KR20230090117A - Phamaceutical composition for preventing or treating chronic obstructive pulmonary disease comprising evodiamine as an active ingredient - Google Patents

Abstract

The present invention relates to a composition for preventing or treating chronic obstructive pulmonary disease containing ebodiamine as an active ingredient. The composition containing the ebodiamine of the present invention as an active ingredient inhibits the expression of inflammatory cytokines TNF-α, IL-1β, and IL-6 and the expression of chemokines CCL2 and CXCL10, thereby reducing lung tissue damage and inflammatory response. Since it effectively suppresses, it is expected to be widely used for the treatment and prevention of chronic obstructive pulmonary diseases including emphysema.

Description

Pharmaceutical composition for preventing or treating chronic obstructive pulmonary disease comprising evodiamine as an active ingredient

The present invention relates to a composition for preventing or treating chronic obstructive pulmonary disease containing ebodiamine as an active ingredient.

Chronic obstructive pulmonary disease (COPD) is characterized by an abnormal inflammatory response in the lungs caused by inhalation of harmful particles (such as cigarettes) or gases, accompanied by not completely reversible and progressive airflow limitation. I mean respiratory disease. These chronic obstructive pulmonary diseases include emphysema, chronic bronchitis, and the like.

Chronic obstructive pulmonary disease is one of the top 10 causes of death in Korea, with an increasing incidence and mortality rate due to an increase in smoking rate and an extension of average life expectancy. In emphysema, one of the chronic obstructive pulmonary diseases, damage to the alveoli in the peripheral airways and the walls of the interalveolar spaces in the terminal bronchioles lose their elasticity and are destroyed, resulting in abnormal and permanent expansion. is a disease that The main cause of emphysema is known to be smoking, but it can also be caused by air pollution or exposure to hazardous work environments, and it is known that the rate of progression is accelerated when accompanied by respiratory infections. In the main symptom of emphysema, alveolar enlargement due to tissue destruction, expression of proteolytic enzymes such as matrix metalloproteinase (MMP) and neutrophil elastase are activated, whereas proteolytic enzymes It is known that the expression of α1-antitrypsin, which inhibits it, is reduced. In addition, smoking induces the secretion of various cytokines and chemokines from macrophages and epithelium in the alveoli, activating the inflammatory response, and cytokines/chemokines (TNF-α, IL- 1β, IL-8, MCP-1, IFN-γ, etc.), reactive oxygen species (ROS), etc. are increased, which accelerates tissue damage due to increased expression and activation of proteolytic enzymes. is known to be Emphysema is a disease in which the lung structure is irreversibly damaged and cannot be fundamentally treated. As currently used therapeutic agents, only symptomatic treatment methods according to symptoms such as steroid preparations for relieving inflammatory reactions, bronchodilators for relieving respiratory distress, and expectorants are used.

Therefore, in order to effectively treat chronic obstructive pulmonary disease, there is an urgent need to develop a therapeutic agent capable of reducing the rate of damage to lung structures or inhibiting lung tissue damage.

Domestic registered patent 10-1647029

The present invention was made to solve the above problems in the prior art, and it was confirmed that a composition containing evodiamine as an active ingredient effectively inhibits lung tissue damage and inflammatory response, thereby preventing or treating chronic obstructive pulmonary disease. The effect was confirmed, and based on this, the present invention was completed.

Accordingly, an object of the present invention is to provide a pharmaceutical composition for preventing or treating chronic obstructive pulmonary disease, comprising evodiamine or a pharmaceutically acceptable salt thereof as an active ingredient.

Another object of the present invention is to provide a food composition for preventing or improving chronic obstructive pulmonary disease, comprising evodiamine or a pharmaceutically acceptable salt thereof as an active ingredient.

However, the technical problem to be achieved by the present invention is not limited to the above-mentioned problems, and other problems not mentioned will be clearly understood by those skilled in the art from the following description.

In order to achieve the object of the present invention, the present invention provides a pharmaceutical composition for preventing or treating chronic obstructive pulmonary disease, comprising evodiamine or a pharmaceutically acceptable salt thereof as an active ingredient.

In addition, the present invention provides a food composition for preventing or improving chronic obstructive pulmonary disease, comprising evodiamine or a pharmaceutically acceptable salt thereof as an active ingredient.

In addition, the present invention provides a method for preventing or treating chronic obstructive pulmonary disease, comprising administering to a subject a composition containing the evodiamine or a pharmaceutically acceptable salt thereof as an active ingredient.

In addition, the present invention provides a preventive or therapeutic use of a composition containing evodiamine or a pharmaceutically acceptable salt thereof as an active ingredient for chronic obstructive pulmonary disease.

In addition, the present invention provides a use of evodiamine or a pharmaceutically acceptable salt thereof for the preparation of a drug for preventing or treating chronic obstructive pulmonary disease.

In one embodiment of the present invention, the chronic obstructive pulmonary disease may be emphysema, but is not limited thereto.

In another embodiment of the present invention, the evodiamine may inhibit an increase in lung compliance by elastase, but is not limited thereto.

In another embodiment of the present invention, the evodiamine may be characterized in that it inhibits a decrease in lung elasticity caused by elastase, but is not limited thereto.

In another embodiment of the present invention, the evodiamine is selected from the group consisting of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6). It may be characterized by inhibiting the expression of caine, but is not limited thereto.

