KR20160029957A - Pharmaceutical Composition for Temporomandibular Joint Disorders comprising Stromal Vascular Fraction - Google Patents

Abstract

The present invention relates to a pharmaceutical composition for preventing or treating temporomandibular disorders, comprising stem cells. In addition, the present invention relates to a kit for treating the temporomandibular disorders, comprising the composition. The stem cells according to the present invention effectively alleviate inflammation, and can comprehensively treat complex symptoms of the temporomandibular disorders, thereby being able to be used as a novel agent for preventing or treating the temporomandibular disorders which are classified as an incurable disease which does not have a fundamental treating method.

Description

[0001] The present invention relates to a pharmaceutical composition for preventing or treating temporomandibular joint disorders including stem cells,

The present invention relates to a pharmaceutical composition for preventing or treating temporomandibular joint disorders comprising stem cells.

The present invention also relates to a kit for treating temporomandibular joint disorders comprising the above composition.

The jaw joint is just before both ears and connects the jaw to the head bone, and plays a very important role in performing the function of the jaw such as chewing or talking. The jaw joint acts as the central axis of all jaw movements and is supported by the jaw muscles and ligaments, in which the disk between the jaw joints serves as a cushion between the bones and the bones. These muscles, ligaments, discs, and jawbone are combined to perform complex activities. The occurrence of abnormalities in the temporomandibular joint function by various factors is referred to as temporomandibular joint disorders (TMJ) or TMJ.

The temporomandibular joint disorder that develops in the jaw joint starts from simple arthritis at first, and gradually destroys the joint structure due to chronic inflammation if not treated appropriately. These symptoms are alleviated and worsened repeatedly, resulting in obstacles to swallowing, chewing, and vocalization, which are inherent features of the jaw, resulting in incongruity of the facial skeleton, which degrades the quality of life of the patient.

Especially, patients with severe jaw joint disorders have severe jaw joint and head and neck pain, limitation of opening, difficulty in chewing, facial deformity, and must be reconstructed through surgery. However, many patients do not receive appropriate treatment due to technical or economic reasons. to be.

The number of patients with any of the symptoms of jaw joint disorders (arthralgia, joint noise, or open mouth) is estimated to be 40-50% of the whole population. In particular, the number of patients who require treatment at a professional medical institution reaches 5% of the total population, and a large number of patients with diabetes mellitus population exist in Korea. Among them, 5% of the patients with severe jaw joint disorders that require surgery, and also the number of days with advanced disease related to stress is increasing trend.

Recently, the development of regenerative medicine and tissue engineering technology has been actively investigated for the improvement of inflammation and tissue regeneration using stem cells, and various attempts have been made to apply it to clinical applications. There have been many studies to prevent the progression of jaw joint disorders However, there is no way to treat and prevent the inflammation that has occurred other than the symptomatic therapy from the characteristic change, and it is considered to be a typical incurable disease.

Accordingly, the present inventors have made intensive efforts to develop a therapeutic agent for a disorder of the jaw joint classified as an intractable disease, and as a result, they have found that the stem cell is very effective for disorder of the jaw joint and have completed the present invention.

Application number: 1020097018749

One aspect of the present invention is to provide a pharmaceutical composition for preventing or treating temporomandibular joint disorder comprising stem cells.

Further, one aspect of the present invention is to provide a kit for treating a jaw joint disorder comprising the composition.

One aspect of the present invention is to provide a pharmaceutical composition for preventing or treating temporomandibular joint disorder comprising stem cells.

Hereinafter, the present invention will be described in detail.

In the present invention, the jaw joint comprises a musculoskeletal system such as a lower jaw bone, a head bone, a disc (disc), ligament, and surrounding muscles therebetween, and a jaw joint disorder refers to a defect in the jaw joint of the anterior portion of the ear. Temporomandibular joint disorders are often referred to as "temporomandibular disorders" and are often referred to as "temporomandibular disorders" or "cranial mandibular disorders", often involving head and neck pain disorders.

