KR20240018292A - Anti-inflammatory composition - Google Patents

Abstract

The present specification relates to anti-inflammatory compositions and compositions for preventing, alleviating or treating inflammatory diseases. The composition according to one aspect of the present invention is an inflammatory disease mouse model induced by lipopolysaccharide (LPS) or cecal ligation puncture (CLP), In an inflammatory disease model induced by ligand treatment of toll-like receptors and an inflammatory disease model infected with typhoid fever, Mycobacterium tuberculosis, and BCG bacteria, respectively, inflammatory cytokines such as IL-1b, IL-6, and TNF-α, and CXCL5 and CXCL10 It has anti-inflammatory activity that suppresses the expression level of chemokines and has an excellent effect in preventing, alleviating or treating inflammatory diseases caused by bacterial infections or autoimmune diseases, and the composition can be used as a therapeutic agent, food, food additive, feed additive, and disinfectant. , can be usefully used in cosmetics, etc.

Description

Anti-inflammatory composition

The present specification relates to anti-inflammatory compositions and compositions for preventing, alleviating or treating inflammatory diseases.

Inflammation is one of the most important health problems worldwide. Inflammation is generally a localized protective response of body tissues against host invasion by foreign substances or harmful stimuli. Causes of inflammation include infectious causes such as bacteria, viruses, and parasites; Physical causes such as burns or irradiation; Chemicals such as toxins, drugs or industrial agents; It may be a condition associated with an immune response, such as allergies and autoimmune reactions, or oxidative stress. Inflammation is characterized by pain, redness, swelling, heat, and ultimate loss of function of the affected area. These symptoms are the result of a complex series of interactions between cells of the immune system. The cellular response results in the creation of an interactive network of several groups of inflammatory mediators: proteins (e.g. cytokines, enzymes (e.g. proteases, peroxidases), major basic proteins, and adhesion molecules (ICAMs). VCAM), lipid mediators (e.g., eicosanoids, prostaglandins, leukotrienes, platelet-activating factor (PAF)), reactive oxygen species (e.g., hydroperoxide, superoxide anion O ^{2 -} , nitric oxide ( NO), etc.). However, most of these mediators of inflammation are also regulators of normal cellular activity. Accordingly, dysfunction of the inflammatory response can result in uncontrolled damage (i.e. infection) to the host, thus leading to chronic inflammation. This causes inflammatory diseases that are partially mediated by excessive production of several of the above-mentioned mediators.

Autoimmune disease, one of the inflammatory diseases, is characterized by the immune system attacking its own organs and causing a spontaneous response. These reactions are due to the recognition of auto-antigens by T lymphocytes, which triggers humoral (autoantibody production) and cellular (increased cytotoxic activity of lymphocytes and macrophages) immune responses. Autoimmune diseases include: rheumatic diseases, psoriasis, systemic dermatomyositis, multiple sclerosis, lupus erythematosus, or aggravated immune response due to antigens, such as asthma, drug or food allergy, etc. These diseases are all limiting and chronic diseases, and in some cases are fatal, and there is currently no effective treatment method to treat the diseases. Therefore, any drug, medicine, or medium that can relieve or alleviate the disease during its progression can be considered an important solution for the patient's health.

Meanwhile, 'sepsis' induces a systemic inflammatory response due to excessive secretion of inflammatory mediators, including inflammatory cytokines, by pathogens such as bacteria and fungi that invade the living body, ultimately leading to tissue and organ damage and loss of function of white blood cells. It is a serious acute infectious inflammatory disease that paralyzes the ability to defend against infectious bacteria.

Sepsis is a typical acute inflammatory disease caused by excessive activity of various types of inflammatory immune cells, such as macrophages, monocytes, and neutrophils. These immune cells normally play a role in defense of the body by eliminating external infectious bacteria, but in a sepsis environment, they become excessively activated and produce various inflammatory cytokines or inflammatory mediators that are harmful to the living body, worsening the symptoms in a short period of time and making them uncontrollable. This causes abnormalities in vital vital organs and organs, leading to systemic shock and death. Severe sepsis is called a modern-day plague and is a major cause of death around the world. It kills 215,000 people every year in the United States alone, and is the third leading cause of death after cardiovascular disease and cancer.

Treatment of sepsis has emerged as a very important problem in relation to the gradual increase in pathogens resistant to various antibiotics, but a suitable treatment for it has not yet been developed.

Accordingly, while researching compounds with anti-inflammatory activity through a new mechanism of action, the present inventors confirmed that the compound according to the present invention, its salt, stereoisomer, solvate or hydrate had anti-inflammatory activity and completed the present invention. .