In another embodiment of the present invention, the evodiamine may be characterized in that it inhibits the expression of one or more chemokines selected from the group consisting of CCL2 (chemokine ligand 2) and CXCL10 (C-X-C motif chemokine ligand 10), but is limited thereto it is not going to be

The composition containing the ebodiamine of the present invention as an active ingredient inhibits the expression of inflammatory cytokines TNF-α, IL-1β, and IL-6 and the expression of chemokines CCL2 and CXCL10, thereby reducing lung tissue damage and inflammatory response. Since it effectively suppresses, it is expected to be widely used for the treatment and prevention of chronic obstructive pulmonary diseases including emphysema.

1 shows the toxicity of evodiamine according to an embodiment of the present invention to mouse lung epithelial cells.
Figure 2 shows the result of NO production according to evodiamine treatment in terms of nitrite concentration in mouse-derived alveolar macrophages according to an embodiment of the present invention.
Figure 3 shows the results of measuring reactive oxygen species (ROS) according to evodiamine treatment in mouse-derived alveolar macrophages according to an embodiment of the present invention.
Figure 4 is isolated mRNA from mouse-derived alveolar macrophages according to an embodiment of the present invention, expression of inflammatory cytokines (TNF, IL-1β and IL-6) and chemokines (CCL2 and CXCL10) according to evodiamine treatment It shows the result of confirming the inhibitory effect.
Figure 5 shows the results of confirming the effect of reducing lung compliance and increasing lung elasticity according to evodiamine treatment using FlexiVent RM in an emphysema mouse model according to an embodiment of the present invention.
Figure 6 shows the results of confirming the effect of evodiamine treatment on lung tissue damage inhibition through H&E staining in an emphysema mouse model according to an embodiment of the present invention.
7 shows the result of confirming the weight change of the mouse by evodiamine in the emphysema mouse model according to an embodiment of the present invention.
Figure 8 shows the results of confirming changes in hepatotoxicity (GPT/GOT) and kidney toxicity (BUN/creatinine) indicators in the emphysema mouse model according to an embodiment of the present invention.

In one embodiment of the present invention, mouse-derived lung epithelial cells were treated with evodiamine, and then cell viability was analyzed to confirm that evodiamine had low cytotoxicity (see Example 1).

In another example of the present invention, mouse-derived alveolar macrophages were treated with evodiamine, and then the NO production inhibitory effect was confirmed to confirm the anti-inflammatory effect of evodiamine (see Example 2).

In another embodiment of the present invention, after treatment of mouse-derived alveolar macrophages with evodiamine, the inhibition of reactive oxygen species production was confirmed to confirm the antioxidant effect of evodiamine (see Example 3).

In another example of the present invention, mouse-derived alveolar macrophages were treated with evodiamine, and then the effect of inhibiting the expression of inflammatory cytokines and chemokines was confirmed (see Example 4).

In another example of the present invention, the effect of reducing lung compliance, increasing lung elasticity, and reducing lung tissue damage according to evodiamine treatment in an emphysema mouse model was confirmed (see Example 5).

In another example of the present invention, it was confirmed that there was no change in the weight of the mouse and no difference in hepatotoxicity and renal toxicity indices according to evodiamine treatment in an emphysema mouse model (see Example 6).

Therefore, in the absence of effective therapeutic agents and treatment methods for chronic obstructive pulmonary disease, the evodiamine of the present invention is expected to be effectively used for the treatment or prevention of chronic obstructive pulmonary disease.

Hereinafter, the present invention will be described in detail.

The present invention provides a pharmaceutical composition for preventing or treating chronic obstructive pulmonary disease, comprising evodiamine or a pharmaceutically acceptable salt thereof as an active ingredient.

As used herein, “chronic obstructive pulmonary disease (COPD)” refers to an abnormal inflammatory response of the lungs caused by inhalation of harmful particles (such as cigarettes) or gases, accompanied by an abnormal inflammatory response that is not completely reversible and gradually progresses. It refers to a respiratory disease characterized by airflow limitation, and according to an embodiment of the present invention, it may be emphysema, but is not limited thereto.

As used herein, “emphysema” refers to damage to the alveoli in the peripheral airway and the walls of the interalveolar space of the terminal bronchioles to lose their elasticity and be destroyed, resulting in abnormal and permanent expansion, resulting in reduced oxygen-carbon dioxide exchange capacity of the lungs and breathing It is a disease that causes trouble. Emphysema progresses when the secretion of various cytokines and chemokines is induced in alveolar macrophages and epithelium due to smoking, etc., and an inflammatory response is activated. Emphysema increases the production of cytokines/chemokines (TNF-α, IL-1β, IL-8, MCP-1, IFN-γ, etc.) and reactive oxygen species (ROS) by inflammation-related cells. As a result, the expression of proteolytic enzymes is increased and activated, accelerating tissue damage.

In the present specification, "evodiamine" may be represented by Formula 1 below, (1S) -21-methyl-3,13,21-triazapentacyclo[11.8.0.0 ^2,10 .0 ^4,9 . It may have the IUPAC name of 0 ^15,20 ]henicosa-2(10),4,6,8,15,17,19-heptaen-14-one. Also, the molecular weight may be 303.4 and the chemical formula may be C ₁₉ H ₁₇ N ₃ O. In addition, the ebodiamine may be chemically synthesized by a method known in the field to which the present invention belongs, or a commercially available material may be used, and may be obtained by extraction and separation, but is not limited thereto.