Temporomandibular Joint Disorders (TMJ) is a term that includes several clinical problems of the masticatory and temporomandibular joints.

The most common symptom of temporomandibular joint pain is pain, which usually occurs in the masticatory muscles, the anterior part of the ear, or the jaw joint. This is characterized by chewing chewing food or other jaw function. Common jaw joint disorders patients complain of chin, ear, head or facial pain, limitation of jaw movement, asymmetrical jaw shape, various noise (clicking, jaw joint) ), Gratings, crepitus, etc.) can occur. In addition, bad oral habits such as irritation, non-painful masticatory hypertrophy and abnormal mowing (teeth wear) can also occur. In other words, temporomandibular joint disorders are mainly composed of disorders of the jaw joint itself and head and neck muscular disorders (including tension headache).

Symptoms of jaw joint disorders are very diverse and can be caused by multiple causes. In other words, unconscious habits such as genetic predisposition, past trauma, irregularity of bone, malnutrition, changes in endocrine factors, irregularity, irritability, bite and jaw tightness, posture, sleeping habits, occupational habits, It can be attributed to social factors such as midnight, occupation, financial, and friends as well as behavioral factors such as level, smoking, drinking, drug overuse and managerial use. Especially, emotional, It is a disease greatly affected by emotional disorders.

Accordingly, the jaw joint disorder of the present invention is a concept including both jaw joint inflammation, temporomandibular joint pain, head and neck pain, limitation of opening, difficulty in chewing and facial deformity.

In the present invention, a stem cell is a precursor cell having an ability to self-replicate and to differentiate into various tissues by a suitable signal, and forms organs of the human body from the developmental stage and functions of organs and tissues after growth It plays an important role in restoration. Stem cells are embryonic stem cells obtained from early blastocysts and adult stem cells obtained from embryonic adult or placenta

Adult stem cells are transplanted in vivo and have the specificity to differentiate according to their long-term characteristics and the flexibility to cross-differentiate into different kinds of cells from their original cell characteristics, and the potential for differentiation into various cells have.

The above may preferably be a stem cell fraction. Stromal Vascular Fraction (SVF) is a vascular matrix that connects the gaps of cells in a tissue. It refers to a group of cells in the connective tissue that act as a signaling and nutrient supply between cells and cells.

The stem cells may be autologous or other stem cells.

The stem cells may be derived from tissues such as fat, uterus, bone marrow, muscle, placenta, umbilical cord blood, hair follicle or skin, preferably stem cells derived from adipose tissue .

The adipose tissue-derived stem cells can be derived from, and are not limited to, adipose tissue isolated from the subcutaneous, testicular, mesentery, peritoneal, facial, thigh, or calf.

In particular, in the best embodiment of the present invention, the stem cells may be autologous fat-derived stem cells.

In addition, the stem cell fraction can be obtained by collagenase treatment and centrifugation on adipose tissue isolated from a living body.

The stem cells include not only adipocytes but also preadipocytes, mesenchymal stem cells (MSC), endothelial progenitor cells, T cells, B cells, mast cells, macrophages, and cells Growth-related cytokines, and the like.

The stem cells may include 1 x 10 ² to 1 x 10 ¹⁰ cells / ml of mesenchymal stem cells.

The stem cells according to the present invention can effectively treat inflammation effectively and can treat complex symptoms of jaw joint disorders comprehensively and thus can be used as a new preventive or therapeutic agent for jaw joint disorders which are separated into incurable diseases because there is no fundamental treatment method .

The term "prevention" of the present invention means all actions that inhibit or delay the onset of the jaw joint disorder by the administration of the pharmaceutical composition according to the present invention, and "treatment" Means any act that improves or improves the symptoms caused by the disability.

The term "pharmaceutical composition" of the present invention can be used in combination with a "cell therapy composition ", and it is intended to encompass a pharmaceutical composition which is used for treatment, diagnosis and prevention with cells and tissues prepared by isolation, The US FDA regulation) is intended to treat, diagnose, and treat patients through a series of actions, such as the extracorporeal selection of live, homologous, or xenogeneic cells to alter the function of a cell or tissue, Means drugs used for prevention purposes.

