Composition for regenerating bone tissue or cartilage tissue comprising hydrogel

Abstract

The present invention relates to a composition or hydrogel for preventing or treating osteoarticular diseases. The composition or hydrogel according to one aspect exhibits excellent tissue regeneration effects that simultaneously induce structural and functional recovery of bone tissue and cartilage tissue, promotes bone regeneration in local bone defects caused by trauma or diseases, and can also be usefully applied to the treatment of traumatic osteoarticular diseases.

Description

Composition for regenerating bone tissue or cartilage tissue containing a hydrogel

[0001] The present invention relates to a composition for regenerating bone tissue or cartilage tissue comprising a hydrogel.

[0002] Bone tissue can be easily damaged by various causes, such as spinal injury, fractures, infections, and tumors. While natural healing is possible in cases like simple fractures, sufficient tissue repair through natural healing alone is difficult if the bone defect exceeds a certain extent. The healing rate of bone tissue slows down depending on the size of the defect, the presence of inflammation, and local circulatory disorders. If appropriate treatment is not provided, it can lead to long-term functional impairments such as delayed union, nonunion, and malunion. Furthermore, problems frequently arise where alveolar bone loss occurs during tooth extraction due to periodontal disease or trauma, making prosthetic and implant procedures difficult.

[0003] Meanwhile, cartilage tissue is an avascular tissue lacking blood vessels, nerves, and lymphatic vessels; consequently, its natural regenerative capacity is extremely low due to the very limited supply of cells following injury. Although some nearby chondrocytes or stem cells are supplied to the cartilage defect site, restoration to normal cartilage tissue is not effectively achieved due to insufficient migration distance and volume. Furthermore, damaged cartilage is vulnerable to mechanical pressure, making it susceptible to further injury and prone to expanding damage, thus highlighting the importance of early regenerative treatment.

[0004] Currently, clinical treatment for bone and cartilage tissue damage relies primarily on surgical procedures. For bone tissue damage and defects, invasive methods such as surgical reduction and internal fixation are used, but these are accompanied by various side effects including pain, infection, soft tissue damage, and delayed recovery. In cases of extensive bone defects or those accompanied by inflammation, it is difficult to expect sufficient tissue recovery through conventional surgery alone. For cartilage tissue, procedures such as artificial joint replacement, arthrochondroplasty, and microfracture are applied; however, these methods have limitations, including scarring from incisions, the burden of the procedure, and incomplete regeneration into less durable fibrocartilage rather than normal hyaline cartilage.

[0005] As such, existing treatments for bone and cartilage damage have various problems, such as invasiveness, limited tissue regeneration, and the possibility of re-injury; therefore, there is a need for the development of new non-invasive treatment strategies that can restore tissue structure and function more safely and effectively.

[0006] Accordingly, the inventors developed a hydrogel composition for the regeneration of bone tissue or cartilage tissue comprising fibrinogen, laminin, and hyaluronic acid, and applied it to damaged lesions in bone and joint areas, and confirmed an excellent tissue regeneration effect, thereby proving that the composition can be usefully utilized for the prevention or treatment of damage to bone tissue and cartilage tissue.

[0007] One aspect provides a composition for regenerating bone tissue or cartilage tissue comprising fibrin and / or fibrinogen; laminin or a laminin-derived peptide or protein; and / or hyaluronic acid or a salt thereof as active ingredients.

[0008] Another aspect provides a composition for regenerating bone tissue or cartilage tissue comprising, as an active ingredient, a hydrogel or hydrogel patch comprising fibrin and / or fibrinogen; laminin or a laminin-derived peptide or protein; and / or hyaluronic acid or a salt thereof.

[0009] Another aspect provides a topical composition for the regeneration of bone tissue or cartilage tissue comprising fibrin and / or fibrinogen; laminin or a laminin-derived peptide or protein; and / or hyaluronic acid or a salt thereof as active ingredients.

[0010] Another aspect provides a topical composition for the regeneration of bone tissue or cartilage tissue comprising, as an active ingredient, a hydrogel or hydrogel patch comprising fibrin and / or fibrinogen; laminin or a laminin-derived peptide or protein; and / or hyaluronic acid or a salt thereof.

[0011] Another aspect provides a pharmaceutical composition for the prevention or treatment of bone or cartilage diseases comprising fibrin and / or fibrinogen; laminin or a laminin-derived peptide or protein; and / or hyaluronic acid or a salt thereof as active ingredients.

[0012] Another aspect provides a pharmaceutical composition for the prevention or treatment of bone or cartilage diseases comprising, as an active ingredient, a hydrogel or hydrogel patch comprising fibrin and / or fibrinogen; laminin or a laminin-derived peptide or protein; and / or hyaluronic acid or a salt thereof.

[0013] Another aspect provides a method for preventing or treating bone disease or cartilage disease, comprising the step of administering a hydrogel or hydrogel patch containing fibrin or fibrin and fibrinogen; laminin or a laminin-derived peptide or protein; and hyaluronic acid or a salt thereof to an individual in need thereof.

[0014] Another aspect is to provide a hydrogel or hydrogel patch comprising fibrin or fibrin and fibrinogen; laminin or laminin-derived peptide or protein; and hyaluronic acid or its salt for use in the manufacture of medicines for the prevention or treatment of bone disease or cartilage disease.

[0015] Another aspect is to provide a hydrogel or hydrogel patch for the treatment or prevention of bone disease or cartilage disease comprising fibrin or fibrin and fibrinogen; laminin or laminin-derived peptide or protein; and hyaluronic acid or its salt.