The present inventors conducted research to develop a substance with anti-inflammatory activity. As a result of the study, it was confirmed that the compound represented by the following formula (1), its possible salt, stereoisomer, solvate or hydrate, exhibits an anti-inflammatory effect that suppresses inflammation in an inflammation model.

Therefore, in one aspect, an object of the present invention is to provide an anti-inflammatory composition comprising a compound represented by the following formula (1), a salt, stereoisomer, solvate or hydrate thereof as an active ingredient:

[Formula 1]

In Formula 1,

R ₁ and R ₂ are independently -(CH ₂ ) _n1 -R _a , n1 is an integer of 1 to 3, and R _a is unsubstituted or fluoro-substituted phenyl;

R ₃ is -(CH ₂ ) _n2 - or -O-CH ₂ -CH(OH)-CH ₂ -, and n2 is an integer of 1 to 3;

R ₄ is -CH ₂ -CH ₂ - or -CH ₂ -CH ₂ -O-CH ₂ -CH ₂ -.

In another aspect, an object of the present invention is to provide a composition for preventing, alleviating or treating inflammatory diseases, comprising the compound represented by Formula 1, a salt, stereoisomer, solvate or hydrate thereof as an active ingredient.

One aspect of the present invention provides an anti-inflammatory composition comprising a compound represented by the following formula (1), a salt, stereoisomer, solvate or hydrate thereof as an active ingredient:

[Formula 1]

In Formula 1,

R ₁ and R ₂ are independently -(CH ₂ ) _n1 -R _a , n1 is an integer of 1 to 3, and R _a is unsubstituted or fluoro-substituted phenyl;

R ₃ is -(CH ₂ ) _n2 - or -O-CH ₂ -CH(OH)-CH ₂ -, and n2 is an integer of 1 to 3;

R ₄ is -CH ₂ -CH ₂ - or -CH ₂ -CH ₂ -O-CH ₂ -CH ₂ -.

Another aspect of the present invention provides a composition for preventing, alleviating, or treating inflammatory diseases, comprising the compound represented by Formula 1, a salt, stereoisomer, solvate, or hydrate thereof as an active ingredient.

A composition comprising the compound represented by Formula 1, a salt, stereoisomer, solvate or hydrate thereof as an active ingredient according to one aspect of the present invention is used to treat mice with inflammatory disease induced by lipopolysaccharide (LPS) or cecal ligation puncture (CLP). model, an inflammatory disease model induced by ligands of toll-like receptors, inflammatory cytokines such as IL-1b, IL-6, and TNF-α, and CXCL5 and CXCL10 in an inflammatory disease model infected with Salmonella, Mycobacterium tuberculosis, and BCG bacteria, respectively. It has anti-inflammatory activity that suppresses the expression level of chemokines such as, and has an excellent effect in preventing, alleviating, or treating inflammatory diseases caused by bacterial infections or autoimmune diseases, and the composition can be used as a therapeutic agent, food, food additive, or feed additive. , it can be useful as a disinfectant, cosmetics, etc.

1A and 1B are graphs showing changes in survival rate when a compound according to one aspect of the present invention is treated with an inflammatory disease mouse model induced by LPS treatment.
Figure 2 is a graph showing the change in survival rate when a compound according to one aspect of the present invention is treated with a mouse model of inflammatory disease induced by cecal ligation puncture (CLP).
Figures 3a to 3d show the lungs (FIGS. 3a and 3c) and spleen (FIGS. 3b and 3c) of the mouse model of inflammatory disease induced by LPS treatment when the compound according to one aspect of the present invention is treated with the mouse model. 3d) is a graph showing the expression levels of IL-1b and IL-6.
Figures 4a to 4e are graphs showing the expression levels of inflammatory cytokines and chemokines when a compound according to one aspect of the present invention is treated with intraperitoneal macrophages (PM) in which an inflammatory response is induced with LPS.
Figures 5a to 5d are graphs showing the expression levels of inflammatory cytokines and chemokines when a compound according to an aspect of the present invention is treated with bone marrow-derived macrophages (BMDMs) in which an inflammatory response is induced with LPS.
Figures 6a to 6f show the expression levels of IL-1b and IL-6 when treating intraperitoneal macrophages (PM) in which an inflammatory response is induced with a compound according to one aspect of the present invention with the ligands of toll-like receptor (TLR). This is a graph representing .
Figures 7a and 7b are graphs showing the expression levels of IL-1b and IL-6 when HeLa cells infected with Salmonella Typhimurium are treated with a compound according to one aspect of the present invention.
Figures 8a to 8c are graphs showing the expression levels of IL-1b, IL-6, and CXCL5 when the compound according to one aspect of the present invention is treated with Mycobacterium tuberculosis- infected bone marrow-derived macrophages (BMDM).
Figures 9a to 9c are graphs showing the expression levels of IL-1b, IL-6, and CXCL5 when the compound according to one aspect of the present invention is treated with intraperitoneal macrophages (PM) infected with Mycobacterium tuberculosis .
Figure 10 is a graph showing the expression level of TNF-α when a mouse model infected with BCG bacteria (bacille de Calmette-Guerin) is treated with a compound according to one aspect of the present invention.