The cell treatment composition of the present invention may further comprise a pharmaceutically acceptable carrier. The term "pharmaceutically acceptable" as used herein refers to a substance which is not toxic to cells or humans exposed to the composition. The carrier can be used without limitation as long as it is known in the art such as a buffer, a preservative, an anhydrous agent, a solubilizer, an isotonic agent, a stabilizer, a base, an excipient, a lubricant and a preservative. The pharmaceutical compositions of the present invention may be prepared according to techniques commonly used in the form of various formulations. The cell therapeutic agent, which is a composition of the present invention, can be administered through any route as long as it can induce migration to a disease site. In some cases, it may be possible to consider loading into a vehicle with means for directing the stem cells to the lesion.

Thus, the compositions of the present invention can be administered topically (including buccal, sublingual, skin and intravenous), parenterally (subcutaneously, intradermally, intramuscularly, intradermally, intravenously, intraarterially, intraarticularly and intraperitoneally) , And is preferably administered parenterally, most preferably directly at the site of onset. In one embodiment, the stem cells can be administered to a subject by suspension in a suitable diluent at a concentration of about 1 × 10 ² to 1 × 10 ¹⁰ cells / ml, which protects and maintains the cells, It is used for easy use. The diluent may be physiological saline, phosphate buffer solution, buffer solution such as HBSS, plasma, cerebrospinal fluid, or blood component. In addition, the pharmaceutical composition can be administered by any device so that the active substance can migrate to the target cell. Preferred modes of administration and formulations are injections. The injectable solution may be an aqueous solvent such as a physiological saline solution, a ring gel solution, a Hank solution or a sterilized aqueous solution, a vegetable oil such as an olive oil, a higher fatty acid ester such as ethyl oleic acid and a non-aqueous solution such as ethanol, benzyl alcohol, propylene glycol, polyethylene glycol or glycerin A non-penetrating agent known in the art suitable for the barrier to be passed may be used for mucosal permeation, and a stabilizer for preventing deterioration may include ascorbic acid, sodium hydrogen sulfite, BHA, tocopherol, EDTA And a pharmaceutical carrier such as an emulsifier, a buffer for controlling pH, mercury nitrate, a preservative for inhibiting the growth of microorganisms such as thimerosal, benzalkonium chloride, phenol, cresol and benzyl alcohol.

In another embodiment, the present invention provides a method for preventing or treating temporomandibular joint disorder by administering to a mammal a pharmaceutical composition for preventing or treating the temporomandibular joint disorder, comprising administering the composition to a subject in need thereof can do.

The stem cells may be treated before or after washing.

The term "individual" of the present invention means all animals including humans that have developed or may develop jaw joint disorders as described above, and the diseases can be effectively prevented or treated by administering the pharmaceutical composition of the present invention to a subject. The pharmaceutical composition of the present invention can be administered in parallel with existing therapeutic agents.

The term "administering" of the present invention means introducing a predetermined substance into a patient in an appropriate manner, and the administration route of the composition can be administered through any conventional route as long as it can reach the target tissue. But are not limited to, intraperitoneal, intravenous, intramuscular, subcutaneous, intradermal, oral, topical, intranasal, intrathecal, rectal.

In addition, the pharmaceutical composition of the present invention may be administered by any device capable of moving the active substance to the target cell. The preferred modes of administration and formulations are intravenous, subcutaneous, intradermal, intramuscular, and drip injections. The injectable solution may be a non-aqueous solvent such as an aqueous solvent such as a physiological saline solution or a ring gel solution, a vegetable oil, a higher fatty acid ester (e.g., oleic acid), an alcohol (e.g., ethanol, benzyl alcohol, propylene glycol, glycerin, etc.) (For example, ascorbic acid, sodium hydrogen sulfite, sodium pyrophosphate, BHA, tocopherol, EDTA and the like), an emulsifier, a buffer for pH control, a microbial growth inhibitor And a pharmaceutical carrier such as a preservative (e.g., mercury nitrate, thimerosal, benzalkonium chloride, phenol, cresol, benzyl alcohol, etc.).