[0016] One aspect provides a composition for regenerating bone tissue or cartilage tissue comprising fibrin and / or fibrinogen; laminin or a laminin-derived peptide or protein; and / or hyaluronic acid or a salt thereof as active ingredients.

[0017] Another aspect provides a composition for regenerating bone tissue or cartilage tissue comprising, as an active ingredient, a hydrogel or hydrogel patch comprising fibrin and / or fibrinogen; laminin or a laminin-derived peptide or protein; and / or hyaluronic acid or a salt thereof.

[0018] Another aspect provides a topical composition for the regeneration of bone tissue or cartilage tissue comprising fibrin and / or fibrinogen; laminin or a laminin-derived peptide or protein; and / or hyaluronic acid or a salt thereof as active ingredients.

[0019] Another aspect provides a topical composition for the regeneration of bone tissue or cartilage tissue comprising, as an active ingredient, a hydrogel or hydrogel patch comprising fibrin and / or fibrinogen; laminin or a laminin-derived peptide or protein; and / or hyaluronic acid or a salt thereof.

[0020] Another aspect provides a quasi-drug composition for the regeneration of bone tissue or cartilage tissue comprising fibrin and / or fibrinogen; laminin or a laminin-derived peptide or protein; and / or hyaluronic acid or a salt thereof as active ingredients.

[0021] Another aspect provides a quasi-drug composition for the regeneration of bone tissue or cartilage tissue comprising, as an active ingredient, a hydrogel or hydrogel patch comprising fibrin and / or fibrinogen; laminin or a laminin-derived peptide or protein; and / or hyaluronic acid or a salt thereof.

[0022] Another aspect provides a pharmaceutical composition for the prevention or treatment of bone or cartilage diseases comprising fibrin and / or fibrinogen; laminin or a laminin-derived peptide or protein; and / or hyaluronic acid or a salt thereof as active ingredients.

[0023] Another aspect provides a pharmaceutical composition for the prevention or treatment of bone or cartilage diseases comprising, as an active ingredient, a hydrogel or hydrogel patch comprising fibrin and / or fibrinogen; laminin or a laminin-derived peptide or protein; and / or hyaluronic acid or a salt thereof.

[0024] Another aspect provides a method for preventing or treating osteoarthritis, comprising the step of administering the above-mentioned hydrogel, hydrogel patch, or composition to an individual.

[0025] Another aspect provides the use of the above hydrogel, hydrogel patch, or composition in the manufacture of a formulation for the prevention or treatment of osteoarthritis.

[0026] Another aspect provides the use of the above hydrogel, hydrogel patch, or composition for the prevention or treatment of osteoarthritis.

[0027] The terms of the present specification, hydrogel, hydrogel patch, or composition may comprise fibrin and / or fibrinogen; laminin or laminin-derived peptide or protein; and / or hyaluronic acid or its salt. For example, the hydrogel, hydrogel patch, or composition may comprise fibrin or fibrinogen; laminin or laminin-derived peptide or protein; and hyaluronic acid or its salt.

[0028] As used herein, the term “osseous tissue” refers to tissue distributed throughout the body of humans or animals that performs functions such as structural support of bones, organ protection, weight bearing, and maintenance of the microenvironment. Osteous tissue is composed of compact bone and trabecular bone and includes osteocytes, osteoblasts, osteoclasts, and an extracellular matrix based on inorganic and organic substances. Osteous tissue may be damaged or defective due to various factors such as trauma, infection, inflammation, tumors, and tooth extraction; in particular, if the extent of damage or defect to the periosteum exceeds a certain level, sufficient recovery cannot be achieved through natural healing alone. Specifically, alveolar bone, as part of the jawbone, functions to support teeth and is included in general osseous tissue in terms of its histological composition and biological characteristics. Alveolar bone defects can also occur due to periodontitis, infection, trauma, or bone resorption following tooth extraction, and such defects require bone regeneration, just like other bone tissue defects. Therefore, the bone tissue defined in this specification includes all types of bone tissue, including alveolar bone. Specifically, the bone tissue encompasses various forms of bone tissue constituting the skeleton, such as long bone, compact bone, spongy bone, and alveolar bone, and includes long bone areas damaged by fracture, bone tissue affected around joints during dislocation, and alveolar bone areas where defects have occurred due to periodontitis, abscesses, etc.

[0029] The term "cartilage tissue" as used in this specification refers to connective tissue present in various parts of the human or animal body, such as the ends of bones, joint surfaces, the thorax, bronchi, nasal cavity, external ear, and intervertebral discs, which provides structural support, elasticity, and shock absorption functions for joints. Cartilage tissue is an avascular tissue composed primarily of an extracellular matrix consisting of collagen fibers and proteoglycans, and is composed of chondroblasts and chondrocytes; it possesses the biological characteristic of having very limited natural healing and cell regeneration capabilities after injury.

[0030] The above cartilage tissues include articular cartilage on the joint surface, fibrocartilage constituting the intervertebral disc, costal cartilage, bronchial cartilage, elastic cartilage of the nasal cavity and external ear, and hyaline cartilage and subchondral structures around the joint that are damaged due to trauma or dislocation are also included in the cartilage tissues to which the present invention applies.

[0031] The term "regeneration" as used in this specification generally refers to the process in which, when a living organism loses a part of its body or its function, it attempts to reconstruct the tissue or organ of that part to restore it to its original state or recover its function. This regenerative ability is stronger in organisms that have simpler systems and a lower degree of phylogenetic evolution.