[Formula 1]

The evodiamine may inhibit an increase in lung compliance by elastase, but is not limited thereto. Specifically, the ebodiamine may restore an increase in lung compliance by elastase to a normal level, but is not limited thereto. The normal level means that the increase in lung compliance by elastase is reduced, and there is no significant difference from the lung compliance in the control group not treated with elastase.

In addition, the evodiamine may inhibit a decrease in lung elasticity caused by elastase, but is not limited thereto. Specifically, the ebodiamine may restore a decrease in lung elasticity caused by elastase to a normal level, but is not limited thereto. The normal level means that the decrease in lung elasticity caused by elastase is increased so that there is no significant difference from the lung elasticity of the control group not treated with elastase.

In addition, the ebodiamine may inhibit the expression of inflammation-related cytokines (TNF-α, IL-1β, IL-6, etc.) or chemokines (CCL2, CXCL10, etc.), but is not limited thereto.

The lung compliance refers to a change in volume according to a change in unit pressure in the lung. In the case of emphysema, the walls of the interalveolar space lose their elasticity and are destroyed, resulting in abnormal permanent dilatation, resulting in increased lung compliance.

The lung elasticity (tissue elastance) is the ability to breathe by the muscle contraction action of the diaphragm, and when the lung elasticity is reduced, the airway collapses and airflow obstruction occurs. In the case of emphysema, the walls of the interalveolar space lose their elasticity and are destroyed, resulting in a decrease in lung elasticity.

The present invention may also include a pharmaceutically acceptable salt of ebodiamine as an active ingredient. As used herein, the term "pharmaceutically acceptable salt" includes salts derived from pharmaceutically acceptable inorganic acids, organic acids, or bases.

Examples of suitable acids are hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, perchloric acid, fumaric acid, maleic acid, phosphoric acid, glycolic acid, lactic acid, salicylic acid, succinic acid, toluene-p-sulfonic acid, tartaric acid, acetic acid, citric acid, methanesulfonic acid, formic acid , benzoic acid, malonic acid, gluconic acid, naphthalene-2-sulfonic acid, benzenesulfonic acid and the like. Acid addition salts can be prepared by conventional methods, for example, by dissolving a compound in an aqueous solution of excess acid and precipitating the salt using a water-miscible organic solvent such as methanol, ethanol, acetone or acetonitrile. It can also be prepared by heating equimolar amounts of the compound and an acid or alcohol in water and then evaporating the mixture to dryness, or suction filtering the precipitated salt.

Salts derived from suitable bases may include, but are not limited to, alkali metals such as sodium and potassium, alkaline earth metals such as magnesium, and ammonium. An alkali metal or alkaline earth metal salt can be obtained, for example, by dissolving a compound in an excess alkali metal hydroxide or alkaline earth metal hydroxide solution, filtering the undissolved compound salt, and then evaporating and drying the filtrate. At this time, as the metal salt, it is particularly suitable for pharmaceutical purposes to prepare a sodium, potassium or calcium salt, and the corresponding silver salt can be obtained by reacting an alkali metal or alkaline earth metal salt with a suitable silver salt (eg, silver nitrate).

The content of ebodiamine in the composition of the present invention can be appropriately adjusted according to the symptoms of the disease, the progress of the symptoms, the condition of the patient, etc., for example, 0.0001 to 99.9% by weight, or 0.001 to 50% by weight based on the total weight of the composition. It may be, but is not limited thereto. The content ratio is a value based on the dry amount after removing the solvent.

The ebodiamine in the composition of the present invention is 1 to 500 μM, 1 to 400 μM, 1 to 300 μM, 1 to 200 μM, 1 to 150 μM, 2 to 500 μM, 2 to 400 μM, 2 to 300 μM, 2 to 200 μM, 2 to 150 μM, 5 to 500 μM, 5 to 400 μM, 5 to 300 μM, 5 to 200 μM, 5 to 150 μM, 7 to 500 μM, 7 to 400 μM, 7 to 300 μM, 7 to 200 μM, 7 to 150 μM, or 7 to 120 μM, but may be included at a concentration, but is not limited thereto.

The pharmaceutical composition according to the present invention may further include suitable carriers, excipients and diluents commonly used in the manufacture of pharmaceutical compositions. The excipient may be, for example, one or more selected from the group consisting of a diluent, a binder, a disintegrant, a lubricant, an adsorbent, a moisturizer, a film-coating material, and a controlled release additive.

The pharmaceutical compositions according to the present invention are powders, granules, sustained-release granules, enteric granules, solutions, eye drops, elsilic agents, emulsions, suspensions, spirits, troches, perfumes, and limonadese, respectively, according to conventional methods. , tablets, sustained-release tablets, enteric tablets, sublingual tablets, hard capsules, soft capsules, sustained-release capsules, enteric capsules, pills, tinctures, soft extracts, dry extracts, fluid extracts, injections, capsules, perfusate, It can be formulated and used in the form of an external agent such as a warning agent, lotion, pasta agent, spray, inhalant, patch, sterile injection solution, or aerosol, and the external agent is a cream, gel, patch, spray, ointment, warning agent , lotion, liniment, pasta, or cataplasma may have formulations such as the like.