In addition, the present invention provides a kit for treating a jaw joint disorder comprising the composition.

The kit may take the form of a bottle, a tub, a sachet, an envelope, a tube, an ampoule, etc., which may be partly or wholly made of plastic, glass, paper, foil, / RTI > The container may be fitted with a cap which is initially part of the container or can be fully or partially detachable, which may be attached to the container by mechanical, adhesive, or other means. The container may also be equipped with a stopper, which is accessible to the contents by the injection needle. The kit may include an external package, and the external package may include instructions for use of the components.

The stem cells according to the present invention can effectively treat inflammation effectively and can treat complex symptoms of jaw joint disorders comprehensively and thus can be used as a new preventive or therapeutic agent for jaw joint disorders which are separated into incurable diseases because there is no fundamental treatment method .

FIG. 1 is a photograph showing injection of stem cells obtained in a mouse model of jaw arthritis inflammation.
2 shows the damaged disk before the stem cell treatment, and the right side of FIG. 2 shows the recovered disk after the stem cell fraction treatment.
3 shows the infiltration of inflammatory cells before the stem cell treatment, and the right side of FIG. 3 shows the recovery of inflammatory cell infiltration after the stem cell fraction treatment.
FIG. 4 is a chart showing that, in the case of a patient having lost the effusion, not only the disappearance of the inflammatory exudate (Grade 3? Grade 0) but also the regeneration of the condyle.
Figure 5 is a schematic diagram of a clinical trial design.

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

<Example 1> Examination of therapeutic effect in a jaw joint inflammation model

1-1. Obtaining and administering stem cells

The stem cells to be used in the experiment of the present invention were obtained by the following method.

Process number Process Name Contents One Getting the province The abdomen of the rat is opened to acquire adipose tissue. 2 Fatty tissue
relay The adipose tissue obtained in Step 1 is transferred to a sterilized syringe or a sterilized bottle and then transferred to an adipocyte fractionation chamber. 3 Tissue cleaning Transfer the adipose tissue to a 50 ml tube and wash the blood and other buffer by centrifugation by adding phosphorylation buffer solution at a ratio of 1: 1. 4 Enzyme treatment 0.075% collagenase in the weight of the washed fat tissue of step 3 is added, and the reaction is carried out at 37 degrees for 45 minutes. 5 wash Wash-out three times with phosphorylation buffer solution. 6 Obtain SVF The washed tissue of step 5 is centrifuged at 2,000 rpm for 10 minutes to obtain the SVF pellet. 7 percolation The pellet of step 6 removes impurities and RBC with a 70 um mesh filter. 8 SVF cleaning The SVF obtained in step 7 is washed three times with the final injection water. 9 Cell quantification The SVF of Step 8 is subjected to morphological analysis using an inverted microscope and the cell viability and the yield of stem cells are confirmed through an automated cell measuring device. 10 charge
label 1x10 ⁷ cells / cc in the water for injection so that it can be administered to the Rat of Step 9, and the solution is placed in a 1cc syringe. 11 SVF forwarding The final SVF procedure manager in step 10 is informed and confirmed receipt.

In addition, the stem cells obtained by the above method were administered to a mouse model of jaw joint inflammation. FIG. 1 is a photograph showing injection of stem cells obtained in a mouse model of jaw arthritis inflammation.

The stem cells obtained by the above method are injected in 0.2 cc of a 1 cc syringe in both sides of the Rat.

1-3. Surgical findings for temporomandibular joint inflammation model

Tissue inflammation model was sacrificed and tissues were dissected to confirm the relief of jaw arthritis.

The results are shown in Fig.

In Fig. 2, the left photograph shows the damaged disk before the stem cell treatment, and the right photograph of Fig. 2 shows the recovered disk after the stem cell treatment.