[0032] The term "bone disease" in this specification refers to any bone tissue-related disease requiring damage, loss, or regeneration of bone tissue. Specifically, the bone disease may include conditions in which bone defects, osteolysis, microfractures, fractures, or structural deformations occur in bone tissue due to various causes such as trauma, inflammation, infection, tumors, metabolic disorders, and periodontal disease. In particular, long bone, spongy bone, and compact bone, as well as alveolar bone supporting teeth, are types of bone tissue that can easily be damaged by tooth extraction, infection, abscesses, etc., and are therefore included in the bone diseases defined in this specification.

[0033] In one embodiment, the bone disease may be one or more selected from the group consisting of fracture, bone defect, osteolysis, alveolar bone loss / defect, osteomyelitis, delayed union or non-union, traumatic bone injury, and degenerative bone conditions, but is not limited thereto.

[0034] The term “cartilage disease” in this specification refers to any cartilage-related pathological condition requiring damage, loss, degeneration, or structural or functional regeneration of cartilage tissue. Such cartilage disease may include conditions in which articular cartilage or the subchondral region is damaged due to trauma, degenerative changes, inflammation, infection, or mechanical load, resulting in cartilage defects, chondromalacia, full- or partial-thickness injury, or cartilage degeneration. Additionally, cartilage lesions resulting from damage to surrounding joint structures during dislocation or ligament injury are also included in the cartilage disease defined in this specification.

[0035] In one embodiment, the cartilage disease may be one or more selected from the group consisting of cartilage injury, cartilage defect, chondromalacia, and traumatic joint injury.

[0036] In this specification, the term "pharmaceutical effective dose" may mean any amount of a composition used in the course of practicing the invention provided herein that is sufficient for the alleviation, inhibition of progression, prevention, or treatment of a disease, disorder, or pathological condition, or one or more of its symptoms. The level of said effective dose may be determined by factors including the patient's health condition, the type and severity of the disease, sensitivity to the drug, the method of administration, the route of administration, and other factors well known in the medical field.

[0037] The terms “administering,” “applying,” “introducing,” and “implanting” are used interchangeably and may mean the placement of a patch or composition according to one embodiment into an object by a method or route resulting in at least partial localization of the patch or composition according to one embodiment to a desired site.

[0038] In this specification, the term "patch" may refer to a means having a certain shape that can be applied, attached, or contacted to a target area.

[0039] In one embodiment, the hydrogel, hydrogel patch, or composition may have properties intermediate between solid and liquid. The hydrogel, hydrogel patch, or composition may be amorphous, spherical, hemispherical, disc-shaped, or cylindrical. Additionally, for example, the diameter of the hydrogel patch may be 0.05 mm to 10 cm, 0.1 mm to 5 cm, 0.1 mm to 3 cm, or 0.2 mm to 1.5 cm, and may be provided in such a size or shape. Additionally, the hydrogel patch may be applied, implanted, attached, or in contact with a target site (e.g., a site of tissue damage) and deformed to conform to the shape of the damaged site.

[0040] In another embodiment, the hydrogel, hydrogel patch, or composition may be solid (including powder), semi-solid, or liquid. Additionally, for example, the hydrogel, hydrogel patch, or composition may undergo a reversible phase transition (e.g., depending on a change in temperature) to a solid (including powder), semi-solid, or liquid state. Since the hydrogel patch may undergo a reversible phase transition depending on ambient conditions such as temperature conditions, the hydrogel patch according to one embodiment may be produced and provided in a solid (including powder) or liquid state, and then converted into a hydrogel patch before, during, or after administration to a target site for use. For example, the hydrogel patch may be provided in a sol state containing fibrinogen, laminin, and hyaluronic acid, and the hydrogel patch according to one embodiment may be manufactured and used by using a substance (e.g., thrombin) that allows the user to convert fibrinogen into fibrin. Accordingly, a hydrogel patch according to one embodiment may be provided in the form of a prodrug, for example, a solid (powder), liquid (sol), or semi-solid composition comprising fibrin and / or fibrinogen; laminin; and / or hyaluronic acid. A composition provided in the form of a prodrug may be manufactured or modified into a hydrogel patch in vivo to function. Additionally, a hydrogel patch according to one embodiment may further comprise thrombin, or thrombin may be provided together as a kit.

[0041] In another embodiment, the hydrogel, hydrogel patch, or composition may be porous. Specifically, the surface of the hydrogel patch may have porosity (micropores). Without being limited to any specific theory, a hydrogel patch according to one embodiment may enhance the interaction between active substances by having porosity.

[0042] In one embodiment, the hydrogel, hydrogel patch, or composition may comprise fibrin and / or fibrinogen. The final pharmacological substance acting in vivo may comprise fibrin, but fibrinogen may be used instead of fibrin in the form of a prodrug. Additionally, depending on the amount of substance that converts fibrin to fibrinogen, the hydrogel, hydrogel patch, or composition according to one embodiment may partially comprise fibrinogen or thrombin. Accordingly, the present specification may additionally provide a prodrug comprising fibrinogen; laminin; and / or hyaluronic acid. The above fibrin or fibrinogen may be included at a concentration of 0.1 to 50 mg / ml, 0.1 to 25 mg / ml, 0.5 to 25 mg / ml, 1 to 25 mg / ml, 1 to 20 mg / ml, 5 to 25 mg / ml, 0.5 to 15 mg / ml, 1 to 15 mg / ml, 3 to 15 mg / ml, 5 to 15 mg / ml, 7 to 12 mg / ml, 8 to 25 mg / ml, 12 to 25 mg / ml, 15 to 24 mg / ml, 18 to 24 mg / ml, or 18 to 22 mg / ml.

[0043] The above fibrinogen glycoprotein is a hexamer composed of soluble α, β, and γ subunits produced in hepatocytes of the liver. Fibrinogen reacts with the enzyme thrombin to undergo a phase transition from soluble to insoluble fibrin polymer fibers.