Carriers, excipients and diluents that may be included in the pharmaceutical composition according to the present invention include lactose, dextrose, sucrose, oligosaccharide, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil.

When formulated, it is prepared using diluents or excipients such as commonly used fillers, extenders, binders, wetting agents, disintegrants, and surfactants.

Corn starch, potato starch, wheat starch, lactose, sucrose, glucose, fructose, di-mannitol, precipitated calcium carbonate, synthetic aluminum silicate, phosphoric acid as additives for tablets, powders, granules, capsules, pills, and troches according to the present invention Calcium monohydrogen, calcium sulfate, sodium chloride, sodium bicarbonate, purified lanolin, microcrystalline cellulose, dextrin, sodium alginate, methylcellulose, sodium carboxymethylcellulose, kaolin, urea, colloidal silica gel, hydroxypropyl starch, hydroxypropylmethyl Excipients such as cellulose (HPMC) 1928, HPMC 2208, HPMC 2906, HPMC 2910, propylene glycol, casein, calcium lactate, Primogel; Gelatin, gum arabic, ethanol, agar powder, cellulose phthalate acetate, carboxymethyl cellulose, calcium carboxymethyl cellulose, glucose, purified water, sodium caseinate, glycerin, stearic acid, sodium carboxymethyl cellulose, sodium methyl cellulose, methyl cellulose, microcrystalline cellulose, dextrin Binders such as hydroxycellulose, hydroxypropyl starch, hydroxymethylcellulose, purified shellac, starch arc, hydroxypropylcellulose, hydroxypropylmethylcellulose, polyvinyl alcohol, and polyvinylpyrrolidone may be used, Hydroxypropyl Methyl Cellulose, Corn Starch, Agar Powder, Methyl Cellulose, Bentonite, Hydroxypropyl Starch, Sodium Carboxymethyl Cellulose, Sodium Alginate, Calcium Carboxymethyl Cellulose, Calcium Citrate, Sodium Lauryl Sulfate, Silicic Anhydride, 1-Hydroxy Propyl cellulose, dextran, ion exchange resin, polyvinyl acetate, formaldehyde-treated casein and gelatin, alginic acid, amylose, guar gum, sodium bicarbonate, polyvinylpyrrolidone, calcium phosphate, gelled starch, gum arabic, disintegrants such as amylopectin, pectin, sodium polyphosphate, ethyl cellulose, white sugar, magnesium aluminum silicate, di-sorbitol solution, and light anhydrous silicic acid; Calcium stearate, magnesium stearate, stearic acid, hydrogenated vegetable oil, talc, lycopod, kaolin, petrolatum, sodium stearate, cacao butter, sodium salicylate, magnesium salicylate, polyethylene glycol 4000, polyethylene glycol 6000, liquid paraffin, hydrogenated soybean oil (Lubri wax), aluminum stearate, zinc stearate, sodium lauryl sulfate, magnesium oxide, macrogol, synthetic aluminum silicate, silicic anhydride, higher fatty acid, higher alcohol, silicone oil, paraffin oil, polyethylene glycol fatty acid ether, starch, A lubricant such as sodium chloride, sodium acetate, sodium oleate, dl-leucine, light anhydrous silicic acid; may be used.

Additives for the liquid formulation according to the present invention include water, dilute hydrochloric acid, dilute sulfuric acid, sodium citrate, sucrose monostearate, polyoxyethylene sorbitol fatty acid esters (tween esters), polyoxyethylene monoalkyl ethers, lanolin ethers, Lanolin esters, acetic acid, hydrochloric acid, aqueous ammonia, ammonium carbonate, potassium hydroxide, sodium hydroxide, prolamine, polyvinylpyrrolidone, ethyl cellulose, sodium carboxymethyl cellulose, and the like may be used.

In the syrup according to the present invention, a solution of white sugar, other sugars, or a sweetener may be used, and aromatics, coloring agents, preservatives, stabilizers, suspending agents, emulsifiers, thickeners, etc. may be used as necessary.

Purified water may be used in the emulsion according to the present invention, and emulsifiers, preservatives, stabilizers, fragrances, etc. may be used as needed.

Acacia, tragacantha, methylcellulose, carboxymethylcellulose, carboxymethylcellulose sodium, microcrystalline cellulose, sodium alginate, hydroxypropylmethylcellulose, HPMC 1828, HPMC 2906, HPMC 2910, etc. and, if necessary, surfactants, preservatives, stabilizers, colorants, and fragrances may be used.

Injections according to the present invention include distilled water for injection, 0.9% sodium chloride injection, IV injection, dextrose injection, dextrose + sodium chloride injection, PEG, lactated IV injection, ethanol, propylene glycol, non-volatile oil-sesame oil , solvents such as cottonseed oil, peanut oil, soybean oil, corn oil, ethyl oleate, isopropyl myristate, and benzene benzoate; solubilizing agents such as sodium benzoate, sodium salicylate, sodium acetate, urea, urethane, monoethylacetamide, butazolidine, propylene glycol, twins, nijuntinamide, hexamine, and dimethylacetamide; buffers such as weak acids and their salts (acetic acid and sodium acetate), weak bases and their salts (ammonia and ammonium acetate), organic compounds, proteins, albumin, peptone, and gums; tonicity agents such as sodium chloride; Stabilizers such as sodium bisulfite (NaHSO ₃ ), carbon dioxide gas, sodium metabisulfite (Na ₂ S ₂ O ₅ ), sodium sulfite (Na ₂ SO ₃ ), nitrogen gas (N ₂ ), and ethylenediaminetetraacetic acid; Sulfating agents such as sodium bisulfide 0.1%, sodium formaldehyde sulfoxylate, thiourea, ethylenediamine disodium tetraacetate, acetone sodium bisulfite; analgesics such as benzyl alcohol, chlorobutanol, procaine hydrochloride, glucose, and calcium gluconate; Suspending agents such as Siemesis sodium, sodium alginate, Tween 80, aluminum monostearate may be included.

