KR20240103830A - Injection-type hyaluronic acid hydrogel drug delivery system comprising vitamin D for the treatment of tendinopathy or muscle damage and manufacturing method thereof - Google Patents

Abstract

The present invention relates to a pharmaceutical composition for preventing or treating musculoskeletal diseases comprising hyaluronic acid or a salt thereof, a crosslinking agent, and vitamins, and a method for manufacturing the same. the composition of the present invention can induce sustained release of drugs and increase stability by dispersing fat-soluble vitamin d in hydrophilic hyaluronic acid hydrogel, and can be used as a complex agent for preventing and treating tendonopathy by solving cytotoxicity exhibited at high concentrations.

Description

Hyaluronic acid hydrogel drug delivery system comprising vitamin D for the treatment of tendinopathy or muscle damage and manufacturing method thereof {Injection-type hyaluronic acid hydrogel drug delivery system comprising vitamin D for the treatment of tendinopathy or muscle damage and manufacturing method thereof}

The present invention relates to a pharmaceutical composition for preventing or treating musculoskeletal diseases containing hyaluronic acid or a salt thereof, a cross-linking agent and vitamins, and a method for producing the same.

Tendinopathy is a disease caused by degenerative changes in collagen within the tendon tissue. When the tendon tissue is damaged due to tendinopathy, repetitive sprains occur or when the area is overused, recovery of the tendon tissue occurs. This will not come true. In addition, tendinopathy may cause abnormal alignment of collagen fibers within the tendon tissue, fibrinoid or myxoid degeneration of collagen, and fibrosis, and the weakened tendon tissue may rupture in severe cases.

Although this tendinopathy is a very common clinical disease, there is no clear treatment yet. Treatments for tendinopathy known to date include conservative treatments such as stretching, physical therapy, anti-inflammatory medication, steroid injections, extracorporeal shock wave therapy, and autologous platelet-rich plasma injections. For most of them, the scientific basis has not been clearly revealed. Meanwhile, extreme stages of tendinopathy can be resolved with surgical treatment, but the development of non-surgical treatments that can treat early or mid-stage tendinopathy is urgently needed.

Therefore, under this background, the inventors of the present invention have developed a technology for continuously releasing an appropriate amount of vitamin D in the body over a certain period of time and controlling the rate using a biocompatible, biodegradable, natural polymer-derived hydrogel, and utilizing this to produce vitamin D. An injectable hyaluronic acid hydrogel treatment containing D was developed.

One aspect provides a pharmaceutical composition for preventing or treating musculoskeletal diseases comprising hyaluronic acid or a salt thereof, a cross-linking agent, and vitamins.

Another aspect provides a method for producing a composition for preventing or treating musculoskeletal diseases comprising hyaluronic acid or a salt thereof, a cross-linking agent, and vitamins.

Another aspect provides a method for preventing or treating musculoskeletal disorders, comprising administering the composition to a subject other than a human.

One aspect is to provide a pharmaceutical composition for preventing or treating musculoskeletal diseases comprising hyaluronic acid or a salt thereof, a cross-linking agent, and vitamins.

The composition of the present invention can induce sustained release of the drug and increase its stability by dispersing fat-soluble vitamin D in the hydrophilic hyaluronic acid hydrogel, and solves the cytotoxicity that appears at high concentrations to treat musculoskeletal diseases including tendinopathy. It can be used as a treatment for prevention and treatment.

As used herein, the term "hyaluronic acid and its salts" is a linear polysaccharide with a negative charge and composed of a glycosaminoglycan structure, a polymer of disaccharides made of glucuronic acid and N-acetyl glucosamine. It means and his salt. Specifically, the hyaluronic acid of the present invention may have a crosslinking rate of 0.01 to 10% to form a hydrogel, but is not particularly limited thereto. The salt of hyaluronic acid may be any one or more selected from the group consisting of sodium hyaluronate, potassium hyaluronate, calcium hyaluronate, magnesium hyaluronate, zinc hyaluronate, cobalt hyaluronic acid, and tetrabutylammonium hyaluronic acid, but is specifically limited thereto. It doesn't work.