[0044] The above-mentioned thrombin enzyme is a serine protease that transforms soluble fibrinogen into insoluble fibrin. The above-mentioned thrombin can play a role in gelling a hydrogel in a sol state by converting fibrinogen into fibrin.

[0045] In this specification, the term "laminin" may refer to an extracellular matrix protein constituting the basal lamina, specifically a heterotrimeric protein composed of α, β, and γ subunits. Accordingly, the laminin may include not only the full-length laminin protein but also laminin-derived peptides or proteins. For example, the laminin may be laminin-1, laminin-2, laminin-3, laminin-4, laminin-5A, laminin-5B, laminin-6, laminin-7, laminin-8, laminin-9, laminin-10, laminin-11, laminin-12, laminin-14, or laminin-15. In addition, the laminin may be included at a concentration of 1 to 100 μg / ml, 2 to 80 μg / ml, 5 to 50 μg / ml, 5 to 25 μg / ml, 8 to 20 μg / ml, 8 to 15 μg / ml, 10 to 70 μg / ml, 20 to 50 μg / ml, or 20 to 40 μg / ml.

[0046] In this specification, the term “hyaluronic acid” refers to a natural polymer found in various tissues, such as joint fluid, cartilage, connective tissue, and skin, as one of the representative glycosaminoglycans present in the body.

[0047] The above hyaluronic acid is a disaccharide polysaccharide composed of glycosidic bonds having β-(1→4) and β-(1→3) changes between D-glucuronic acid and N-acetyl-D-glucosamine, and has various molecular weights depending on the length of the disaccharide bonds. In one embodiment, the molecular weight of the hyaluronic acid may be 5,000 to 20,000,000 Da. More specifically, the molecular weight of the hyaluronic acid is 0.5 to 4.0 x 10⁻⁶. 6 Da, 1.0 to 2.0 x 10 6Da, or 1.5 to 1.8 x 10⁻⁶ 6 Da may be present. The hyaluronic acid may be included at a concentration of 10 μg / ml to 10 mg / ml, 10 μg / ml to 5 mg / ml, 50 μg / ml to 5 mg / ml, 100 μg / ml to 3 mg / ml, 200 μg / ml to 1 mg / ml, 500 μg / ml to 5 mg / ml, 500 μg / ml to 8 mg / ml, 500 μg / ml to 4 mg / ml, 500 μg / ml to 3 mg / ml, 300 μg / ml to 3 mg / ml, 1 mg / ml to 8 mg / ml, 1 mg / ml to 5 mg / ml, or 1 mg / ml to 3 mg / ml.

[0048] In other embodiments, the hydrogel, hydrogel patch, or composition may further include a cell growth factor. The cell growth factor may be a neuronal growth factor, a vascular endothelial growth factor, a fibroblast growth factor, a bone morphogenetic protein, an epidermal growth factor, a hepatocyte growth factor, a transforming growth factor, or a combination thereof. More specifically, the growth factors may include placental growth factor, macrophage colony-stimulating factor, granulocyte macrophage colony-stimulating factor, neuropilin, fibroblast growth factor (FGF)-1, FGF-2 (bFGF), FGF-3, FGF-4, FGF-5, FGF-6, erythropoietin, BMP-2, BMP-4, BMP-7, BMP-9, TGF-beta, IGF-1, osteopontin, pleotropin, activin, endothelin-1, and combinations thereof. The above neuronal growth factor may include one or more selected from the group consisting of BDNF (Brain-derived neurotropic factor), GDNF (Glial cell derived neurotropic factor), CNTF (Ciliary neurotropic factor), bFGF (basic fibroblast growth factor), cAMP (cyclic adenocyne monophosphate), NT (Neurotropin), NT3 (Neurotropin-3), NT4 (Neurotropin-4), T3 (Triiodo-L-Thyronine), SHH (Sonic hedgehog), and PDGF (Platelet-derived growth factor). The above vascular endothelial growth factor may include vascular endothelial growth factor (VEGF)-A, VEGF-A, VEGF-B, VEGF-C, VEGF-D, or VEGF-E, but is not limited thereto.

[0049] In one embodiment, the cell growth factor is neuronal growth factor, vascular endothelial growth factor, fibroblast growth factor (FGF), bone morphogenetic protein (BMP), epidermal growth factor (EGF), hepatocyte growth factor (HGF), transforming growth factor (TGF), placental growth factor (PIGF), macrophage colony-stimulating factor (M-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), neuropilin, FGF-1, FGF-2, FGF-3, FGF-4, FGF-5, FGF-6, erythropoietin (EPO), BMP-2, BMP-4, BMP-7, BMP-9, TGF-β, IGF-1, osteopontin, pleiotrophin, activin, endothelin-1, BDNF, GDNF, It may be CNTF, cAMP, NT, NT-3, NT-4, T3, SHH, PDGF, VEGF-A, VEGF-B, VEGF-C, VEGF-D, VEGF-E, or a combination thereof.

[0050] The concentration of the cell growth factor included in the hydrogel, hydrogel patch, or composition may vary depending on the type of growth factor, but generally, it may be included at a concentration of 1 ng / ml to 1,000 ng / ml or 0.1 μM to 100 μM. The cell growth factor may play a role in enhancing the tissue damage recovery effect of a hydrogel patch or composition containing fibrin, laminin, and hyaluronic acid.

[0051] In other embodiments, the hydrogel, hydrogel patch, or composition may include or substantially not include collagen. Without being limited to any particular theory, the collagen may be included as a component of the composition or may not be substantially included, but in one aspect, not including it may be more advantageous than including it.