The left disc before the procedure is thick, cloudy and deformed due to inflammation, but the disc on the right side is thin and transparent after the procedure.

1-4. Confirmation of inflammatory cell infiltration recovery effect

In addition, it was confirmed whether inflammatory cell infiltration in the temporomandibular joint inflammation model was recovered after stem cell treatment.

After removing the temporomandibular joint discs and mandibular condyles of Rat, fix them with 4% paraformaldehyde and decolorize them with 15% EDTA. The specimens were immobilized on paraffin and cut into sagittal sections at intervals of 5 μm. Then, they were stained with H-E and observed with an electron microscope.

The results are shown in Fig.

3 shows the infiltration of inflammatory cells before the stem cell treatment, and the right side of FIG. 3 shows the recovery of inflammatory cell infiltration after the stem cell fraction treatment.

The left upper part of the image shows the proliferation of the synovial lining cells and the invasion of the mononucleated cells. However, the above picture shows that the above findings are decreased after the procedure. In addition, the lower left image shows that the blood vessels expand and develop due to inflammation, and sub-synovial lipid droplets are observed in the lower part of the synovial membrane. .

<Example 3> Clinical preliminary experiment for patients with jaw arthritis

We then performed preliminary clinical studies in three patients with refractory degenerative jaw arthritis who did not respond to conservative treatment.

As a result, in the preliminary clinical study, all three patients showed significant improvement of the opening disorder (32.6 ± 2.5 mm → 44.6 ± 2.3 mm) and chronic pain relief (VAS 6.7 → VAS 1.3).

On follow-up MRI findings, complete loss of inflammatory effusion (grade 3 → grade 0) in joint pain, significant reduction of inflammatory effusion (grade 3 → grade 1) in 2 patients, and slight decrease in 1 patient (Grade 3 → grade 2).

Particularly, as shown in Fig. 4, in the case of the patient having lost the effusion, not only the disappearance of inflammatory effusion (Grade 3? Grade 0) but also the regeneration of the condyle were observed.

Loss of the effusion or reconstruction of the condylar is a very encouraging phenomenon in the treatment of temporomandibular joint, which is not confirmed by the postoperative MRI findings of dexamethasone or hyaluronic acid injected after the conventional treatment of the jaw.

&Lt; Example 2 > Examination of therapeutic effect in patients with jaw arthritis

2-1. Clinical trial patient information

Because the incidence of female patients is overwhelmingly high and the frequency of progression to osteoarthritis of the jaw is high, female patients are selected first. The characteristics of clinical studies are chosen to select adult age groups , And women aged 19 years or older. Patients with less than normal opening (less than 40 mm), chronic arthralgia, and MRI findings were excluded because of the following: 1) patients with inflammatory exudates in the joint, 2) patients with displaced discs, 3) Patients who showed all three signs of degenerative changes on the surface were included.

In addition, patients with significant symptomatic improvement in normal standard therapy (physical therapy, device therapy, medication) except jaw joint cleansing, pregnant and lactating women, taking oral steroids for at least two weeks within the past month Patients who did not participate more than once in the planned follow-up after the operation of the temporomandibular joint rinse were enrolled in the study. Patients who had experienced, had other banned treatment, jaw joint ankylosis, anatomical structural abnormality, , 60 patients were excluded from the study, except those who were deemed unable to comply with the requirements of the trial and those who were deemed unsuitable for clinical trials. 30 were randomly assigned to the test group and 30 to the control group.

2-2. Design of clinical trials

The clinical trial was designed as shown in Fig.

Specifically, patients who did not show significant symptom improvement in the standard treatment methods (physiotherapy, device therapy, medication) except temporomandibular cleansing among patients with temporomandibular joint disorder were performed general cleansing . In the experimental group, autologous fat-derived stem cell transplantation was performed if the authors agree to autologous fat-derived stem cell transplantation and if the subject agrees in writing to participate in clinical trials.