Hereinafter, the present invention will be described in detail.

In one aspect, the present invention provides an anti-inflammatory composition comprising a compound represented by the following formula (1), a salt, stereoisomer, solvate or hydrate thereof as an active ingredient.

[Formula 1]

In Formula 1,

R ₁ and R ₂ are independently -(CH ₂ ) _n1 -R _a , n1 is an integer of 1 to 3, and R _a is unsubstituted or fluoro-substituted phenyl;

R ₃ is -(CH ₂ ) _n2 - or -O-CH ₂ -CH(OH)-CH ₂ -, and n2 is an integer of 1 to 3;

R ₄ is -CH ₂ -CH ₂ - or -CH ₂ -CH ₂ -O-CH ₂ -CH ₂ -.

In one aspect of the present invention, the compound may be selected from the group consisting of the following compounds 1 to 3:

1) 2-((3,4-diphenethoxybenzyl)amino)ethan-1-ol,

2) ( R )-1-(3,4-bis((4-fluorobenzyl)oxy)phenoxy)-3-((2-hydroxyethyl)amino)propan-2-ol, and

3) 2-(2-((3,4-bis(benzyloxy)benzyl)amino)ethoxy)ethan-1-ol.

In one aspect of the invention, a “salt” may be a “pharmaceutically acceptable salt,” and “pharmaceutically acceptable” means avoiding significant toxic effects when used in typical medicinal dosages, means that it is capable of being approved or approved by a government or equivalent regulatory agency for use in animals, more specifically in humans, or is listed in a pharmacopoeia or is recognized as being listed in other general pharmacopoeia.

In one aspect of the present invention, “pharmaceutically acceptable salt” means a salt according to one aspect of the present invention that is pharmaceutically acceptable and has the desired pharmacological activity of the parent compound. The salts are (1) formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, etc.; or acetic acid, propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, 3-(4-hydroxybenzoyl) Benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethane-disulfonic acid, 2-hydroxyethanesulfonic acid, benzenesulfonic acid, 4-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, 4-Toluenesulfonic acid, camphorsulfonic acid, 4-methylbicyclo [2,2,2]-oct-2-ene-1-carboxylic acid, glucoheptonic acid, 3-phenylpropionic acid, trimethylacetic acid, tert -acid addition salts formed with organic acids such as butylacetic acid, lauryl sulfate, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic acid, and muconic acid; or (2) a salt formed when an acidic proton present in the parent compound is replaced.

In one aspect of the invention, “stereoisomers” specifically include optical isomers (e.g., essentially pure enantiomers, essentially pure diastereomers, or mixtures thereof). rather, it includes conformational isomers (i.e., isomers that differ only in the angle of one or more chemical bonds) or geometric isomers (e.g., cis-trans isomers).

In one aspect of the present invention, "essentially pure", when used in relation to, for example, enantiomers or partial isomers, means that the specific compound, for example, the enantiomer or partial isomer, is about 90% or more, specifically about 90% or more. It means present at least 95%, more specifically at least about 97% or at least about 98%, even more specifically at least about 99%, and even more specifically at least about 99.5% (w/w).

In one aspect of the present invention, “hydrate” refers to a compound to which water is bound, and is a broad concept that includes embedded compounds in which there is no chemical bond between water and the compound.

In one aspect of the present invention, “solvate” refers to a higher-order compound formed between a solute molecule or ion and a solvent molecule or ion.