[0052] Additionally, the hydrogel, hydrogel patch, or composition may not substantially comprise cells. In this specification, "substantially does not comprise" means that collagen or cells are included to an extent that they do not affect the activity or pharmacological activity of the hydrogel, hydrogel patch, or composition, or are not included at all. The hydrogel, hydrogel patch, or composition according to one embodiment may be distinguished from cell therapies generally used for the regeneration of damaged tissue by not substantially comprising cells.

[0053] In one embodiment, the hydrogel, hydrogel patch, or composition may be composed of fibrin and / or fibrinogen; laminin or a laminin-derived peptide; and hyaluronic acid or a salt thereof. Additionally, one or more of the components mentioned herein may be additionally included. For example, the hydrogel, hydrogel patch, or composition may be composed of fibrin and / or fibrinogen; laminin or a laminin-derived peptide; hyaluronic acid or a salt thereof; optionally thrombin; optionally collagen; and optionally a cell growth factor.

[0054] The dosage of the above composition according to one embodiment may be 0.01 mg to 10,000 mg, 0.1 mg to 1,000 mg, 1 mg to 100 mg, 0.01 mg to 1,000 mg, 0.01 mg to 100 mg, 0.01 mg to 10 mg, or 0.01 mg to 1 mg. However, the dosage may be prescribed differently depending on factors such as the formulation method, administration method, patient's age, weight, gender, pathological condition, food, time of administration, route of administration, excretion rate, and response sensitivity, and a person skilled in the art may appropriately adjust the dosage by considering these factors. The number of administrations may be one or two or more within the range of clinically acceptable side effects, and regarding the administration site, it may be administered at one or two or more sites. For animals other than humans, the above dosage may be administered at the same dose per kg as for humans, or calculated based on the ratio of organ volumes (e.g., heart, etc.) between the target animal and humans (e.g., average value). Possible routes of administration may include oral, sublingual, parenteral (e.g., subcutaneous, intramuscular, intra-arterial, intraperitoneal, intradural, or intravenous), rectal, local (including transdermal), inhalation, and injection, or insertion of an implantable device or substance. Examples of animals targeted for treatment according to one embodiment include various animal species including humans, and specifically include mammals such as humans, monkeys, mice, rats, rabbits, sheep, cattle, dogs, horses, camels, pigs, sugar gliders, etc., as well as birds (avian species) such as parrots and reptilian species such as lizards.

[0055] A pharmaceutical composition according to one embodiment may include pharmaceutically acceptable carriers and / or additives. For example, it may include sterile water, physiological saline, a common buffer (phosphoric acid, citric acid, other organic acids, etc.), a stabilizer, a salt, an antioxidant (ascorbic acid, etc.), a surfactant, a suspending agent, an isotonic agent, or a preservative. For topical administration, it may also include combinations with organic materials such as biopolymers, inorganic materials such as hydroxyapatite, specifically collagen matrices, polylactic acid polymers or copolymers, polyethylene glycol polymers or copolymers, and chemical derivatives thereof.

[0056] A pharmaceutical composition according to one embodiment may appropriately include, if necessary depending on the method of administration or dosage form, a suspending agent, a solubilizing agent, a stabilizer, an isotonic agent, a preservative, an anti-adsorption agent, a surfactant, a diluent, an excipient, a pH adjuster, an analgesic agent, a buffer, a reducing agent, an antioxidant, etc. Pharmaceutically acceptable carriers and formulations suitable for the present invention, including those exemplified above, are described in detail in the literature [Remington's Pharmaceutical Sciences, 19th ed., 1995]. A pharmaceutical composition according to one embodiment may be prepared in a unit dose form or contained in a multi-dose container by formulating using a pharmaceutically acceptable carrier and / or excipient according to a method that can be easily carried out by a person skilled in the art to which the invention pertains. In this case, the dosage form may be in the form of a solution, suspension, or emulsion in an oil or aqueous medium, or in the form of a powder, granule, tablet, or capsule.

[0057] The above composition may be formulated into an oral or parenteral administration formulation. The oral administration formulation may be a granule, powder, liquid, tablet, capsule, dry syrup, or a combination thereof. The parenteral administration formulation may be an injection or a topical application.

[0058] The term “topical preparation for skin” in this specification refers to a formulation for local application by applying directly to or attaching to the skin. The topical preparation for skin may be a cream, gel, ointment, skin emulsifier, skin suspension, transdermal patch, drug-containing bandage, lotion, or a combination thereof. The topical preparation for skin may be appropriately formulated as needed with ingredients commonly used in topical preparations for skin such as cosmetics, pharmaceuticals, or quasi-pharmaceuticals, for example, aqueous ingredients, oily ingredients, powder ingredients, alcohols, moisturizers, thickeners, UV absorbers, whitening agents, preservatives, antioxidants, surfactants, fragrances, colorants, various skin nutrients, or combinations thereof. The above topical preparation may also appropriately incorporate metal chelating agents such as disodium edetate, trisodium edetate, sodium citrate, sodium polyphosphate, sodium metaphosphate, and gluconic acid; caffeine, tannin, bellapamil, licorice extract, glablidin, hot water extract of the fruit of *calin*; various herbal medicines; preparations such as tocopherol acetate, glycyrrhizic acid, tranexamic acid and its derivatives or salts; and sugars such as vitamin C, magnesium ascorbate phosphate, ascorbate glucoside, arbutin, kojic acid, glucose, fructose, and trehalose.