The suppository according to the present invention includes cacao butter, lanolin, witapsol, polyethylene glycol, glycerogelatin, methylcellulose, carboxymethylcellulose, a mixture of stearic acid and oleic acid, subanal, cottonseed oil, peanut oil, palm oil, cacao butter + Cholesterol, Lecithin, Lannet Wax, Glycerol Monostearate, Tween or Span, Imhausen, Monolen (Propylene Glycol Monostearate), Glycerin, Adeps Solidus, Buytyrum Tego-G -G), Cebes Pharma 16, Hexalide Base 95, Cotomar, Hydroxycote SP, S-70-XXA, S-70-XX75 (S-70-XX95), Hyde Hydrokote 25, Hydrokote 711, Idropostal, Massa estrarium (A, AS, B, C, D, E, I, T), Massa-MF, Masupol, Masupol-15, Neosupostal-N, Paramound-B, Suposiro (OSI, OSIX, A, B, C, D, H, L), suppository type IV (AB, B, A, BC, BBG, E, BGF, C, D, 299), Supostal (N, Es), Wecobi (W, R, S, M, Fs), testosterone triglyceride base (TG-95, MA, 57) and The same mechanism can be used.

Solid preparations for oral administration include tablets, pills, powders, granules, capsules, etc., and these solid preparations contain at least one excipient, for example, starch, calcium carbonate, sucrose, etc. ) or by mixing lactose and gelatin. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used.

Liquid preparations for oral administration include suspensions, solutions for oral administration, emulsions, syrups, etc. In addition to water and liquid paraffin, which are commonly used simple diluents, various excipients such as wetting agents, sweeteners, aromatics, and preservatives may be included. there is. Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solvents, suspensions, emulsions, freeze-dried formulations, and suppositories. Propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate may be used as non-aqueous solvents and suspending agents.

The pharmaceutical composition according to the present invention is administered in a pharmaceutically effective amount. In the present invention, "pharmaceutically effective amount" means an amount sufficient to treat a disease with a reasonable benefit / risk ratio applicable to medical treatment, and the effective dose level is the type of patient's disease, severity, activity of the drug, It may be determined according to factors including sensitivity to the drug, administration time, route of administration and excretion rate, duration of treatment, drugs used concurrently, and other factors well known in the medical field.

The pharmaceutical composition according to the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, may be administered sequentially or simultaneously with conventional therapeutic agents, and may be administered single or multiple times. Considering all of the above factors, it is important to administer an amount that can obtain the maximum effect with the minimum amount without side effects, which can be easily determined by a person skilled in the art to which the present invention belongs.

The pharmaceutical composition of the present invention can be administered to a subject by various routes. All modes of administration can be envisaged, eg oral administration, subcutaneous injection, intraperitoneal administration, intravenous injection, intramuscular injection, paraspinal space (intrathecal) injection, sublingual administration, buccal administration, intrarectal insertion, vaginal It can be administered by intraoral insertion, ocular administration, otic administration, nasal administration, inhalation, spraying through the mouth or nose, dermal administration, transdermal administration, and the like.

The pharmaceutical composition of the present invention is determined according to the type of drug as an active ingredient together with various related factors such as the disease to be treated, the route of administration, the age, sex, weight and severity of the disease of the patient.

In the present invention, "individual" means a subject in need of treatment of a disease, and more specifically, a human or non-human primate, mouse, rat, dog, cat, horse, cow, etc. of mammals.

In the present invention, "administration" means providing a given composition of the present invention to a subject by any suitable method.

In the present invention, “prevention” refers to any action that suppresses or delays the onset of a desired disease, and “treatment” means that the desired disease and its resulting metabolic abnormality are improved or improved by administration of the pharmaceutical composition according to the present invention. All actions that are advantageously altered are meant, and "improvement" means any action that reduces a parameter related to a target disease, for example, the severity of a symptom, by administration of the composition according to the present invention.

In addition, it provides a food composition for preventing or improving chronic obstructive pulmonary disease, comprising evodiamine or a pharmaceutically acceptable salt thereof as an active ingredient.

In the present invention, the term "acceptable salt in food science" includes salts derived from organic acids, inorganic acids, or bases acceptable in food science.

When the evodiamine of the present invention is used as a food additive, the evodiamine can be added as it is or used together with other foods or food ingredients, and can be appropriately used according to a conventional method. The mixing amount of the active ingredient may be appropriately determined according to the purpose of use (prevention, health or therapeutic treatment). In general, evodiamine of the present invention may be added in an amount of 15% by weight or less, or 10% by weight or less based on the raw material when preparing food or beverage. However, in the case of long-term intake for the purpose of health and hygiene or health control, the amount may be less than the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount greater than the above range.