As used herein, the term "hydrogel" refers to a gel using water as a dispersion medium, which means that the hydrosol loses its fluidity due to cooling or the hydrophilic polymer with a three-dimensional network structure and microcrystalline structure contains water. It can be formed by expansion or expansion. Therefore, in the present invention, the hyaluronic acid-based hydrogel may be manufactured by introducing a crosslinkable group into hyaluronic acid and then crosslinking it through reaction with a crosslinking agent.

As used herein, the term “cross-linking agent” refers to a substance that connects specific atoms in a linear molecule, and may refer to a substance for cross-linking the hyaluronic acid. Specifically, the crosslinking agent is butanediol diglycidyl ether (1,4-butandiol diglycidyl ether, BDDE), ethylene glycol diglycidyl ether (EGDGE), and hexanediol diglycidyl ether. Diyl ether (1,6-hexanediol diglycidyl ether), propylene glycol diglycidyl ether, polypropylene glycol diglycidyl ether, polyterdramethylene ether Polytetramethylene glycol diglycidyl ether, neopentyl glycol diglycidyl ether, polyglycerol polyglycidyl ether, diglycerol polyglycerol ether ( diglycerol polyglycidyl ether), glycerol polyglycidyl ether, tri-methylpropane polyglycidyl ether, bisepoxypropoxyethylene (1,2-(bis(2,3-epoxypropoxy) ethylene), pentaerythritol polyglycidyl ether, and sorbitol polyglycidyl ether. More specifically, butanediol diglycidyl. It may be ether (BDDE), but is not particularly limited thereto.

In the present invention, the vitamin may be any one or more selected from the group consisting of vitamin A, vitamin B group, vitamin C, vitamin D group, vitamin E, vitamin K group, and vitamin P. Specifically, the vitamin is vitamin D, more specifically, any one selected from the group consisting of cholecalciferol, calcifediol, calcitriol, ergocalciferol, and combinations thereof. It may be more than one, and more specifically, it may be cholecalciferol. Additionally, the vitamin may be concentrated cholecalciferol powder with a titer of 90,000 IU/g to 120,000 IU/g, which may be included in 0.01 to 10% by weight of the hydrogel, but is not particularly limited thereto.

In one example, 1mM or 2mM of vitamin D3 (cholecalciferol) was dispersed in a PBS solution containing 0.4% Tween 80 and added to the hyaluronic acid hydrogel cross-linked with 20 mg/ml BDDE in a ratio of 50:50. A hyaluronic acid-based hydrogel containing 1 mM vitamin D3 was prepared.

As used herein, the term “musculoskeletal disease” refers to pain or damage occurring in the musculoskeletal system, including muscles, nerves, tendons, ligaments, bones and surrounding tissues. Specifically, the musculoskeletal disease may be a tendon disease or a ligament disease, and more specifically, the tendon disease may include Achilles tendon disease, patellar tendon disease, lateral epicondylitis, medial epicondylitis, plantar fasciitis, rotator cuff tendon disease, tenosynovitis, and tendonitis. , tenosynovitis, tendon injury, tendon detachment, and tendinopathy, and the ligament disease is cruciate ligament injury, ankle ligament injury, collateral ligament injury, quadriceps femoris ligament rupture, coronary ligament sprain, ligament rupture, and It may be any one or more selected from the group consisting of sprains. More specifically, the musculoskeletal disease may be tendinopathy, but is not particularly limited thereto.

In addition, the pharmaceutical composition for preventing or treating musculoskeletal diseases may further include a surfactant. The surfactant is sorbitan monolaurate (Tween 20), polyoxyethylene sorbitan monopalmitate (Tween 40), polyoxyethylene sorbitan monostearate (Tween 60), and polyoxyethylene sorbitan monooleate (Tween 80), and polyoxyethylene sorbitan trioleate (Tween 85) and their derivatives. Specifically, it may be polyoxyethylene sorbitan monooleate (Tween 80). , but is not particularly limited thereto.

As used herein, the term “prevention” refers to any action that suppresses or delays musculoskeletal diseases by administering the pharmaceutical composition.

As used herein, the term “treatment” refers to any action that improves or beneficially changes the symptoms of a musculoskeletal disease by administering the pharmaceutical composition.

