A method for volumetric expansion of a composite matrix, osteogenic compositions and uses

Abstract

This invention discloses a method for expanding the volume of a composite matrix, an osteogenic composition, and its applications. The components of this invention are entirely autologous, posing no risk of immune rejection or disease transmission. The composite matrix in the osteogenic composition provides a bone conduction scaffold, while the volume-expanding agent provides bone-inducing signals while simultaneously expanding the composite matrix, thus providing a dual osteogenic mechanism. This invention requires only a conventional centrifuge and sterile containers, making it suitable for outpatient surgery. It also eliminates the need to purchase expensive bone powder, offering advantages such as ease of operation and low cost. Furthermore, the materials of this invention exhibit good stability; the composite matrix, after being encapsulated by the volume-expanding agent, does not easily dissipate and conforms well to the defect morphology.

Description

Technical Field

[0001] This invention relates to the field of biomedical materials technology, and more specifically to a method for expanding the volume of a composite matrix, an osteogenic composition, and its applications. Background Technology

[0002] Bone defect repair is a common challenge in clinical fields such as dental implantology, maxillofacial surgery, trauma orthopedics, and spinal fusion. To restore bone volume and structural integrity, bone augmentation materials are often used. Ideal bone augmentation materials should possess good biocompatibility, osteoconductivity, and osteoinductive properties, while minimizing the risk of immune rejection and disease transmission.

[0003] Currently, commonly used bone augmentation materials in clinical practice include autologous bone, allogeneic / xenogeneic bone, and synthetic bone. Autologous bone possesses excellent osteogenic capacity, but it suffers from donor site complications and limited bone harvest. While allogeneic / xenogeneic bone avoids the need for autologous bone harvesting, it raises concerns about immune rejection and ethical controversies. Synthetic bone substitutes (such as hydroxyapatite and β-tricalcium phosphate) offer advantages in biocompatibility, but they lack bioactive factors, exhibit weak osteoinductive capacity, and their degradation rate is difficult to match with new bone formation. Therefore, there is still an urgent need for a completely autologous, easy-to-operate, and highly efficient bone augmentation method.

[0004] The information in the background section is merely intended to illustrate the general background of the invention and should not be construed as an admission or implication in any way that such information constitutes prior art known to those skilled in the art. Summary of the Invention

[0005] To address at least some of the technical problems existing in the prior art, the present invention provides a bone augmentation composite material that requires no exogenous bone material, no complex equipment, and can be prepared intraoperatively. Specifically, the present invention includes the following:

[0006] A first aspect of the present invention provides a method for expanding the volume of a composite matrix, comprising the step of contacting the composite matrix with a volume-expanding agent, thereby causing the volume-expanding agent to diffuse or penetrate into the interior of the composite matrix to expand its volume, wherein the volume-expanding agent is obtained from blood by centrifugation, the composite matrix comprises inorganic components, organic components, and cellular components, wherein the organic components include at least one of collagen, osteocalcin, osteopontin, proteoglycans, and glycoproteins, and the cellular components include at least one of osteocytes, osteoblasts, and osteoclasts.

[0007] In some embodiments, the method for causing volume expansion of a composite matrix according to the present invention further includes the steps of first contacting the composite matrix with physiological saline and then contacting it with a volume expansion agent, wherein the physiological saline does not cause or substantially does not cause volume expansion of the composite matrix.

[0008] In some embodiments, according to the method for causing the composite matrix to expand in volume according to the present invention, the inorganic component includes at least one of hydroxyapatite, calcium phosphate, calcium carbonate and magnesium phosphate.

[0009] In some embodiments, according to the method for expanding the volume of a composite matrix according to the present invention, the composite matrix is ​​in the form of granules or powder.

[0010] In some embodiments, according to the method for causing volume expansion of a composite matrix according to the present invention, the composite matrix and the volume expansion agent are derived from the same subject.

[0011] In some embodiments, according to the method for expanding the volume of a composite matrix according to the present invention, the volume expanding agent is liquid rather than gel or solidified.

[0012] In some embodiments, the method for expanding the volume of a composite matrix according to the present invention further includes a step of solidifying the composition to form a stable structure without adding blood or a coagulant.

[0013] In a second aspect, the present invention provides an osteogenic composition prepared by any of the methods described above.

[0014] A third aspect of the invention provides the use of the osteogenic composition in the preparation of medical materials for osteogenic purposes.

[0015] In some embodiments, according to the application described in the invention, the medical material is used for oral bone augmentation.