There is no particular limitation on the type of food. Examples of foods to which the above substances can be added include meat, sausages, bread, chocolates, candies, snacks, confectionery, pizza, ramen, other noodles, gums, dairy products including ice creams, various soups, beverages, tea, drinks, There are alcoholic beverages and vitamin complexes, and includes all health functional foods in a conventional sense.

The health beverage composition according to the present invention may contain various flavoring agents or natural carbohydrates as additional components, like conventional beverages. The aforementioned natural carbohydrates are monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, polysaccharides such as dextrins and cyclodextrins, and sugar alcohols such as xylitol, sorbitol and erythritol. As the sweetener, natural sweeteners such as thaumatin and stevia extract, or synthetic sweeteners such as saccharin and aspartame may be used. The proportion of the natural carbohydrate is generally about 0.01-0.20 g, or about 0.04-0.10 g per 100 mL of the composition of the present invention.

In addition to the above, the composition of the present invention contains various nutrients, vitamins, electrolytes, flavors, colorants, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, A carbonation agent used in carbonated beverages and the like may be contained. In addition, the composition of the present invention may contain fruit flesh for preparing natural fruit juice, fruit juice beverages and vegetable beverages. These components may be used independently or in combination. The ratio of these additives is not critical, but is generally selected in the range of 0.01-0.20 parts by weight per 100 parts by weight of the composition of the present invention.

Hereinafter, the following examples are presented to aid understanding of the present invention. However, the following examples are provided to more easily understand the present invention, and the content of the present invention is not limited by the following examples.

[Example]

Example 1. Confirmation of Toxicity of Ebodiamine to Normal Cells

In order to confirm the cytotoxicity of evodiamine, 4x10 ⁴ mouse-derived lung epithelial cells were added to each well of a 96-well plate, and then treated with evodiamine at concentrations of 10, 20, and 50 μM, and incubated at 37°C and 5% CO. ₂ conditions were cultured for 24 hours in a cell incubator. Cell viability was measured using a cell counting kit-8 according to the instructions provided by the manufacturer. Cell viability was calculated as a percentage of the absorbance of the experimental group relative to the absorbance of the control group (experimental group treated with medium only). The results are shown in Figure 1.

As shown in FIG. 1 , even when evodiamine was treated at a concentration of 50 μΜ, normal cells showed a survival rate of 50% or more, confirming that evodiamine had low cytotoxicity.

Example 2. Confirmation of the anti-inflammatory effect of evodiamine

Mouse-derived alveolar macrophages, MH-S, were treated with evodiamine (10, 20, 50 μΜ) and LPS (lipopolysaccharide) for 20 hours, and NO released in the medium was evaluated in terms of nitrite concentration. Specifically, MH-S cells were added to each well of a 24-well plate in an amount of 2.5x10 ⁵ , and after 1 day, various concentrations of evodiamine were treated with 0.5 μg/ml of LPS and treated at 37° C. and 5% CO ₂ conditions. It was cultured for 20 hours in a phosphorus cell incubator. After incubation, 50 μL of the medium was placed in a 96-well plate, 1% sulfanilamide solution prepared in 5% phosphoric acid solution was added, and after incubation at room temperature for 10 minutes, 0.1% N-1-naphthylethylenediamine diamine was added. 50 μL of the hydrochloride solution was treated, and absorbance was measured at 540 nm to quantitatively measure the degree of color development. Diluted sodium nitrite solutions of various concentrations were reacted under the same conditions, and a calibration curve was prepared to convert NO in the medium to nitrite concentration. The results are shown in Figure 2.

As shown in Figure 2, evodiamine inhibited NO production by LPS. This means that evodiamine has an anti-inflammatory effect.

Example 3. Confirmation of antioxidant effect of ebodiamine

Mouse-derived alveolar macrophages, MH-S, were treated with evodiamine (10, 50, 100μΜ) and LPS (lipopolysaccharide) for 20 hours, and intracellular reactive oxygen species (ROS) were then treated with H ₂ DCF-DA was measured using Specifically, MH-S cells were added to each well of a 96-well black 96-well plate with a transparent bottom at 5x10 ⁴ cells, and after 1 day, various concentrations of evodiamine were treated with 0.5 μg/mL of LPS and 37° C., 5% It was cultured for 20 hours in a cell incubator under CO ₂ conditions. After incubation, the cells were treated with a 20 μM H ₂ DCF-DA solution and incubated for 1 hour in a cell incubator at 37° C. under 5% CO ₂ conditions, and fluorescence was measured at an excitation wavelength of 485 nm and an emission wavelength of 535 nm. . The results are shown in Figure 3.

As shown in Figure 3, evodiamine inhibited ROS production by LPS. This means that evodiamine has an antioxidant effect.

