WO2020017915A1 - Pharmaceutical composition comprising phytoncide for preventing or treating sarcopenia - Google Patents

Abstract

Provided according to an embodiment of the present invention are a pharmaceutical composition comprising a phytoncide as an effective ingredient for preventing or treating sarcopenia and a food composition comprising a phytocide as an effective ingredient for preventing or alleviating sarcopenia.

Description

Pharmaceutical composition for the prevention or treatment of myopathy containing phytoncide

The present invention relates to a pharmaceutical composition for the prevention or treatment of myopathy containing phytoncide as an active ingredient.

The concept of sarcopenia began with the introduction of the term 'sarcopenia' by Irwin Rosenberg in 1989, where the Greek word "sarco" means muscle and "penia" means reduced. Muscular dystrophy refers to the decrease in muscle strength associated with aging with a decrease in muscle mass. "Muscle" here means skeletal muscle and has nothing to do with smooth muscle. In other words, myotropia mainly refers to a loss of skeletal muscle mass distributed in the extremities, and muscle loss due to acute diseases such as cachexia and flu, which are significant muscle loss states in the late stages of malignant tumors. wasting, or primary muscle disease.

Recently, the prevalence of osteoporosis and muscular dystrophy is increasing rapidly as the age group of 65 and older increases rapidly. It is estimated that a gradual decrease in muscle mass occurs after the 40s and decreases by 8% every 10 years until the 70s. After that, it is known that a more rapid decrease may occur and may occur up to 15% every 10 years. have. Many follow-up studies have shown that the physiological changes that occur in older people vary and, in general, decrease in muscle mass and bone density with age.

On the other hand, there are three major methods of treatment of muscular dystrophy. The first is exercise. Exercise has been reported to increase skeletal muscle protein synthesis in the short term and increase muscle strength or mobility in older adults. However, it is inadequate for long-term treatment. Second, testosterone or anabolic steroid can be used as a drug treatment, but this induces masculinity in females, and in men, side effects such as prostate symptoms. Other approved regimens include dehydroepiandrosterone (DHEA) and growth hormone, which have been reported to be therapeutic in areas containing SARMs (Selective Androgen Receptor Modulators). In addition, although diet is known as a treatment, nutritional assessments indicate that malnutrition and modern eating habits are inadequate to maintain adequate total body mass.

Myostatin is a polypeptide growth factor belonging to the superfamily of TGF-β. TGF-β has a large amount of isoforms, which are known to be involved in proliferation, apoptosis, differentiation, bone formation and maintenance of cells (Massague & Chen, 2000). Myostatin belongs to growth differentiation factor (GDF) No. 8, is involved in the growth and development of tissues, and acts by activating the Smad signaling system. In addition, the p21 gene has been reported to inhibit the proliferation of cell cycle and progenitor cells, thereby affecting bone formation and regeneration. Myostatin is produced mainly in skeletal muscle cells, causing muscle loss and muscle strength by the method of self-secretion. In addition, myostatin inhibits the expression of IGF-1 or Follistatin, which is involved in muscle hypertrophy, and causes protein synthesis and proliferation in myoblasts. It is known to suppress.

Against this background, the present inventors have made a diligent effort to find a substance that can treat myostatin by inhibiting myostatin expression that causes muscle loss and muscle strength. The present invention was completed by discovering that the cedrol) component can be used for preventing or treating myostatin by inhibiting myostatin expression.

It is an object of the present invention to provide a pharmaceutical composition for the prevention or treatment of myopathy containing phytoncide as an active ingredient.

Another object of the present invention relates to a food composition for preventing or improving myopathy containing phytoncide as an active ingredient.

However, the problem to be solved by the present invention is not limited to the above-mentioned problem, another task that is not mentioned will be clearly understood by those skilled in the art from the following description.

According to one embodiment of the present invention, there is provided a pharmaceutical composition for preventing or treating myopathy, containing phytoncide as an active ingredient.

According to one side, the phytoncide may contain caryophyllene oxide or cedrol.

According to one side, the pharmaceutical composition may inhibit the expression of myostatin mRNA or protein.

According to one side, the myostatin mRNA or protein expression may be due to bleomycin (bleomycin).

According to one side, the myostatin mRNA or protein expression may be due to UV.

According to one side, the pharmaceutical composition may inhibit the expression of MURF1 (Muscle RING-finger protein-1) mRNA or protein.

According to one side, the pharmaceutical composition may inhibit the expression of Foxo3 (forkhead box O3) mRNA or protein.

According to another embodiment of the present invention, there is provided a food composition for preventing or improving myopathy, containing phytoncide as an active ingredient.