The pharmaceutical composition may further include a pharmaceutically acceptable carrier. As used herein, the term “pharmaceutically acceptable carrier” refers to a carrier or diluent that does not irritate the subject being administered and does not inhibit the biological activity and properties of the pharmaceutical composition. Acceptable pharmaceutical carriers in compositions formulated as liquid solutions include those that are sterile and biocompatible, such as saline solution, sterile water, Ringer's solution, buffered saline solution, albumin injection solution, dextrose solution, maltodextrin solution, glycerol, ethanol, and One or more of these ingredients can be mixed and used, and other common additives such as antioxidants, buffers, and bacteriostatic agents can be added as needed. In addition, diluents, dispersants, surfactants, binders, and lubricants can be additionally added to formulate injectable formulations such as aqueous solutions, suspensions, emulsions, etc., pills, capsules, granules, or tablets.

The pharmaceutical composition may be in various oral or parenteral dosage forms. When formulated, it can be prepared using diluents or excipients such as commonly used fillers, extenders, binders, wetting agents, disintegrants, and surfactants. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, etc. These solid preparations contain one or more compounds and at least one excipient, such as starch, calcium carbonate, sucrose, or lactose ( It is prepared by mixing lactose, gelatin, etc. Additionally, in addition to simple excipients, lubricants such as magnesium stearate, talc, etc. are also used. Liquid preparations for oral administration include suspensions, oral solutions, emulsions, and syrups. In addition to the commonly used simple diluents such as water and liquid paraffin, various excipients such as wetting agents, sweeteners, fragrances, and preservatives may be included. there is. Preparations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Non-aqueous solvents and suspensions may include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable esters such as ethyl oleate.

The pharmaceutical composition is selected from the group consisting of tablets, pills, powders, granules, capsules, suspensions, oral solutions, emulsions, syrups, sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, suppositories and injections. It can have any one dosage form, and more specifically, it can be an injection.

Another aspect is to provide a method for producing a composition for preventing or treating musculoskeletal disorders comprising hyaluronic acid or a salt thereof, a cross-linking agent, and vitamins. The same parts as described above also apply to the manufacturing method.

Specifically, the manufacturing method includes preparing a dispersion by adding vitamins to a solution containing a surfactant; Preparing a cross-linked hyaluronic acid hydrogel by reacting hyaluronic acid or a salt thereof with a cross-linking agent; And it may include the step of mixing the cross-linked hyaluronic acid hydrogel and the dispersion.

Another aspect is to provide a method for preventing or treating musculoskeletal diseases, comprising administering a composition containing hyaluronic acid or a salt thereof, a cross-linking agent, and vitamins to an entity other than a human. The same parts as described above also apply to the method of preventing or treating musculoskeletal diseases.

The subject may be a mammal other than a human that has or is likely to develop a musculoskeletal disease, and may specifically be a mammal such as a cow, horse, sheep, pig, goat, camel, antelope, dog, or cat. , but is not particularly limited thereto.

The administration route or method of administration of the pharmaceutical composition of the present invention may be intravenous injection, intramuscular or ligament injection, subcutaneous injection, intradermal injection, intratracheal injection, and topical skin application, and more specifically, intramuscular or ligament injection, fiber- It may be an injection into the bone junction, muscle-ligament junction, etc. The formulation of the pharmaceutical composition according to an embodiment of the present invention is not limited thereto, and may be formulated in various forms according to any administration route and method as long as it can reach the target disease area.

The composition of the present invention induces sustained release of the drug and increases stability by dispersing fat-soluble vitamin D in the hydrophilic hyaluronic acid hydrogel, and prevents and treats tendinopathy by solving cytotoxicity that appears at high concentrations. It can be used as a combination preparation for:

Figure 1 is a schematic diagram of injecting an injectable hyaluronic acid hydrogel treatment containing vitamin D into the affected area for the purpose of improving and treating tendinopathy.
Figure 2 shows the results of measuring complex viscosity and tanδ in the frequency range of 0.02 to 1 Hz using a rheometer to analyze the rheological properties of the hydrogel.
Figure 3 shows the results of verifying the amount of the drug through HPLC for 150 days at 4°C to test the stability of the therapeutic drug in the hydrogel when stored for a long period of time.
Figure 4 shows the results of verifying the amount of the drug through HPLC for 150 days at 25°C to test the stability of the therapeutic drug in the hydrogel when stored for a long period of time.