[0059] In addition, the above composition may be a quasi-drug composition. The term "quasi-drug" refers to an article that falls under one of the following categories: fibers, rubber products, or similar items used for the purpose of treating, alleviating, managing, or preventing diseases in humans or animals; items that have a weak effect on the human body or do not act directly on the human body and are not instruments or machines, or similar items; and preparations used for sterilization, insecticidal, and similar purposes for preventing infection. It excludes articles used for the purpose of diagnosing, treating, alleviating, managing, or preventing diseases in humans or animals that are not instruments, machines, or devices, and articles used for the purpose of exerting pharmacological effects on the structure and function of humans or animals that are not instruments, machines, or devices. It may also include topical skin preparations and personal hygiene products. When the above hydrogel, hydrogel patch, or composition is added to a quasi-drug composition for the purpose of regenerating bone tissue or cartilage tissue, the above hydrogel, hydrogel patch, or composition may be added as is or used together with other quasi-drug ingredients, or may be used appropriately according to conventional methods. The mixture of active ingredients can be appropriately determined according to the purpose of use (prevention, health, or therapeutic treatment).

[0060] Another aspect provides a method for preparing a hydrogel patch by adding thrombin to a composition in a sol state comprising fibrinogen; laminin or a laminin-derived peptide; and hyaluronic acid or a pharmaceutically acceptable salt thereof.

[0061] The above method may also further include the step of incorporating a cell growth factor into the composition in the sol state.

[0062] The above method may also include a step of gelling by adding thrombin, and then gelling by adding thrombin again. The gelling may be performed at 10 to 40°C for 5 minutes to 3 hours. The hydrogel patch may be produced in various shapes or sizes depending on the shape of the mold.

[0063] The above method may also include the step of low-temperature preservation or cryopreservation of the hydrogel patch in a solution at 4 to -210°C. The solution may include DMSO (Dimethyl sulfoxide), but any solution that does not substantially alter the chemical or physical properties of the hydrogel patch may be used. The form or activity of the hydrogel patch does not substantially change even when low-temperature preservation or cryopreservation.

[0064] The above hydrogel patch, fibrin and / or fibrinogen, laminin, hyaluronic acid, or cell growth factor are as described above.

[0065] Another aspect provides a method for low-temperature preservation or cryopreservation of the above hydrogel patch in a solution (e.g., DMSO) at 4 to -210°C.

[0066] The terms, methods, effects, etc. described for the above inventions apply equally between each invention.

[0067] A composition or hydrogel according to one aspect exhibits an excellent tissue regeneration effect that simultaneously induces structural and functional recovery of bone tissue and cartilage tissue, promotes bone regeneration in local bone defects caused by trauma or disease, and can be usefully applied to the treatment of traumatic bone and joint diseases.

[0068] Figure 1 is an image captured from X-ray images, local tissue conditions, and video footage of a clinical animal with persistent motor dysfunction and pain due to a hind limb fracture, taken before and after hydrogel administration, and the animal recovering to normal walking 13 days after hydrogel administration.

[0069] Figure 2 is a radiographic image of a clinical animal with a gait disorder due to cruciate ligament dislocation before hydrogel administration, a photograph showing the gait disorder, and a video captured of the animal recovering to normal gait after hydrogel administration.

[0070] Figure 3 is a radiographic image confirming the alveolar bone defect and bone dissolution site that existed before hydrogel administration, and a photograph showing the state of regeneration progressing through the reduction of the defect space and the recovery of alveolar bone density and contour 13 days after hydrogel administration.

[0071] The present invention will be explained in detail below through reference examples and embodiments. However, the following reference examples and embodiments are merely illustrative of the present invention and should not be construed as limiting the present invention.

[0072]

[0073] Example 1. Preparation of a hydrogel patch

[0074] 1.1. Hydrogel Formulation

[0075] Fibrinogen was dissolved in physiological saline or PBS at a sol state at 20 mg / ml, hyaluronic acid was dissolved in physiological saline or PBS at 5 mg / ml, and thrombin was dissolved in physiological saline or PBS at 200 U / ml. Subsequently, sol-state hydrogels were prepared by combining sol-state fibrinogen at 1 mg / ml, 5 mg / mL, or 10 mg / ml; laminin at 5 µg / ml, 10 µg / ml, 30 µg / ml, or 50 µg / ml; and hyaluronic acid at 0.1 mg / ml, 0.5 mg / ml, 1.0 mg / ml, or 1.5 mg / ml.

[0076] 1.2. Manufacture of Hydrogel Patch

[0077] Thrombin in a sol state was mixed with fibrinogen, laminin, and hyaluronic acid prepared in Example 1-1, and then dispensed into a sterile Parafilm or petridish. Subsequently, a hydrogel patch was prepared through a sol-gel phase transition by curing at room temperature or 37°C.

[0078]

[0079] Experimental Example 1. Confirmation of fracture healing effect of hydrogel

[0080] The tissue recovery and motor function improvement effects of the hydrogel according to one embodiment of the present invention were evaluated on clinical animals, prairie dogs, which suffer from motor dysfunction and persistent pain due to hind limb fractures.

[0081] Specifically, the subject prairie dog was unable to walk due to a fracture in the long bone region of its hind leg, and severe inflammation and edema were observed in the soft tissue surrounding the fracture site. In X-ray images prior to hydrogel administration, a fracture line was clearly observed, and findings accompanied by tissue damage and an inflammatory response around the fracture site were confirmed. 200 µl to 5 ml of hydrogel (410 µg / kg) prepared by the method of Example 1 was injected into the peri-fracture soft tissue, and the ability to walk, recovery of mobility, pain response, and tissue condition were observed for 13 days after administration. The evaluation results are shown in Table 1 and Figure 1.