Example 4. Confirmation of the inhibitory effect of cytokine and chemokine expression related to inflammation of ebodiamine

Mouse-derived alveolar macrophages, MH-S, were treated with evodiamine (Evo) and LPS (lipopolysaccharide) for 20 hours, and mRNA was isolated to analyze changes in the expression of inflammatory cytokines and chemokines. Specifically, MH-S cells were treated with evodiamine along with 0.5 μg/mL of LPS, and cultured for 20 hours in a cell incubator at 37°C and 5% CO ₂ conditions. After culturing, the cells were disassembled with an easy-BLUE solution (Intron), and RNA was isolated from the cells using a phenol-chloroform method. Then, the amount and purity of RNA was determined by measuring absorbance at 260 nm, 280 nm and 230 nm, and then 1-2 μg of RNA was converted into cDNA using a first-strand cDNA synthesis kit, and 1/10 to 1 /50 diluted cDNA was amplified using nTaq-HOT DNA polymerase mixed with SYBR Green I dye, dNTP solution, and gene-selective primers. Amplification of cDNA was carried out by repeating the conditions of pretreatment at 95 ° C for 10 minutes, incubation at 95 ° C for 10 seconds and at 60 ° C for 26 seconds in total 50 times, and to verify the selectivity of SYBR Green dye incorporation into amplified cDNA. A melting curve analysis was performed for After determining the CT (cycle threshold) value by monitoring the amplification of cDNA in real time, gene expression changes were confirmed by the 2 ^-ΔΔCt method with reference to the amplification level of the GAPDH gene. The results are shown in FIG. 4 . In addition, the sequences of the primers used in the experiment are shown in Table 1 below.

As shown in Figure 4, evodiamine significantly inhibited the expression of inflammatory cytokines TNF-α, IL-1β and IL-6 and chemokines CCL2 and CXCL10.

Gene Forward Sequence (5'-3') Reverse Sequence (5'-3') Il6 CTC TGG GAA ATC GTG GAA AT CCA GTT TGG TAG CAT CCA TC Il1b AAC CTG CTG GTG TGT GAC GTT C CAG CAC GAG GCT TTT TTG TTG T Tnf -α CAG CCG ATG GGT TGT ACC TT TGT GGG TGA GGA GCA CGT AGT Ccl2 TTA AAA ACC TGG ATC GGA ACC AA GCA TTA GCT TCA GAT TTA CGG GT Cxcl10 TGC CCA CGT GTT GAG ATC ATT TGT GCG TGG CTT CAC TCC AG Gapdh CAT GTT CCA GTA TGA CTC CAC TC GGC CTC ACC CCA TTT GAT GT

Example 5. Confirmation of the effect of ebodiamine on emphysema

5.1. Confirmation of lung function

To confirm the effect of evodiamine on the deterioration of lung function due to emphysema in vivo, after oral administration of 20 mg/kg of evodiamine to FVB mice once, porcine pancreatic elastase (PPE; porcine pancreatic 0.25 U of elastase, a proteolytic enzyme that causes lung tissue destruction, was administered once into the airway. Then, evodiamine was administered 6 times a week for 3 weeks. Then, in order to confirm lung function, lung compliance and lung elasticity (tissue elastance, ability to breathe by muscle contraction of the diaphragm) of the mice were measured using the FlexiVent RM. The results are shown in FIG. 5 .

As shown in FIG. 5, when only PPE was treated, lung compliance increased and lung elasticity decreased, resulting in a marked decrease in lung function, but in the experimental group in which evodiamine was administered together with PPE, lung compliance decreased again and lung elasticity was increased again, confirming that lung function was restored. Through the above results, it was confirmed that ebodiamine has preventive and therapeutic effects on emphysema.

5.2. H&E staining

In order to confirm the effect of ebodiamine administration on damage to lung tissue, the lungs of two mice treated in the same manner as in Example 5.1 were separated, and the separated lungs were treated with 10% formalin for 24 hours. After fixation, a paraffin block was prepared using paraffin. Then, the prepared paraffin block was cut to a thickness of 4 μm, attached to a silane coated slide, and dried. The dried slides were put in a dry oven at 65° C. to deparaffinize for 16 hours, and washed by immersing them in a xylene solution twice for 5 minutes each. Then, the fixed lung tissue sections were deparaffinized and hydrated with 100%, 95%, and 70% alcohol, and washed with distilled water. Then, it was stained for 1 minute and 30 seconds using a Harris hematoxylin solution, washed by immersion with a 1% HCl alcohol solution and ammonia, and then stained for 30 seconds using an eosin solution. Then, after sequential dehydration using 95% ethanol and 100% ethanol, final washing using a xylene solution was carried out, and the cells were sealed, observed under a microscope, and photographs were obtained. In addition, this was quantitatively expressed as a mean linear intercept value. The results are shown in FIG. 6 .

As shown in FIG. 6, lung tissue of the control group administered only with PPE collapsed, but in the case of the experimental group administered with evodiamine together with PPE, it was confirmed that lung tissue damage was reduced.

Example 6. Confirmation of in vivo toxicity of ebodiamine

Changes in body weight and blood hepatotoxicity and kidney toxicity indicators of mice treated in the same manner as in Example 5.1 were confirmed. Specifically, the mouse weight change was confirmed by measuring the weight of the mouse at least once a week. In addition, changes in hepatotoxicity and nephrotoxicity indicators in the blood were measured as follows. After blood was collected from the euthanized mouse through cardiac sampling, the blood was coagulated by culturing at 4°C, and serum was collected by centrifugation at 3,000 rpm for 10 minutes at 4°C. ALT (alanine aminotransferase, GPT), AST (aspartate aminotransferase, GOT), BUN (blood urea nitrogen), and creatinine levels in serum were measured using a veterinary hematology analyzer. The results are shown in Figures 7 and 8.