In one aspect of the invention, the composition increases survival in mouse models of inflammatory disease induced by lipopolysaccharide (LPS) or cecal ligation puncture (CLP), inflammatory cytokines such as IL-1b and IL-6, and CXCL5. And it has anti-inflammatory activity that suppresses the expression level of chemokines such as CXCL10, and is effective in preventing, alleviating, or treating inflammatory diseases such as sepsis (Experimental Examples 1 and 2). In addition, the composition has anti-inflammatory activity by suppressing the expression of inflammatory cytokines such as IL-1b and IL-6 even when treated with the composition in mouse models of inflammatory diseases such as autoimmune diseases induced by Toll-like receptor treatment. It has the effect of preventing, alleviating, or treating inflammatory diseases related to the receptor (Experimental Example 3). Furthermore, when the composition is treated with cells or mice infected with typhoid fever, Mycobacterium tuberculosis, and BCG bacteria, respectively, it acts as an anti-inflammatory agent that inhibits the expression levels of inflammatory cytokines such as IL-1b, IL-6, and TNF-α, and chemokines such as CXCL5. Because it is active, it has the effect of preventing, alleviating, or treating inflammatory diseases caused by bacterial infection (Experimental Example 4).

In one aspect of the invention, the inflammation may be caused by overexpression of inflammatory cytokines or chemokines. Specifically, the inflammatory cytokines are a group consisting of interleukin-1b (IL-1b), interleukin-6 (IL-6), and tumor necrosis factor alpha (TNF-α). It may be one or more selected from, but is not limited to. Additionally, specifically, the chemokine may be one or more selected from the group consisting of C-X-C motif chemokine 5 (CXCL5) and C-X-C motif chemokine 10 (CXCL10), but is not limited thereto. In addition, specifically, the overexpression of the inflammatory cytokine or chemokine is 5% or more, 10% or more, 15% or more, 20% or more, 25% or more, 30% or more. % or more, 35% or more, 40% or more, 45% or more, 50% or more, 55% or more, 60% or more, 65% or more, 70% or more, 75% or more, 80% or more, 85% or more, 90% or more , it may be an increase of more than 95% or more than 100%.

In one aspect of the present invention, the composition may have anti-inflammatory activity against inflammation caused by bacterial infection. Specifically, the composition may have anti-inflammatory activity against inflammation caused by gram-negative bacteria or gram-positive bacteria. More specifically, the gram-negative bacteria include Salmonella species, Yersinia species, Escherichia species, Chlamydia species, Xanthomonas species, and Erwinia species ( Erwinia species), Pseudomonas species ( Pseudomonas species), and Ralstonia species ( Ralstonia species) may be one or more selected from the group consisting of, more specifically, Salmonella typhimurium , Yersinia pestis , Yersinia pseudotuberculosis , Yersinia enterocolitica , Escherichia coli , Chlamydia species, Xanthomonas campestris, Pseudomonas syringae ( Pseudomonas syringae ) and Ralstonia solanacearum. More specifically, the composition has anti-inflammatory activity against inflammation caused by Salmonella typhimurium . It may be. Or more specifically, the Gram-positive bacteria include Mycobacterium species, Streptococcus species, Staphylococcus species, Enterococcus species, and Lactobacillus species. It may be one or more selected from the group consisting of, and more specifically, the composition may have anti-inflammatory activity against inflammation caused by one or more selected from the group consisting of Mycobacterium tuberculosis and Streptococcus pyogenes . . According to one embodiment of the present invention, the composition according to one embodiment of the present invention is applied to cells or mice infected with Salmonella typhimurium , Mycobacterium tuberculosis , or BCG bacteria (bacille de Calmette-Guerin). Alternatively, as the expression levels of inflammatory cytokines and chemokines in mice were reduced compared to the control group, it was found that there was an excellent anti-inflammatory effect on inflammation caused by infection with gram-negative or gram-positive bacteria (Experimental Example 4).

In one aspect of the present invention, the composition may inhibit the expression of inflammatory cytokines or chemokines. Specifically, the composition is selected from the group consisting of interleukin-1b (IL-1b), interleukin-6 (IL-6), and tumor necrosis factor alpha (TNF-α). It may be to suppress the expression of one or more inflammatory cytokines. More specifically, the composition increases the expression level of the inflammatory cytokines by 10% or more, 15% or more, 20% or more, 25% or more, 30% or more, 35% or more, 40% or more, 45% or more, It may be, but is not limited to, more than 50%, more than 55%, more than 60%, more than 65%, more than 70%, more than 75%, more than 80%, more than 85%, or more than 90%. Or, specifically, the composition may inhibit the expression of one or more chemokines selected from the group consisting of C-X-C motif chemokine 5 (CXCL5) and C-X-C motif chemokine 10 (CXCL10). there is. More specifically, the composition increases the expression level of the chemokine by 10% or more, 15% or more, 20% or more, 25% or more, 30% or more, 35% or more, 40% or more, 45% or more, 50% or more compared to the control group. It may be, but is not limited to, more than 55%, more than 60%, more than 65%, more than 70%, more than 75%, more than 80%, more than 85%, or more than 90%.