The pharmaceutical composition for the prevention or treatment of myotropenia containing the phytoncide of the present invention as an active ingredient inhibits the expression of myostatin, which directly affects muscle loss and muscle loss, and thus prevents or treats more fundamental myotropin. Can be represented.

It is to be understood that the effects of the present invention are not limited to the above effects, and include all effects deduced from the configuration of the invention described in the detailed description or claims of the present invention.

Figure 1 confirms the effect of inhibiting myostatin mRNA and protein expression increased by UV and bleomycin when treated with cariophyllene oxide.

Figure 2 confirms the effect of inhibiting myostatin mRNA and protein expression increased by UV and bleomycin during cedrol treatment.

Figure 3 confirms the effect of inhibiting the increased MURF1 protein expression by UV during treatment with cariophyllene oxide and cedrol.

Figure 4 confirms the effect of inhibiting Foxo3 protein expression increased by UV during treatment with cariophyllene oxide and cedrol.

Hereinafter, exemplary embodiments will be described in detail with reference to the accompanying drawings. However, various changes may be made to the embodiments so that the scope of the patent application is not limited or limited by these embodiments. It is to be understood that all changes, equivalents, and substitutes for the embodiments are included in the scope of rights.

The terminology used herein is for the purpose of description and should not be construed as limiting. Singular expressions include plural expressions unless the context clearly indicates otherwise. In this specification, terms such as "comprise" or "have" are intended to indicate that there is a feature, number, step, operation, component, part, or combination thereof described on the specification, and one or more other features. It is to be understood that the present invention does not exclude the possibility of the presence or the addition of numbers, steps, operations, components, components, or a combination thereof.

Unless defined otherwise, all terms used herein, including technical or scientific terms, have the same meaning as commonly understood by one of ordinary skill in the art. Terms such as those defined in the commonly used dictionaries should be construed as having meanings consistent with the meanings in the context of the related art, and shall not be construed in ideal or excessively formal meanings unless expressly defined in this application. Do not.

In addition, in the description with reference to the accompanying drawings, the same components will be given the same reference numerals regardless of the reference numerals and duplicate description thereof will be omitted. In the following description of the embodiment, if it is determined that the detailed description of the related known technology may unnecessarily obscure the gist of the embodiment, the detailed description thereof will be omitted.

According to one embodiment of the present invention, there is provided a pharmaceutical composition for preventing or treating myopathy, containing phytoncide as an active ingredient.

As used herein, the term "phytoncide" refers to a bactericidal substance that plants produce to defend themselves from pests or microorganisms. Phytoncide is the main ingredient of isoprenoids (isoprenoid or terpene) compounds, which have a unique odor, and are known to be the most contributing factors to various forest bath effects such as cardiopulmonary function, vascular function, respiratory function and skin sterilization.

The phytoncide may contain caryophyllene oxide or cedrol, which is an isoprenoid-derived component. The structures of cariophyllene oxide and cedrol are as shown in the following formulas (1) and (2), respectively.

Isoprenoid is a hydrocarbon composed of a plurality of isoprenes (C ₅ H ₈ ), which are monoterpenes having two, three or four isoprenes, sesquiterpenes and diterpenes. 0.01 to% terpene is synthesized through photosynthesis of solar energy. In general, isoprenoids are obtained in the form of essential oils by steam distillation, compression or extraction.

The composition of the present invention can inhibit the expression of mRNA or protein of myostatin, MUFR1, and Foxo3, which are known as indices of myopathy, in particular myostatin expression by bleomycin or UV stimulation. Can be suppressed. The bleomycin is an anti-tumor antibiotic and is used as an effective anticancer agent for squamous cell carcinoma or malignant lymphoma, or is known as an aging-inducing factor.

Therefore, the composition of the present invention can be usefully used for the prevention or treatment of myopathy by aging, chemotherapy and the like.

As used herein, the term "prevention" means any action that inhibits or delays myopathy by administration of the composition.

As used herein, the term "treatment" means any action that improves or beneficially changes the symptoms of myotropia by administration of the composition.

The pharmaceutical composition of the present invention may be prepared by a method known in the pharmaceutical field for use as a medicament, and may be mixed with pharmaceutically acceptable carriers, excipients, diluents, stabilizers, preservatives, etc. to powders, granules, tablets, It may be prepared and used in the form of a capsule or injection. In addition, the composition may be prepared in a sustained release formulation including a base used for sustained release purposes in addition to the active ingredient so that the release of the active ingredient occurs slowly.

