claim 12, wherein the organic solvent is chloroform-methanol.

14. A method according to claim 7, 8 or 9, wherein the bone material is treated, before or after the protein cross-linking agent treating step (c), with an agent which reduces a content of undesired antigens.

15. A method according to claim 14, wherein the agent which reduces a content of undesired antigens is a detergent or an enzyme.

16. A method of producing a shaped biologically absorbable implant material for filling or closing soft-tissue cavities or for replacing soft-tissue parts, which method comprises:
(a) decalcifying shaped native protein-containing spongiosa bone material of animal origin with hydrochloric acid as a decalcifying agent until a calcium content of no more than 80 mMol/g wet weight is attained, (b) washing the decalcified material until the material becomes free of the decalcifying agent, (c) treating the washed material with a protein cross-linking agent selected from the group consisting of an aldehyde, a polyepoxide compound, a diisocyanate compound and a carbodiimide, (d) repeatedly washing the thus-treated material to remove the cross-linking agent, and (e) finally drying and sterilizing the material, wherein (i) the product material has such a spongy consistency that a pore volume thereof is from 55 to 95% and (ii) the produced material is compressible in a dry state but resumes its original shape in the presence of moisture.

17. A use of the implant material as defined in any one of claims 1 through 4, as a replacement of a trachea piece.

18. A use of the implant material as defined in any one of claims 1 through 4, for filling or closing a soft-tissue cavity.