In the experimental group, they were injected via intracapsular injection at a dose of 2 cc per stem (1 × 10 7 / cc of stem cells) per day, 24 hours after the procedure.

2-3. Acquisition of fat-derived stem cell fraction from patient

The adipose-derived stem cells isolated from the patients were obtained and treated as shown in Table 2 below.

Process number Process Name Contents One Getting the province Fat tissue is obtained through liposuction in the subject's abdomen, thighs, and oral tissues. A 1.5-2.0 mm incision is made and lidocaine, a local anesthetic, and fepretin, a vasoconstrictor, are administered, followed by inhalation of the fat using a cannula.
: Tumescent method 2 Fatty tissue
relay The adipose tissue obtained in Step 1 is transferred to a sterilized syringe or a sterilized bottle and then transferred to an adipocyte fractionation chamber. 3 Tissue cleaning Transfer the adipose tissue to a 50 ml tube and wash the blood and other buffer by centrifugation by adding phosphorylation buffer solution at a ratio of 1: 1. 4 Enzyme treatment 0.075% collagenase in the weight of the washed fat tissue of step 3 is added, and the reaction is carried out at 37 degrees for 45 minutes. 5 wash Wash-out three times with phosphorylation buffer solution. 6 Obtain SVF The washed tissue of step 5 is centrifuged at 2,000 rpm for 10 minutes to obtain the SVF pellet. 7 percolation The pellet of step 6 removes impurities and RBC with a 70 um mesh filter. 8 SVF cleaning The SVF obtained in step 7 is washed three times with the final injection water. 9 Cell quantification The SVF of Step 8 is subjected to morphological analysis using an inverted microscope and the cell viability and the yield of stem cells are confirmed through an automated cell measuring device. 10 charge
label In order to be administered to the patient of Step 9, the dose is 1 × 10 ⁷ cells / cc in the water for injection and is placed in a 2 cc syringe. 11 SVF forwarding The final SVF procedure manager in step 10 is informed and confirmed receipt.

<Example 3> Analysis of composition of stem cells

The composition of the stem cell fraction was confirmed, and the average value of individual composition was as follows.

Adipose derived stem cell: about 20%

Adipocyte: about 20%

Endothelial progenitor cell: about 20%

Growth factor: about 20%

Red blood cell: about 20%

Normal saline: 2,000 μl

Claims (10)

A pharmaceutical composition for preventing or treating temporomandibular joint disorder comprising stem cells. The method according to claim 1,
Wherein the stem cell is derived from a tissue selected from the group consisting of fat, uterus, bone marrow, muscle, placenta, umbilical cord blood, hair follicle, and skin. The method of claim 2,
The pharmaceutical composition for preventing or treating temporomandibular joint disorder, wherein the adipose-derived stem cell is derived from a self or a teratoma. The pharmaceutical composition according to claim 2, wherein the adipose-derived stem cells are derived from adipose tissue isolated from subcutaneous, testicular, mesenteric, peritoneal, facial, thigh, or calf. The method according to claim 1,
The pharmaceutical composition for preventing or treating temporomandibular joint disorder, wherein the stem cell is an autologous fat-derived stem cell. The method according to claim 1,
Wherein the stem cell can be treated before or after the rinsing. The stem cell according to claim 1, wherein the stem cell is selected from the group consisting of adipocytes, preadipocytes, mesenchymal stem cells (MSC), endothelial progenitor cells, T cells, B cells, A cell growth factor, a phagocytic cell, a phagocyte, and a cell growth factor. [Claim 7] The pharmaceutical composition for preventing or treating temporomandibular joint disorder according to claim 7, wherein the mesenchymal stem cells are contained at 1 × 10 ² to 1 × 10 ¹⁰ cells / mL. The pharmaceutical composition according to claim 1, wherein the temporomandibular joint disorder is selected from the group consisting of jaw joint inflammation, temporomandibular joint pain, head and neck pain, limitation of opening, difficulty in chewing and facial deformity. A kit for treating jaw joint disorders, comprising the composition of claims 1 to 9.

Images