[0016] This invention utilizes a naturally derived volume-expanding agent to expand the composite matrix, thereby enabling the filling of larger bone defects with only a small amount of composite matrix. This is significant for expensive or scarce composite matrices, especially those derived from autologous bone. In some embodiments, this invention can achieve bone defect repair entirely using autologous materials, fully leveraging the advantages of autologous materials—no immune rejection and no risk of disease transmission—while simultaneously avoiding the limitation of insufficient autologous material to completely fill bone defects.

[0017] Furthermore, the osteogenic material provided by this invention possesses a dual osteogenic mechanism: the composite matrix provides a bone conduction scaffold, while the volume-expanding agent provides a bone-inducing signal. The osteogenic material obtained by this invention exhibits good stability; the composite matrix is ​​not easily lost after being encapsulated by the volume-expanding agent, conforming well to the defect morphology, and is cost-effective: no expensive bone powder or specialized equipment is required. The osteogenic material of this invention is easy to use, requiring only a conventional centrifuge and sterile containers, making it suitable for outpatient surgery. Attached Figure Description

[0018] Figure 1 The preparation of the liquid volume expander is shown.

[0019] Figure 2 The method for obtaining the composite matrix is ​​illustrated.

[0020] Figure 3 and 4 An electron micrograph of the osteogenic composition according to an embodiment of the present invention is shown.

[0021] Figure 5 An electron microscope image of Comparative Example 1 is shown.

[0022] Figure 6 This illustrates the use of the same tubular structure to obtain the same volume of composite material.

[0023] Figure 7 This illustrates the use of the same tubular structure to obtain equal amounts of composite base material.

[0024] Figure 8 The diagram shows the volume changes after equal amounts of composite material were immersed in physiological saline and liquid volume expander and left to stand (physiological saline on the left, liquid volume expander on the right).

[0025] Figure 9 The diagram shows the volume changes of equal volumes of composite matrix and artificial bone powder after immersion in a liquid volume-expanding agent and standing (autologous bone on the left, artificial bone on the right). Detailed Implementation

[0026] Various exemplary embodiments of the present invention are now described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, characteristics, and implementations of the present invention. Where specific techniques or conditions are not specified in the embodiments, they shall be performed in accordance with the techniques or conditions described in the literature in the art (e.g., refer to J. Sambrook et al., *Molecular Cloning: A Laboratory Manual*, 3rd edition, Science Press, translated by Huang Peitang et al.) or according to the product instructions. Reagents or instruments whose manufacturers are not specified are all commercially available conventional products.

[0027] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, the numerical ranges in this invention should be understood to specifically disclose the upper and lower limits of the range and every intermediate value between them. Every smaller range between any stated value or intermediate value within a stated range, and any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0028] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0029] Methods for causing composite matrix to expand in volume One aspect of the present invention provides a method for expanding the volume of a composite matrix, comprising the step of contacting the composite matrix with a volume-expanding agent, thereby causing the volume-expanding agent to diffuse or penetrate into the interior of the composite matrix to expand its volume, wherein the volume-expanding agent is obtained from blood by centrifugation, the composite matrix comprises inorganic components, organic components, and cellular components, wherein the organic components include at least one of type I collagen, osteocalcin, osteopontin, proteoglycans, and glycoproteins, the cellular components include at least one of osteocytes, osteoblasts, and osteoclasts, and the cellular components optionally include at least one of mesenchymal stem cells, hematopoietic cells, and vascular endothelial cells.

[0030] In some embodiments of the present invention, the composite base material may optionally contain trace amounts of lipids.

[0031] In a preferred embodiment, the method causes the composition to coagulate into a stable structure without the addition of blood or a coagulant.

[0032] In a preferred embodiment, the method further includes the step of first contacting the composite matrix with physiological saline and then with a volume-expanding agent, wherein the physiological saline does not cause or substantially does not cause volume expansion of the composite matrix.

[0033] In one specific implementation, the composite matrix is ​​isolated autologous bone that has not undergone any treatment (e.g., removal of organic components, physical or chemical treatment such as high temperature or high pressure). The size of the autologous bone is not particularly limited and can be the size of a soybean or particles or powder of any suitable size or particle size, as long as it has excellent operability.

[0034] In this invention, the volume expansion refers to the increase in volume of the composite base material after the addition of the volume expansion agent compared to before treatment. Preferably, the volume increases by at least two times compared to before treatment.

[0035] In this invention, the volume expansion agent can be prepared in-house or purchased from commercially available products. Preferably, the volume expansion agent is obtained by centrifuging blood to obtain specific liquid-phase components. In a preferred embodiment, a variable-speed centrifugation program is used to obtain liquid-phase components from the same subject, wherein the liquid-phase components are located at the red-yellow boundary between the erythrocyte layer and the pale yellow serum.