This will be described in more detail through examples below. However, these examples are for illustrative purposes only and the scope of the present invention is not limited to these examples.

Example 1. Preparation of hyaluronic acid hydrogel treatment containing vitamin D3

A hydrogel-type tendinopathy treatment containing vitamin D3 was prepared. Specifically, 2mM of vitamin D3 was dispersed in a PBS solution containing 0.4% Tween 80, and this was added in a 50:50 ratio to a hyaluronic acid hydrogel cross-linked with 20 mg/ml BDDE to obtain 1 or 2mM of vitamin D3. A hydrogel-type therapeutic agent containing

Example 2. Preparation of gelatin hydrogel therapeutic agent containing vitamin A

Vitamin A was used as the supported drug to induce dispersion through Span 80, and a hydrogel-type therapeutic agent was prepared in the same manner as Example 1 above, except that a gelatin-derived hydrogel was used.

Example 3. Preparation of collagen hydrogel therapeutic agent containing vitamin C

Dispersion was induced through F68 using vitamin C as the supported drug, and a hydrogel-type therapeutic agent was prepared in the same manner as Example 1, except that collagen-derived hydrogel was used.

Example 4. Preparation of alginic acid hydrogel therapeutic agent containing vitamin B2

Dispersion was induced through F127 using vitamin B2 as the supported drug, and a hydrogel-type therapeutic agent was prepared in the same manner as Example 1, except that an alginic acid-derived hydrogel was used.

Example 5. Preparation of chitosan hydrogel therapeutic agent containing vitamin K

Vitamin K was used as the supported drug to induce dispersion through Tween 20, and a hydrogel-type therapeutic agent was prepared in the same manner as Example 1 above, except that chitosan-derived hydrogel was used.

Comparative Example 1. Preparation of hyaluronic acid hydrogel injection

A hyaluronic acid hydrogel injection containing 5% cross-linked 20 mg/ml hyaluronic acid was prepared.

Comparative Example 2. Manufacturing of vitamin D injection

A saline-based injection containing 1 mM vitamin D3 was prepared.

Comparative Example 3. Preparation of synthetic polymer injection

A saline-based injection containing 0.4% by weight of Tween 80 was prepared.

Experimental Example 1. Rheological characterization of hydrogel

In order to analyze the rheological properties of the hydrogel of Example 1, complex viscosity and tan δ were measured in a frequency range of 0.02 to 1 Hz using a rheometer. As a result, as shown in Figure 2, the storage modulus (G') was higher than the loss modulus (G"), so it was confirmed that all formulations existed in hydrogel form.

Experimental Example 2. Stability test of therapeutic drug

To test the stability of the therapeutic drug in the hydrogel when stored for a long period of time, the amount of the drug was verified through high-performance liquid chromatography (HPLC) for 150 days at 4℃ and 25℃, respectively. HPLC measured the UV length at 265 nm for 15 minutes with a composition ratio of 70% acetonitrile, 25% methanol, and 5% water. As a result, as shown in Figures 3 and 4, it was confirmed that vitamin D3 was stable for about 150 days when the therapeutic drug was stored at 4°C.

Experimental Example 3. Evaluation of biocompatibility and anti-inflammatory activity

Human tenocytes were cultured in a 24 well-plate (8,000 per well). After damaging the cells with TNF-α, one day later, injectable hyaluronic acid hydrogel cross-linked with vitamin D according to the conditions was injected into the trans-well. After culturing for 3 days, cell survival and death analysis was performed.

division polymer drug Surfactants biocompatibility Anti-inflammatory effect Example 1 Hyaluronic acid Vitamin D3 Tween 80 O O Example 2 gelatin Vitamin A Span 80 △ X Example 3 Collagen Vitamin C F68 O △ Example 4 alginic acid Vitamin B2 F127 △ X Example 5 Chitosan Vitamin K Tween 20 △ X Comparative Example 1 Hyaluronic acid - - X X Comparative Example 2 - Vitamin D3 - X X Comparative Example 3 - - Tween 80 X X

As a result, as shown in Table 1, Comparative Examples 1 to 3 did not show biocompatibility and anti-inflammatory effects at all, and Examples 2, 4, and 5 containing vitamins A, B2, and K did not show any anti-inflammatory effects at all. Meanwhile, in the case of Example 3 containing vitamin C, it was judged to be biocompatible, but the recovery effect on tendon cell inflammation induced by TNF-α was found to be minimal. On the other hand, in the case of Example 1 containing vitamin D3, it was confirmed that both biocompatibility and anti-inflammatory effects were excellent.