In one aspect of the present invention, the compound, its salt, stereoisomer, solvate or hydrate may be 0.001 to 1000 μM based on the total volume of the composition.

In one aspect of the present invention, the dosage of the composition may be 0.01 to 5000 mg/kg/day.

In another aspect, the present invention provides a composition for preventing, alleviating, or treating inflammatory diseases, comprising the compound represented by Formula 1, a salt, stereoisomer, solvate, or hydrate thereof as an active ingredient. The description of the compound, its salt, stereoisomer, solvate, hydrate, anti-inflammatory activity, bacteria, concentration, dosage, etc. is as described above.

In one aspect of the present invention, the inflammatory disease may be a disease caused by excessive production of inflammatory cytokines or chemokines, and the description of inflammatory cytokines and chemokines is as described above.

In one aspect of the present invention, the inflammatory disease may be a disease caused by bacterial infection. Specifically, diseases caused by bacterial infections include sepsis, tuberculosis, pneumonia, food poisoning, dysentery, Pseudomonas aeruginosa infection, impetigo, purulent disease, acute dermatitis, cystitis, vaginitis, meningitis, typhoid fever, paratyphoid fever, tetanus, Lyme disease, Rocky Mountain spotted fever, It may be, but is not limited to, one or more selected from the group consisting of cholera, leprosy, brucellosis, ehrlichiosis, Q fever, scarlet fever, listeriosis, botulism, leptospirosis, plague, typhus, bacteremia, endocarditis, and enteritis. In addition, the disease caused by the bacterial infection may be a disease caused by infection by gram-negative bacteria or gram-positive bacteria, and the description of gram-negative bacteria and gram-positive bacteria is as described above.

In one aspect of the invention, the inflammatory disease may be an autoimmune disease, and the autoimmune disease may be a toll-like receptor (TLR)-related disease. Specifically, the autoimmune disease is an autoimmune disease caused by excessive inflammation, such as dermatitis, allergy, atopy, asthma, conjunctivitis, periodontitis, rhinitis, otitis media, pharyngitis, tonsillitis, pneumonia, stomach ulcer, gastritis, Crohn's disease, Behcet's disease, Colitis, hemorrhoids, gout, ankylosing spondylitis, rheumatic fever, lupus, fibromyalgia, psoriatic arthritis, osteoarthritis, rheumatoid arthritis, periarthritis, tendinitis, tenosynovitis, peritendinitis, myositis, hepatitis, cystitis, nephritis, Sjogren's syndrome ), multiple sclerosis, and acute and chronic inflammatory diseases, but is not limited thereto.

In one aspect of the present invention, the composition may inhibit the expression of inflammatory cytokines or chemokines. Specifically, the composition is selected from the group consisting of interleukin-1b (IL-1b), interleukin-6 (IL-6), and tumor necrosis factor alpha (TNF-α). It may be to suppress the expression of one or more inflammatory cytokines. More specifically, the composition increases the expression level of the inflammatory cytokines by 10% or more, 15% or more, 20% or more, 25% or more, 30% or more, 35% or more, 40% or more, 45% or more, It may be, but is not limited to, more than 50%, more than 55%, more than 60%, more than 65%, more than 70%, more than 75%, more than 80%, more than 85%, or more than 90%. Or, specifically, the composition may inhibit the expression of one or more chemokines selected from the group consisting of C-X-C motif chemokine 5 (CXCL5) and C-X-C motif chemokine 10 (CXCL10). there is. More specifically, the composition increases the expression level of the chemokine by 10% or more, 15% or more, 20% or more, 25% or more, 30% or more, 35% or more, 40% or more, 45% or more, 50% or more compared to the control group. It may be, but is not limited to, more than 55%, more than 60%, more than 65%, more than 70%, more than 75%, more than 80%, more than 85%, or more than 90%.

In one aspect of the present invention, the compound, its salt, stereoisomer, solvate or hydrate may be 0.001 to 1000 μM based on the total volume of the composition.

In one aspect of the present invention, the dosage of the composition may be 0.01 to 5000 mg/kg/day.

In another aspect, the composition according to one aspect of the present invention may be a pharmaceutical, food, or cosmetic composition.