[0036] In this invention, the variable-speed centrifugation program is as follows: acceleration for 25-35 s (e.g., 28-32 s, such as 28, 29, 30, 31, 32 s), centrifugation at 2500-3000 rpm, preferably 2600-2800 rpm for 1-5 min, preferably 1-3 min, followed by centrifugation at 2000-2500 rpm, preferably 2300-2500 rpm for 1-5 min, preferably 3-5 min, followed by centrifugation at 2500-3000 rpm, preferably 2600-2800 rpm for 1-5 min, preferably 1-3 min, and finally deceleration for 30-40 s (e.g., 34-38 s, such as 34, 35, 36, 37, 38 s) to obtain liquid stratified components.

[0037] In this invention, contact between the composite matrix and the volume-expanding agent can be achieved through mixing and soaking or direct dropwise addition. In some embodiments, after contacting the composite matrix with the liquid volume-expanding agent, it is preferable to mix thoroughly and then allow it to stand for 1-5 minutes to allow the liquid volume-expanding agent to be fully adsorbed onto the surface and pores of the bone fragments. In another embodiment, the liquid volume-expanding agent is dropwise added to the composite matrix, and the resulting gel is further mixed evenly.

[0038] In some embodiments, the composite matrix comprises 60-70 parts by weight of inorganic components (e.g., but not limited to minerals), 20-30 parts by weight of organic components (e.g., but not limited to matrix), and 15-25 parts by weight of water. In some embodiments, the composite matrix comprises 2%-10% (typically 2%-5%) of cellular components based on the volume of the composite matrix.

[0039] Osteogenic composition In one aspect, the present invention provides an osteogenic composition prepared by the above method.

[0040] In some embodiments, the pharmaceutical composition further includes a pharmaceutically acceptable carrier or excipient. In this invention, "acceptable" means compatible with other components of the formulation and does not harm the patient.

[0041] Pharmaceutically acceptable carriers or excipients are non-toxic to recipients at the doses and concentrations used, and may include at least one of buffers, antioxidants, preservatives, isotonic agents, stabilizers, and surfactants.

[0042] application In one aspect, the invention provides the use of osteogenic compositions in the preparation of medical materials for osteogenic purposes.

[0043] Osteogenesis can be achieved by administering a therapeutically effective amount of the osteogenic composition to a subject in need. As used herein, the term "effective amount" means the amount of a drug or agent that elicits a biological or pharmaceutical response in a tissue, system, animal, or human, as sought by, for example, an investigator or clinician. Furthermore, the term "therapeuticly effective amount" means the amount that causes improved treatment, cure, prevention, or reduction of disease, condition, or side effects, or reduces the rate of progression of the disease or condition, compared to a corresponding subject who did not receive that amount. The term also includes, within its scope, amounts that effectively enhance normal physiological function. Generally, the effective amount as used herein varies depending on various factors, such as the given drug or composition, pharmaceutical formulation, route of administration, type of disease or condition, subject being treated, etc., but can still be routinely determined by those skilled in the art.

[0044] As used in this invention, the term "subject" refers to any animal (such as a mammal), including but not limited to humans, non-human primates, rodents, and the like who are about to receive a specific treatment. Generally, "subject" and "patient" are used interchangeably in this invention, both referring to the subject of the study.

[0045] The term "treatment" as used in this invention refers to improvement of the condition following a bone defect. The degree of relief or improvement of the bone defect, measured by any standard technique, such as new bone formation, bone volume, bone mineral density, or bone increment, is at least 5%, 10%, 20%, 40%, 50%, 60%, 80%, 90%, 95%, or 100%, compared to an untreated control group under equivalent conditions.

[0046] In a preferred embodiment, the medical material can be used in any scenario suitable for bone defect repair and bone volume reconstruction, such as, but not limited to, bone defect repair and bone volume reconstruction in fields such as dental implants, maxillofacial surgery, and orthopedics.

[0047] In a preferred embodiment, the medical material is used for oral bone augmentation, such as for the repair of vertical and horizontal bone defects in the alveolar ridge.

[0048] In this invention, the specific type of medical material is not particularly limited, and it can be a gel, patch, implant, stent, etc.

[0049] Example The patient's own bone fragments, obtained by grinding or scraping during surgery, are used as the composite matrix for the osteogenic composition. This composite matrix is ​​then thoroughly mixed with a volume-expanding agent extracted from venous blood, utilizing its adhesiveness and sustained-release growth factor properties to form a malleable, highly bioactive bone-increasing composite material.