As shown in Figure 7, no significant body weight change was observed in the mice treated with evodiamine.

As shown in FIG. 8 , blood concentrations of GPT/GOT, an index of hepatotoxicity, and BUN/creatinine, an index of renal toxicity, also showed no significant difference from those of the control group. This means that evodiamine is not toxic.

Through the above results, the composition containing ebodiamine as an active ingredient of the present invention inhibits lung damage and destruction in an emphysema animal model administered with PPE, and effectively reduces the inflammatory response, thereby maintaining lung function and/or confirmed recovery. Through this, it was confirmed that evodiamine can be used as a composition for preventing or treating chronic obstructive pulmonary disease.

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

<110> Seoul National University R&DB Foundation <120> Phamaceutical composition for preventing or treating chronic obstructive pulmonary disease comprising evodiamine as an active ingredient <130> MP21-248 <160> 12 <170> KoPatentIn 3.0 <210> 1 <211> 20 <212> DNA <213> artificial sequence <220> <223> Il6_F <400> 1 ctctgggaaa tcgtgggaaat 20 <210> 2 <211> 20 <212> DNA <213> artificial sequence <220> <223> Il6_R <400> 2 ccagtttggt agcatccatc 20 <210> 3 <211> 22 <212> DNA <213> artificial sequence <220> <223> Il1b_F <400> 3 aacctgctgg tgtgtgacgt tc 22 <210> 4 <211> 22 <212> DNA <213> artificial sequence <220> <223> Il1b_R <400> 4 cagcacgagg cttttttgtt gt 22 <210> 5 <211> 20 <212> DNA <213> artificial sequence <220> <223> Tnf-a_F <400> 5 cagccgatgg gttgtacctt 20 <210> 6 <211> 21 <212> DNA <213> artificial sequence <220> <223> Tnf-a_R <400> 6 tgtgggtgag gagcacgtag t 21 <210> 7 <211> 23 <212> DNA <213> artificial sequence <220> <223> Ccl2_F <400> 7 ttaaaaacct ggatcggaac caa 23 <210> 8 <211> 23 <212> DNA <213> artificial sequence <220> <223> Ccl2_R <400> 8 gcattagctt cagattacg ggt 23 <210> 9 <211> 21 <212> DNA <213> artificial sequence <220> <223> Cxcl10_F <400> 9 tgcccacgtg ttgagatcat t 21 <210> 10 <211> 20 <212> DNA <213> artificial sequence <220> <223> Cxcl10_R <400> 10 tgtgcgtggc ttcactccag 20 <210> 11 <211> 23 <212> DNA <213> artificial sequence <220> <223> Gapdh_F <400> 11 catgttccag tatgactcca ctc 23 <210> 12 <211> 20 <212> DNA <213> artificial sequence <220> <223> Gapdh_R <400> 12 ggcctcaccc catttgatgt 20

Claims (12)

A pharmaceutical composition for preventing or treating chronic obstructive pulmonary disease, comprising evodiamine or a pharmaceutically acceptable salt thereof as an active ingredient.
According to claim 1,
The chronic obstructive pulmonary disease is characterized in that emphysema (emphysema), a pharmaceutical composition for preventing or treating chronic obstructive pulmonary disease.
According to claim 1,
The evodiamine is a pharmaceutical composition for the prevention or treatment of chronic obstructive pulmonary disease, characterized in that inhibits the increase in lung compliance by elastase (elastase).
According to claim 1,
The evodiamine is a pharmaceutical composition for the prevention or treatment of chronic obstructive pulmonary disease, characterized in that for inhibiting the reduction of lung elasticity caused by elastase (elastase).
According to claim 1,
The evodiamine inhibits the expression of one or more inflammatory cytokines selected from the group consisting of TNF-α, IL-1β, and IL-6, a pharmaceutical composition for preventing or treating chronic obstructive pulmonary disease.
According to claim 1,
The evodiamine is a pharmaceutical composition for preventing or treating chronic obstructive pulmonary disease, characterized in that inhibiting the expression of one or more chemokines selected from the group consisting of CCL2 and CXCL10.
A food composition for preventing or improving chronic obstructive pulmonary disease, comprising evodiamine or a pharmaceutically acceptable salt thereof as an active ingredient.
According to claim 7,
The chronic obstructive pulmonary disease is a food composition for preventing or improving chronic obstructive pulmonary disease, characterized in that emphysema (emphysema).
According to claim 7,
The evodiamine is a food composition for preventing or improving chronic obstructive pulmonary disease, characterized in that it inhibits the increase in lung compliance by elastase (elastase).
According to claim 7,
The evodiamine is a food composition for preventing or improving chronic obstructive pulmonary disease, characterized in that for inhibiting the reduction of lung elasticity caused by elastase (elastase).
According to claim 7,
The evodiamine inhibits the expression of one or more inflammatory cytokines selected from the group consisting of TNF-α, IL-1β, and IL-6, a food composition for preventing or improving chronic obstructive pulmonary disease.
According to claim 7,
The evodiamine is a food composition for preventing or improving chronic obstructive pulmonary disease, characterized in that inhibiting the expression of one or more chemokines selected from the group consisting of CCL2 and CXCL10.

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