Pharmaceutically acceptable carriers may further include, for example, carriers for oral administration or carriers for parenteral administration. Carriers for oral administration may include lactose, starch, cellulose derivatives, magnesium stearate, stearic acid and the like. In addition, it may include various drug delivery materials used for oral administration to the peptide formulation. In addition, carriers for parenteral administration may include water, suitable oils, saline, aqueous glucose, glycols, and the like, and may further include stabilizers and preservatives. Suitable stabilizers include antioxidants such as sodium hydrogen sulfite, sodium sulfite or ascorbic acid. Suitable preservatives include benzalkonium chloride, methyl- or propyl-paraben and chlorobutanol. The pharmaceutical composition of the present invention may further include a lubricant, a humectant, a sweetener, a flavoring agent, an emulsifier, a suspension agent, and the like, in addition to the above components.

The diluent may be a non-aqueous solvent such as propylene glycol, polyethylene glycol, vegetable oils such as olive oil and peanut oil, or brine (preferably 0.8% saline), water containing a buffer medium (preferably 0.05M phosphate buffer) An aqueous solvent, such as these, etc. are mentioned, but it is not limited to this.

The excipients include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, anhydrous skim milk, glycerol, propylene, glycol, water , Ethanol and the like, but is not limited thereto.

The stabilizer may include carbohydrates such as sorbitol, mannitol, starch, sucrose, dextran, glutamate, and glucose, and proteins such as animal, vegetable or microbial proteins such as milk powder, serum albumin and casein, but are not limited thereto. no.

The preservative may include thimerosal, merthiolate, gentamicin, neomycin, nistatin, amphotericin B, tetracycline, penicillin, streptomycin, polymyxin B, and the like, but is not limited thereto.

The pharmaceutical compositions of the invention can be administered to any mammal, including humans, by any means, for example orally or parenterally. Parenteral administration methods include intravenous, intramuscular, intraarterial, intramedullary, intradural intracardiac, transdermal, subcutaneous, intraperitoneal, intranasal, intestinal, topical, sublingual or intrarectal administration no.

The pharmaceutical composition of the present invention may be formulated into a preparation for oral or parenteral administration according to the route of administration as described above.

The total effective amount of the pharmaceutical composition of the present invention may be administered to a patient in a single dose, and may be administered by a divided therapeutic regimen administered in multiple doses for a long time. The pharmaceutical composition of the present invention may vary the content of the active ingredient depending on the extent of the disease. This may be determined in consideration of various factors such as the formulation method, the number of administration routes, as well as the age, weight, health condition, symptoms of the disease, administration time and method of the patient. In view of this, one of ordinary skill in the art will be able to determine appropriate effective dosages of the compositions of the present invention. The pharmaceutical composition according to the present invention is not particularly limited in terms of dosage form, route of administration and method of administration as long as the effect of the present invention is shown.

According to another embodiment of the present invention, there is provided a food composition for preventing or improving myopathy, containing phytoncide as an active ingredient.

As used herein, the term “improvement” means any action that at least reduces the parameters associated with the condition being treated, for example, the extent of symptoms.

In addition, when the food composition of the present invention is used as a food additive, the composition may be added as it is or used with other food or food ingredients, and may be appropriately used according to a conventional method. Generally, in the preparation of food or beverages, the compositions of the invention are added in an amount of up to 15% by weight, preferably up to 10% by weight relative to the raw materials. However, in the case of long-term intake for the purpose of health and hygiene or for the purpose of health control, it may be below the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount above the above range.

There is no particular limitation on the kind of the food. Examples of the food to which the substance can be added include dairy products including meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, ice cream, various soups, drinks, tea, drink, Alcoholic beverages and vitamin complexes, and the like and include all of the health foods in the conventional sense.

Hereinafter, the present invention will be described in more detail with reference to Examples. The following examples are described for the purpose of illustrating the present invention, but the scope of the present invention is not limited thereto.

Example 1 Verification of the Inhibitory Effect of Myotropia Induced by UV Irradiation

C2C12 muscle progenitor cells (ATCC, US) were cultured in DMEM containing 10% FBS. Cells were seeded with 3x10 ⁵ cells in a 60-pie dish. The next day after seeding was treated with cetrol and caryophyllene oxide phytoncide-derived isoprenoids were left for 1 hour. After 1 hour, the growth media (10% FBS DMEM) was removed and washed with PBS. After removing PBS, the UV cross-linker (UVP CL-1000) was irradiated with UV of 3J / m² with the plate cap removed (0 day). Incubation in a CO ₂ incubator for 24 hours, the growth media was removed in the same manner PBS wash and then irradiated with UV of 3J / m² (1 day).