As an example, the food composition includes a health functional food composition, and the health functional food composition may be for improving, alleviating, or preventing inflammatory diseases. Specifically, the inflammatory disease may be a disease caused by a bacterial infection, and more specifically, the disease caused by the bacterial infection is sepsis, tuberculosis, pneumonia, food poisoning, dysentery, Pseudomonas aeruginosa infection, impetigo, purulent disease, acute dermatitis, cystitis, and vaginitis. , meningitis, typhoid fever, paratyphoid fever, tetanus, Lyme disease, Rocky Mountain spotted fever, cholera, leprosy, brucellosis, ehrlichiosis, Q fever, scarlet fever, listeriosis, botulism, leptospirosis, plague, typhus, bacteremia, endocarditis and enteritis. It may be one or more selected from the group consisting of: In addition, the disease caused by the bacterial infection may be a disease caused by infection by gram-negative bacteria or gram-positive bacteria, and the description of gram-negative bacteria and gram-positive bacteria is as described above. Alternatively, the inflammatory disease may be an autoimmune disease, and specifically, the autoimmune disease may be a toll-like receptor (TLR)-related disease. More specifically, the autoimmune disease may include dermatitis, allergy, Atopy, asthma, conjunctivitis, periodontitis, rhinitis, otitis media, pharyngitis, tonsillitis, pneumonia, stomach ulcer, gastritis, Crohn's disease, Behcet's disease, colitis, hemorrhoids, gout, ankylosing spondylitis, rheumatic fever, lupus, fibromyalgia, psoriatic arthritis, It may be one or more diseases selected from the group consisting of osteoarthritis, rheumatoid arthritis, periarthritis, tendinitis, tenosynovitis, peritendinitis, myositis, hepatitis, cystitis, nephritis, Sjogren's syndrome, multiple sclerosis, and acute and chronic inflammatory diseases. . Alternatively, the health functional food composition may be used to improve, alleviate, or prevent inflammation.

The health food functional composition includes various types of food additives or functional foods. It can be processed into leached tea, liquid tea, beverage, fermented milk, cheese, yogurt, juice, probiotics and health supplements containing the above composition, and can be used in the form of various other food additives.

As an example, the cosmetic composition may be provided in any formulation suitable for topical application. For example, it may be provided in the form of a solution, an emulsion obtained by dispersing an oil phase in an aqueous phase, an emulsion obtained by dispersing an aqueous phase in an oil phase, a suspension, solid, gel, powder, paste, foam, or aerosol composition. Compositions of this dosage form can be prepared according to conventional methods in the art.

As an example, the cosmetic composition may contain, in addition to the above-mentioned substances, other ingredients that can preferably have a synergistic effect on the main effect within a range that does not impair the main effect. The cosmetic composition according to the present invention may contain a substance selected from the group consisting of vitamins, high molecular weight peptides, high molecular weight polysaccharides, and sphingolipids. In addition, the cosmetic composition may include, as an example, a moisturizer, an emollient, a surfactant, an ultraviolet absorber, a preservative, a disinfectant, an antioxidant, a pH adjuster, an organic and inorganic pigment, a fragrance, a cooling agent, or an antiperspirant. You can. The mixing amount of the above ingredients can be easily selected by a person skilled in the art within the range that does not impair the purpose and effect of the present invention, and the mixing amount may be 0.01 to 5% by weight, specifically 0.01 to 3% by weight, based on the total weight of the composition. there is.

As an example, the pharmaceutical composition may be used for preventing, alleviating, or treating inflammatory diseases. Specifically, the inflammatory disease may be a disease caused by a bacterial infection, and more specifically, the disease caused by the bacterial infection is sepsis, tuberculosis, pneumonia, food poisoning, dysentery, Pseudomonas aeruginosa infection, impetigo, purulent disease, acute dermatitis, cystitis, and vaginitis. , meningitis, typhoid fever, paratyphoid fever, tetanus, Lyme disease, Rocky Mountain spotted fever, cholera, leprosy, brucellosis, ehrlichiosis, Q fever, scarlet fever, listeriosis, botulism, leptospirosis, plague, typhus, bacteremia, endocarditis and enteritis. It may be one or more selected from the group consisting of: In addition, the disease caused by the bacterial infection may be a disease caused by infection by gram-negative bacteria or gram-positive bacteria, and the description of gram-negative bacteria and gram-positive bacteria is as described above. Alternatively, the inflammatory disease may be an autoimmune disease, and specifically, the autoimmune disease may be a toll-like receptor (TLR)-related disease. More specifically, the autoimmune disease may include dermatitis, allergy, Atopy, asthma, conjunctivitis, periodontitis, rhinitis, otitis media, pharyngitis, tonsillitis, pneumonia, stomach ulcer, gastritis, Crohn's disease, Behcet's disease, colitis, hemorrhoids, gout, ankylosing spondylitis, rheumatic fever, lupus, fibromyalgia, psoriatic arthritis, It may be one or more diseases selected from the group consisting of osteoarthritis, rheumatoid arthritis, periarthritis, tendinitis, tenosynovitis, peritendinitis, myositis, hepatitis, cystitis, nephritis, Sjogren's syndrome, multiple sclerosis, and acute and chronic inflammatory diseases. . Alternatively, the pharmaceutical composition may be used to prevent, alleviate, or treat inflammation.