[0050] The specific preparation steps are as follows: Collect venous blood: Draw 5-20 mL of venous blood from the patient before the operation and inject it into a test tube without anticoagulant; Preparation of liquid volume expander by centrifugation: Use a variable speed centrifugation program (e.g., accelerate 0.5 min → 2500-3000 rpm, e.g. 2700 rpm × 1-5 min, e.g. 2 min → 2000-2500 rpm, e.g. 2400 rpm × 1-5 min, e.g. 4 min → 2500-3000 rpm, e.g. 2700 rpm × 1-5 min, e.g. 2 min → decelerate, e.g. less than 1 min, e.g. 30-40 seconds) to obtain layered blood; Extraction of liquid swelling agent: Approximately 0.5-2 mL of the liquid layer above the red blood cell layer and below the pale yellow serum is extracted as the liquid swelling agent. Figure 1 ); Obtaining autologous bone fragments: Fresh autologous bone fragments generated during the procedure are collected using tools such as bone drills and curettes and used as a composite material. Figure 2 The composite matrix includes inorganic components, organic components, and cellular components. The organic components include collagen, osteocalcin, osteopontin, proteoglycans, and glycoproteins, while the cellular components include osteocytes, osteoblasts, and osteoclasts. Mixing and soaking or dripping treatment: Add the above composite base material to the liquid volume expansion agent and let it stand for 1-5 minutes to allow the liquid volume expansion agent to be fully adsorbed on the surface and pores of the bone chips. Alternatively, drip the liquid volume expansion agent onto the composite base material and mix the resulting gel evenly.

[0051] Implantation: The composite material is filled into the bone defect area and can be used in conjunction with a barrier membrane without the need for additional coagulation steps.

[0052] Comparative Example 1 Unlike the examples, this comparative example uses commercially available bone meal as the base material, which has been prepared by processing such as removing organic components. The commercially available bone meal is mixed with a liquid volume expander, and the resulting composition is compared with the composition obtained in the examples.

[0053] Comparative Example 2 Unlike the examples, the reagent used in this comparative example mixing, soaking, or dripping treatment is physiological saline, not a volume-expanding agent, and the resulting composition is compared with the composition obtained in the examples.

[0054] Comparative Example 3 This comparative example uses commercially available artificial bone powder, a volume expander, and blood from the patient's surgical area to obtain an osteogenic composition, and compares the resulting composition with the composition obtained in the examples.

[0055] Comparative Example 4 This comparative example uses only a composite material, namely autologous bone fragments, without using a liquid volume expander. On the one hand, this requires a large amount of autologous bone fragments, significantly increasing the amount needed and making it less practical, limiting its use to small defects. On the other hand, the lack of a high-concentration growth factor sustained-release system provided by a liquid volume expander results in slow bone formation and low volume maintenance in large defects and poorly vascularized areas (such as the maxillary sinus floor).

[0056] Comparative Example 5 This comparative study used only liquid volume expanders, without using composite materials, i.e., autologous bone fragments. The results showed that the liquid volume expanders could solidify into blocks, i.e., they could slowly form a weak gel after standing, but they lacked a bone scaffold structure and therefore could not support the three-dimensional growth of new bone. They were only suitable for soft tissue filling and did not meet the core purpose of "bone augmentation".

[0057] Comparative Example 6 To further verify the effect of the liquid volume expander, this comparative example used another formulation containing growth factors. Unlike the examples, the liquid volume expander was replaced with the following components: Control group 1: Gel membrane / block-like platelet-rich fibrin, a second-generation platelet concentrate product, which is composed of autologous leukocytes and platelet-rich fibrin biomaterials; Control group 2: Platelet-rich plasma, which is a platelet concentrate prepared from autologous whole blood by centrifugation. The platelet concentration is 4-8 times that of normal blood, and it contains multiple growth factors such as PDGF and TGF-β. Preparation methods include density gradient centrifugation, plasma separation and replacement, and blood component separator collection. Control group 3: Liquid platelet-rich fibrin, a liquid platelet-rich preparation.

[0058] Example of effect Figure 3 and 4 The following are electron micrographs of the composite matrix of this application after the addition of liquid volume expander, viewed from different angles. Figure 5Electron micrographs of commercially available bone powder of Comparative Example 1 after the addition of a liquid volume expander are shown. It can be observed that, with differences in the composition and structure of the composite matrix, the porous structure of the composite matrix of this application is more conducive to the entry of the liquid volume expander (e.g., cells contained therein), thereby promoting the formation of an osteogenic structure. Simultaneously, the ordered layered structure enables efficient penetration and loading, allowing for the formation of a stable structure through the slow solidification of the liquid volume expander itself, without the need for the addition of surgical site blood or other coagulants.