[0082] Table 1 summarizes the degree of recovery of hind limb function by category after administration of hydrogel to clinical animals with persistent motor dysfunction and pain due to hind limb fracture, and Figure 1 is an image captured from video data showing the X-ray images, local tissue condition, and recovery to normal walking 13 days after administration of hydrogel to clinical animals with persistent motor dysfunction and pain due to hind limb fracture.

[0083] Evaluation Criteria Before Hydrogel Administration After 13 Days of Hydrogel Administration Pain Reduction Severe pain response upon palpation of the fracture site, inability to bear weight, persistent pain during walking Gradual decrease in signs of pain after administration Recovery of Walking Function Inability to use hind limbs, severe limping, dragging pattern during movement Normal walking and turning ability possible after 13 days Increased Mobility Limited movement distance, avoidance of movement, significantly reduced activity level Movement distance and activity level significantly increased compared to before administration Reduction in Tissue Inflammation Severe soft tissue swelling and redness around the fracture site, accompanied by localized heat Reduction in swelling and redness around the fracture site, confirmation of soft tissue recovery Imaging Improvement Clear fracture line on X-ray, fracture gap and displacement observed Findings of tissue stabilization and reduced inflammation observed at the fracture site

[0084] As shown in Table 1 and Figure 1, when the hydrogel of the present invention was applied, the motor function of the hind limbs of prairie dogs recovered to a normal level within 13 days after administration, and the animals, which were unable to walk at all before administration, showed active movement and natural walking patterns after administration. In addition, the inflammatory redness of the soft tissues observed immediately after injection decreased significantly over time, and X-ray images also confirmed that the condition of the tissues around the fracture stabilized. These results indicate that the hydrogel of the present invention effectively alleviates tissue damage and inflammation caused by fractures, while simultaneously promoting the recovery and regeneration of the missing bone tissue, thereby demonstrating a therapeutic effect that promotes functional recovery.

[0085]

[0086] Experimental Example 2. Confirmation of the dislocation treatment effect of hydrogel

[0087] The pain relief and walking function recovery effects of the hydrogel according to one embodiment of the present invention were evaluated for small dogs exhibiting walking difficulties due to dislocation of the hind limb cruciate ligament.

[0088] Specifically, the subject small dog complained of severe pain due to displacement of the cruciate ligament in the right hind leg, and was unable to bear weight on the affected leg, making daily walking difficult. Radiographic images prior to hydrogel administration confirmed clear displacement of the right cruciate ligament, and clinically, the dog exhibited limping and avoidance behavior while walking.

[0089] For the above small dogs, 200 µl to 5 ml of hydrogel (410 µg / kg) prepared by the method of Example 1 was injected into the injured area around the hind limb cruciate ligament, and walking ability, weight-bearing degree, pain response, and changes in activity were observed for 4 weeks after administration. The evaluation results are shown in Table 2 and Figure 2.

[0090] Table 2 summarizes the degree of clinical recovery by evaluation item after administering hydrogel to clinical animals exhibiting gait impairment due to cruciate ligament dislocation of the hind limb, and Figure 2 is a radiographic image of a clinical animal exhibiting gait impairment due to cruciate ligament dislocation of the hind limb, a photograph showing gait impairment, and a captured image from video footage showing recovery to normal gait after hydrogel administration.

[0091] Evaluation Criteria Before Hydrogel Administration After 4 Weeks of Hydrogel Administration Pain Reduction Clear pain response during walking, avoidance of limb use, significant tenderness Gradual decrease in pain response after administration, significantly alleviated at 4 weeks Recovery of Weight-bearing Inability to bear weight on the affected hind limb, walking with the foot lifted or severe limping Weight-bearing on the affected hind limb, which was impossible before administration, recovered to normal levels Improvement of Walking Function Distinct limping, inability to walk normally Limping disappeared, walking and running movements normalized Increased Activity Level Restricted activity during indoor / outdoor movement, reduced activity due to pain Improved activity during indoor / outdoor movement, disappearance of avoidance behavior

[0092] As shown in Table 2 and Figure 2, the application of the hydrogel of the present invention resulted in a significant improvement in gait impairment caused by anterior cruciate ligament dislocation. In images taken before administration, the affected hind leg was unable to be placed on the ground, but after 4 weeks following administration, a normal gait pattern was restored, and stable walking without limping became possible even during long-distance movement. Furthermore, veterinarian observations confirmed that "normal gait was restored to a level similar to that of a healthy dog." These results indicate that the hydrogel of the present invention stabilizes the joint structure destabilized by anterior cruciate ligament injury and promotes soft tissue stabilization and functional recovery by facilitating the recovery and regeneration of the cartilage tissue forming the joint surface and surrounding soft tissues.

[0093]

[0094] Experimental Example 3. Confirmation of Alveolar Bone Regeneration Effect of Hydrogel

[0095] In guinea pigs with alveolar bone defects formed due to moderate to severe alveolar bone inflammation and abscess, the hydrogel composition of the present invention was evaluated to determine whether it induces bone regeneration of the damaged alveolar bone.

[0096] Specifically, the subject guinea pig showed abscess formation around the alveolar bone, mucosal redness and edema, and soft tissue damage; in particular, a clear bone void with an empty interior was present in the mandibular alveolar bone area due to the abscess. Additionally, in the X-ray images taken before the application of hydrogel, irregular and increased penetrative osteolysis was observed in certain parts of the alveolar bone, along with a clearly observed void formed by the abscess.

[0097] The alveolar bone defect site of the subject guinea pig was first incised to drain pus and remove necrotic tissue, and then 100 µl to 3 ml of hydrogel (410 µg / kg) prepared by the method of Example 1 was directly injected into the defect space and the damaged alveolar bone area. Subsequently, the disappearance of inflammation, the degree of bone regeneration, and changes in the defect space were observed for 13 days, and the results are shown in Table 3 and Figure 3.