Additionally, as an example, the pharmaceutical composition may be provided in any formulation suitable for topical application. For example, it can be administered orally, transdermally, intravenously, intramuscularly, or by subcutaneous injection. As an example, the pharmaceutical composition may be an injection, a solution for external use on the skin, a suspension, an emulsion, a gel, a patch, or a spray, but is not limited thereto. The formulation can be easily prepared according to conventional methods in the field, and contains surfactants, excipients, wetting agents, emulsification accelerators, suspending agents, salts or buffers for adjusting osmotic pressure, colorants, flavorings, stabilizers, preservatives, preservatives or Other commonly used supplements can be used appropriately.

The active ingredient of the pharmaceutical composition according to an embodiment of the present invention will vary depending on the subject's age, gender, weight, pathological condition and severity, route of administration, or the judgment of the prescriber.

From another perspective, the present invention may relate to an anti-inflammatory method comprising administering a composition containing the compound represented by Formula 1, a salt, stereoisomer, solvate or hydrate thereof to an individual in need of anti-inflammatory treatment. In one aspect of the present invention, administration of the method may be performed according to the administration method and administration dose described herein.

From another aspect, the present invention relates to an inflammatory disease comprising administering a composition containing the compound represented by Formula 1, a salt, stereoisomer, solvate or hydrate thereof to an individual in need of prevention, alleviation or treatment of an inflammatory disease. It may relate to a method of preventing, alleviating, or treating inflammation caused by . Specifically, the inflammatory disease may be caused by a bacterial infection, and more specifically, the inflammatory disease may include sepsis, tuberculosis, pneumonia, food poisoning, dysentery, Pseudomonas aeruginosa infection, impetigo, purulent disease, acute dermatitis, cystitis, vaginitis, meningitis, typhoid fever, One selected from the group consisting of paratyphus, tetanus, Lyme disease, Rocky Mountain spotted fever, cholera, leprosy, brucellosis, ehrlichiosis, Q fever, scarlet fever, listeriosis, botulism, leptospirosis, plague, typhus, bacteremia, endocarditis and enteritis. It could be more than that. Alternatively, specifically, the inflammatory disease may be an autoimmune disease, and more specifically, the autoimmune disease may include dermatitis, allergy, atopy, asthma, conjunctivitis, periodontitis, rhinitis, otitis media, pharyngitis, tonsillitis, pneumonia, gastric ulcer, gastritis, and Crohn's disease. Disease, Behcet's disease, colitis, hemorrhoids, gout, ankylosing spondylitis, rheumatic fever, lupus, fibromyalgia, psoriatic arthritis, osteoarthritis, rheumatoid arthritis, periarthritis, tendinitis, tenosynovitis, peritendinitis, myositis, hepatitis, cystitis, nephritis, It may be one or more selected from the group consisting of Sjogren's syndrome, multiple sclerosis, and acute and chronic inflammatory diseases. In one aspect of the present invention, administration of the method may be performed according to the administration method and administration dose described herein.

From another perspective, the present invention may relate to the use of the compound represented by Formula 1, its salt, stereoisomer, solvate or hydrate for preparing an anti-inflammatory pharmaceutical composition.

From another perspective, the present invention may relate to the use of the compound represented by Formula 1, its salt, stereoisomer, solvate or hydrate for producing an anti-inflammatory food composition.

From another perspective, the present invention may relate to the use of the compound represented by Formula 1, its salt, stereoisomer, solvate or hydrate for producing an anti-inflammatory cosmetic composition.

From another perspective, the present invention may relate to the use of the compound represented by Formula 1, a salt, stereoisomer, solvate or hydrate thereof, for preparing a pharmaceutical composition for preventing, alleviating or treating inflammatory diseases.

From another perspective, the present invention may relate to the use of the compound represented by Formula 1, its salt, stereoisomer, solvate or hydrate for producing a food composition for improving, preventing or alleviating inflammatory diseases.

From another perspective, the present invention may relate to the use of the compound represented by Formula 1, its salt, stereoisomer, solvate or hydrate for producing a cosmetic composition for improving, preventing or alleviating inflammatory diseases.

From another perspective, the present invention may relate to the anti-inflammatory use of a composition containing the compound represented by Formula 1, a salt, stereoisomer, solvate or hydrate thereof as an active ingredient.