[0059] Figure 6 This illustrates the use of the same tubular structure to extract equal volumes of bone. Figure 7 This illustrates the use of the same tubular structure to take equal amounts of bone. Figure 8 The volume changes of equal amounts of autologous bone chips after immersion in physiological saline and liquid volume-expanding agent and left to stand are shown. Figure 9 The volume changes of equal volumes of autologous bone fragments and artificial bone powder after immersion in a liquid volume-expanding agent and standing are shown. It can be observed that when the autologous bone fragments, used as the composite matrix, are immersed in physiological saline, their volume remains unchanged compared to their initial volume outside the body. However, when mixed with the liquid volume-expanding agent, the volume expansion rate exceeds 2 times. This is of great significance for practical clinical applications. Currently, the limited amount of bone that can be harvested and the difficulty in obtaining it undoubtedly restrict the application of autologous bone. This is one of the reasons why artificial bone powder is increasingly being used in the field. The osteogenic composition obtained by the method of this invention can significantly reduce the amount of bone harvested, achieving bone increase with less autologous bone. Meanwhile, the composition of Comparative Example 1 cannot obtain the naturally occurring osteogenic signals (such as BMP-2, OCN, etc.) in autologous bone.

[0060] Furthermore, compared to Comparative Example 3, this invention can obtain a coagulated osteogenic composition in almost the same time without adding blood from the patient's surgical area, while the volume of Comparative Example 3 remains almost unchanged. In Comparative Example 6, because Control Group 1 is a gel film / block, it cannot uniformly soak bone chips in a "liquid" form, resulting in low loading efficiency. Control Group 2 requires extrinsic thrombin activation, has rapid growth factor release (<1 day), poor sustained-release properties, and contains anticoagulant residues, which actually inhibits osteogenic formation. Control Group 3 is a liquid platelet-rich preparation obtained by low-speed centrifugation, whose growth factor concentration is lower than that of a liquid volume expander, resulting in a significantly lower osteogenic effect compared to the liquid volume expander.

[0061] Although the invention has been described with reference to exemplary embodiments, it should be understood that the invention is not limited to the disclosed exemplary embodiments. Various adjustments or changes may be made to the exemplary embodiments described in this specification without departing from the scope or spirit of the invention. The scope of the claims should be interpreted in the broadest possible sense to cover all modifications and equivalent structures and functions.

Claims

1. A method for causing volume expansion of a composite base material, characterized in that, The method includes the step of contacting the composite matrix with a volume-expanding agent, thereby causing the volume-expanding agent to diffuse or penetrate into the interior of the composite matrix and expand its volume. The volume-expanding agent is obtained from blood by centrifugation. The composite matrix includes inorganic components, organic components, and cellular components. The organic components include at least one of collagen, osteocalcin, osteopontin, proteoglycans, and glycoproteins. The cellular components include at least one of osteocytes, osteoblasts, and osteoclasts.

2. The method for causing volume expansion of a composite matrix according to claim 1, characterized in that, The method further includes the step of first contacting the composite base material with physiological saline, and then contacting it with a volume-expanding agent, wherein the physiological saline does not cause or substantially does not cause volume expansion of the composite base material.

3. The method for causing volume expansion of a composite matrix according to claim 1, characterized in that, The inorganic components include at least one of hydroxyapatite, calcium phosphate, calcium carbonate, and magnesium phosphate.

4. The method for causing volume expansion of a composite matrix according to claim 1, characterized in that, The composite base material is in granular or powder form.

5. The method for causing volume expansion of a composite matrix according to claim 1, characterized in that, The composite base material and the volume expansion agent were derived from the same subject.

6. The method for causing volume expansion of a composite matrix according to claim 1, characterized in that, The volume expander is liquid, not gel or solid.

7. The method for causing volume expansion of a composite matrix according to claim 1, characterized in that, The method further includes a step of coagulating the composition to form a stable structure without adding blood or a coagulant.

8. An osteogenic composition, characterized in that, It is prepared by the method described in any one of claims 1-7.

9. The use of the osteogenic composition according to claim 8 in the preparation of medical materials for osteogenic purposes.

10. The application according to claim 9, characterized in that, The medical material is used for oral bone augmentation.

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