The description of the present invention described above is for illustrative purposes, and those skilled in the art will understand that the present invention can be easily modified into other specific forms without changing the technical idea or essential features of the present invention. will be. Therefore, the embodiments described above should be understood in all respects as illustrative and not restrictive.

Claims (13)

A pharmaceutical composition for preventing or treating musculoskeletal diseases, comprising hyaluronic acid or a salt thereof, a cross-linking agent, and vitamins.
The composition of claim 1, wherein the musculoskeletal disease is a tendon disease or a ligament disease.
The method of claim 2, wherein the tendon disease consists of Achilles tendon disease, patellar tendon disease, lateral epicondylitis, medial epicondylitis, plantar fasciitis, rotator cuff tendon disease, tenosynovitis, tendonitis, tenosynovitis, tendon damage, tendon dissection, and tendinopathy. One or more selected from the group,
The composition, wherein the ligament disease is at least one selected from the group consisting of cruciate ligament injury, ankle ligament injury, collateral ligament injury, quadriceps femoris ligament rupture, coronary ligament sprain, ligament rupture and sprain.
The composition according to claim 1, wherein the musculoskeletal disease is tendinopathy.
The method of claim 1, wherein the crosslinking agent is butanediol diglycidyl ether (1,4-butandiol diglycidyl ether, BDDE), ethylene glycol diglycidyl ether (EGDGE), and hexanediol. Diglycidyl ether (1,6-hexanediol diglycidyl ether), propylene glycol diglycidyl ether, polypropylene glycol diglycidyl ether, polyether Polytetramethylene glycol diglycidyl ether, neopentyl glycol diglycidyl ether, polyglycerol polyglycidyl ether, diglycerol polygly Serol ether (diglycerol polyglycidyl ether), glycerol polyglycidyl ether, tri-methylpropane polyglycidyl ether, bisepoxypropoxyethylene (1,2-(bis(2,3) A composition that is at least one selected from the group consisting of -epoxypropoxy)ethylene), pentaerythritol polyglycidyl ether, and sorbitol polyglycidyl ether.
The method of claim 1, wherein the salt of hyaluronic acid is at least one selected from the group consisting of sodium hyaluronate, potassium hyaluronate, calcium hyaluronate, magnesium hyaluronate, zinc hyaluronate, cobalt hyaluronate, and tetrabutylammonium hyaluronic acid. A composition.
The composition of claim 1, wherein the vitamin is vitamin D3.
The composition of claim 1, wherein the composition further comprises a surfactant.
The method of claim 8, wherein the surfactant is sorbitan monolaurate (Tween 20), polyoxyethylene sorbitan monopalmitate (Tween 40), polyoxyethylene sorbitan monostearate (Tween 60), and polyoxyethylene sorbitan. A composition comprising at least one selected from the group consisting of carbon monooleate (Tween 80), polyoxyethylene sorbitan trioleate (Tween 85), and derivatives thereof.
A method for preventing or treating musculoskeletal diseases, comprising administering the composition of any one of claims 1 to 9 to an entity other than a human. (a) preparing a dispersion by adding vitamins to a solution containing a surfactant;
(b) preparing a cross-linked hyaluronic acid hydrogel by reacting hyaluronic acid or a salt thereof with a cross-linking agent;
(c) A method for producing a composition for preventing or treating musculoskeletal disorders, comprising mixing the cross-linked hyaluronic acid hydrogel and the dispersion.
The manufacturing method according to claim 11, wherein the musculoskeletal disease is tendinopathy.
The production method according to claim 11, wherein the vitamin is vitamin D3.