The next day, harvested at the same time (2 days) and Western blot was used to measure the expression changes of myostatin, MURF1 and Foxo3, the muscle protein regulatory target proteins in cell lysates. Myostatin uses Anti-GDF8 (Genetex, GTX32624), MURF1 uses anti-MURF1 antibody (Abcam, ab172479), Foxo3 uses anti-Foxo3a antibody (Cell Signaling # 12829) and Beta-actin antibody as a protein loading control Comparative analysis using (Enogene E12-041-4).

RNA was isolated using RNeasy Mini Kit (Qiagen, US), and cDNA was synthesized using AMV Reverse Transcripatse (Promega, US), and then the degree of myostatin mRNA expression was confirmed by cDNA by PCR. PCR conditions were initially denatured at 95 ° C. for 5 minutes,

Denaturation at 95 ° C. for 5 seconds,

Annealing at 62.8 ° C. for 10 seconds and

The final extension was carried out at 72 ° C. for 10 minutes. Detection of mRNA expression was carried out using the following primers.

As a result, as shown in Figures 1 to 4 it was confirmed that the expression of myostatin, MURF1 and Foxo3 protein increased by UV irradiation decreased with treatment with carriophyll oxide and cedrol. In particular, in the case of MURF1 protein expression inhibitory effect, it was confirmed that cedrol is superior to cariophyllene oxide.

Example 2 Validation of Myotropy Inhibitory Effect Induced by Bleomycin

C2C12 muscle progenitor cells (ATCC, US) were cultured in DMEM containing 10% FBS. At this time, the cells were seeded with 3x10 ⁵ cells in a 60-pie dish, and then left for 1 hour by bleomycin treatment after seeding.

Then, treated with isoprenoids cedrol and caryophyllene oxide, respectively, and left for 1 hour, the growth media (10% FBS DMEM) was removed and washed with PBS. After washing, Western blot method was used to measure the expression changes of myostatin and MURF1, the muscle protein regulatory target proteins in cell lysates. The expression of myostatin was analyzed using Anti-GDF8 (Genetex, GTX32624), and the beta-actin antibody (Enogene E12-041-4) as a protein loading control.

RNA was isolated using RNeasy Mini Kit (Qiagen, US), and cDNA was synthesized using AMV Reverse Transcripatse (Promega, US), and then the degree of myostatin mRNA expression was confirmed by cDNA by PCR. PCR conditions were initially denatured at 95 ° C. for 5 minutes,

Denaturation at 95 ° C. for 5 seconds,

Annealing at 62.8 ° C. for 10 seconds and

The final extension was carried out at 72 ° C. for 10 minutes. mRNA expression was confirmed using the same primers as described in Example 1 above.

As a result, as shown in Figures 1 and 2, it was confirmed that the expression of myostatin increased by bleomycin treatment decreased with treatment with cariophyllene oxide and cedrol.

Through the results of the above examples, phytoncide-derived isoprenoids, carophyllene oxide and cedrol, are useful for preventing or treating myotropin by inhibiting the expression of myostatin, MURF1 and Foxo3, which are indicator proteins related to muscle mass control. It can be seen that it can be used.

Although the embodiments have been described with reference to the accompanying drawings, those skilled in the art may apply various technical modifications and variations based on the above. For example, the described techniques may be performed in a different order than the described method, and / or the described components may be combined or combined in a different form than the described method, or replaced or substituted by other components or equivalents. Appropriate results can be achieved.

Therefore, other implementations, other embodiments, and equivalents to the claims are within the scope of the following claims.

Claims (8)

1. A pharmaceutical composition for preventing or treating myopathy, containing phytoncide as an active ingredient.
2. The method of claim 1,

   The phytoncide contains caryophyllene oxide or cedrol, cedrol, a pharmaceutical composition for preventing or treating myopathy.
3. The method of claim 1,

   A pharmaceutical composition for preventing or treating myostatin, which inhibits myostatin mRNA or protein expression.
4. The method of claim 3,

   The myostatin mRNA or protein expression is due to bleomycin (bleomycin), a pharmaceutical composition for preventing or treating myopathy.
5. The method of claim 3,

   The myostatin mRNA or protein expression is due to UV, pharmaceutical composition for the prevention or treatment of myopathy.
6. The method of claim 1,

   MURF1 (Muscle RING-finger protein-1) A pharmaceutical composition for preventing or treating myopathy, which inhibits mRNA or protein expression.
7. The method of claim 1,

   A pharmaceutical composition for preventing or treating myopathy, which inhibits Foxo3 (forkhead box O3) mRNA or protein expression.
8. A food composition for preventing or improving myopathy, containing phytoncide as an active ingredient.

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