[0098] Table 3 compares the clinical changes and bone regeneration status by category before and 13 days after administration of hydrogel in guinea pigs with induced alveolar bone inflammation and abscess, and Figure 3 shows radiographic images of the alveolar bone defect and bone resorption site observed before administration of hydrogel, and a photograph showing the state of regeneration progressing through the reduction of the defect space and the recovery of alveolar bone density and contour 13 days after administration.

[0099] Evaluation Criteria Before Hydrogel Administration 13 Days After Hydrogel Administration Abscess Large amount of abscess and pus present in the alveolar bone area Abscess disappeared, no pus or exudate observed Inflammation Severe redness and swelling of the gingiva and mucosa Reduction in redness and swelling, disappearance of inflammation Mucosa and Soft Tissue Condition Necrotic tissue and irregular mucosa observed Mucosa surface stabilization, epithelialization progressing Oral Hygiene Status Foreign matter and severe odor around the lesion Less area tidied up, odor disappearance Overall Healing Persistent inflammation and tissue damage progressing Distinct tissue recovery, gingival condition near normal

[0100] As shown in Table 3 and Figure 3, prior to hydrogel administration, a hollow space was observed in the alveolar bone area due to abscess formation and tissue damage, and radiographic images also revealed an irregular alveolar bone contour and hypodense areas suspected of bone resorption. Gross examination also clearly showed mucosal redness, edema, abscesses, and necrotic tissue. On the other hand, 13 days after hydrogel administration, radiographic images showed changes suggesting alveolar bone recovery, such as a reduction in the existing hollow space and a denser contour of the surrounding bone tissue. Gross examination also confirmed that the initial abscess and redness had mostly disappeared, the soft tissues around the tooth root had stabilized, and the mucosal surface had recovered uniformly. These results indicate that when the hydrogel composition of the present invention is applied locally to an alveolar bone inflammation lesion, it reduces inflammatory exudation and abscesses and promotes substantial bone regeneration in the defective alveolar bone area.

[0101] Therefore, the composition of the present invention can be usefully applied as a therapeutic agent to induce alveolar bone regeneration and structural recovery in bone defect diseases.

Claims

1. Fibrin or, fibrin and fibrinogen; Laminin or, laminin-derived peptide or protein; and A composition for regenerating bone tissue or cartilage tissue, comprising hyaluronic acid or a salt thereof as an active ingredient.

2. In Claim 1, The above composition is a hydrogel or a hydrogel patch.

3. In Claim 1, A composition comprising the above fibrin or fibrinogen at a concentration of 0.1 to 50 mg / ml, the above laminin at a concentration of 1 to 100 μg / ml, or the above hyaluronic acid at a concentration of 10 μg / ml to 10 mg / ml.

4. In Claim 1, A composition comprising additionally thrombin.

5. In Claim 1, A composition comprising additional cell growth factors.

6. In Claim 5, The above cell growth factors are neuronal growth factor, vascular endothelial growth factor, fibroblast growth factor (FGF), bone morphogenetic protein (BMP), epidermal growth factor (EGF), hepatocyte growth factor (HGF), transforming growth factor (TGF), placental growth factor (PIGF), macrophage colony-stimulating factor (M-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), neuropilin, FGF-1, FGF-2, FGF-3, FGF-4, FGF-5, FGF-6, erythropoietin (EPO), BMP-2, BMP-4, BMP-7, BMP-9, TGF-β, IGF-1, osteopontin, pleiotrophin, activin, endothelin-1, BDNF, GDNF, CNTF, cAMP, A composition that is NT, NT-3, NT-4, T3, SHH, PDGF, VEGF-A, VEGF-B, VEGF-C, VEGF-D, VEGF-E, or a combination thereof.

7. In Claim 2, A composition in which the above hydrogel or hydrogel patch does not contain cells.

8. In Claim 2, A composition in which the above hydrogel or hydrogel patch has porosity on its surface.

9. In Claim 2, A composition in which the above hydrogel or hydrogel patch undergoes a reversible phase transition to a solid, semi-solid, or liquid state depending on temperature.

10. Fibrin or, fibrin and fibrinogen; Laminin or, laminin-derived peptide or protein; and A pharmaceutical composition for the prevention or treatment of bone or cartilage diseases, comprising hyaluronic acid or a salt thereof as an active ingredient.

11. In Claim 10, A pharmaceutical composition wherein the above bone disease is one or more selected from the group consisting of fracture, bone defect, osteolysis, alveolar bone loss / defect, osteomyelitis, delayed union or non-union, traumatic bone injury, and degenerative bone conditions.

12. In Claim 10, A pharmaceutical composition wherein the above cartilage disease is one or more selected from the group consisting of cartilage injury, cartilage defect, chondromalacia, and traumatic joint injury.

13. Fibrin or, fibrin and fibrinogen; Laminin or, laminin-derived peptide or protein; and A method for preventing or treating bone disease or cartilage disease, comprising the step of administering a hydrogel or hydrogel patch containing hyaluronic acid or a salt thereof to an individual in need thereof.

14. For use in the manufacture of medicines for the prevention or treatment of bone or cartilage diseases, Fibrin or, fibrin and fibrinogen; Laminin or, laminin-derived peptide or protein; and Use of a hydrogel or hydrogel patch containing hyaluronic acid or its salt.

15. For the prevention or treatment of bone or cartilage diseases, Fibrin or, fibrin and fibrinogen; Laminin or, laminin-derived peptide or protein; and Use of a hydrogel or hydrogel patch comprising hyaluronic acid or a salt thereof.

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