From another perspective, the present invention may relate to the use of a composition comprising the compound represented by Formula 1, a salt, stereoisomer, solvate or hydrate thereof as an active ingredient for the improvement, prevention, alleviation or treatment of inflammatory diseases. .

From another perspective, the present invention may relate to a composition containing the compound represented by Formula 1, a salt, stereoisomer, solvate or hydrate thereof as an active ingredient for anti-inflammatory purposes.

From another perspective, the present invention may relate to a composition comprising the compound represented by Formula 1, a salt, stereoisomer, solvate or hydrate thereof as an active ingredient for the purpose of improving, preventing, alleviating or treating inflammatory diseases. .

Hereinafter, the configuration and effects of the present invention will be described in more detail through examples, production examples, and experimental examples. However, the following examples, preparation examples, and experimental examples are provided only for illustrative purposes to aid understanding of the present invention, and the scope and scope of the present invention are not limited thereto.

[Preparation example] Preparation of compounds

Example number Chemical formula name constitutional formula Example 1 Compound A 2-((3,4-diphenethoxybenzyl)amino)ethan-1-ol Example 2 Compound B ( R )-1-(3,4-bis((4-fluorobenzyl)oxy)phenoxy)-3-((2-hydroxyethyl)amino)propan-2-ol Example 3 Compound C 2-(2-((3,4-bis(benzyloxy)benzyl)amino)ethoxy)ethane-1-ol

Example 1. Preparation of 2-((3,4-diphenethoxybenzyl)amino)ethan-1-ol (Compound A)

[Scheme 1]

Step 1) Synthesis of 3,4-diphenethoxybenzaldehyde (A-1)

3,4-dihydroxybenzaldehyde (20.0 g, 140 mmol) was dissolved in anhydrous THF (200 mL), then 2-phenylethanol (40.7 g, 330 mmol), triphenylphosphine (PPh ₃ , 102 g, 390 mmol) and diisopropyl azodicarboxylate (DIAD, 79.0 g, 390 mmol) were added slowly to the reaction at 0 °C, and then stirred at 65 °C for 12 hours. When the reaction is completed, the reactant is cooled to room temperature, purified water (100 ml) is added, extracted three times with ethyl acetate (EA, 100 ml), and the separated organic layer is washed again with saturated aqueous sodium chloride solution (50 ml). . Next, anhydrous sodium sulfate (Na ₂ SO ₄ ) was added to the organic layer, stirred, and then filtered under reduced pressure. The filtered solution was concentrated and purified through column chromatography to obtain 3,4-diphenethoxybenzaldehyde ( A-1 , 46.0 g, yield: 92%) in the form of a yellow oil.

¹ H NMR (DMSO-d ₆ , 400 MHz): δ 9.81 (s, 1H), 7.53 (d, 1H), 7.51-7.17 (m, 12H), 4.29-4.20 (m, 4H), 3.09-3.04 ( m, 4H).

Step 2) Synthesis of 2-((3,4-diphenethoxybenzyl)amino)ethan-1-ol (Compound A)

3,4-Diphenethoxybenzaldehyde ( A-1 , 20.0 g, 57.8 mmol) was dissolved in methanol (200 mL), then 2-aminoethanol (3.90 g, 63.9 mmol) was added and stirred at 65°C for 2 hours. do. The reaction was then cooled to room temperature, sodium borohydride (NaBH ₄ , 2.40 g, 36.9 mmol) was added, and stirred at 50°C for 12 hours. When the reaction is completed, the reactant is cooled to room temperature, purified water is added, extracted three times with ethyl acetate (EA, 100 mL), and the separated organic layer is washed again with saturated aqueous sodium chloride solution. Next, anhydrous sodium sulfate (Na ₂ SO ₄ ) was added to the organic layer, stirred, and then filtered under reduced pressure. The filtered solution was concentrated and purified by column chromatography (DCM/MeOH = 20:1) to produce 2-((3,4-diphenethoxybenzyl)amino)ethan-1-ol ( Compound A) as a yellow solid. , 17.0 g, yield: 75%) was obtained.

¹ H NMR (DMSO_d ₆ , 400 MHz): δ 7.21-7.35 (m, 10 H), 6.78-6.95 (m, 3 H), 4.43-4.43 (br, 1 H), 4.09-4.14 (m, 4 H) ), 3.61 (s, 2 H), 3.44-3.47 (m, 2 H), 2.99-3.04 (m, 4 H), 2.53-2.56 (m, 2 H);

LC-MS Calcd.: 391, MS Found: 392 [MS+1].

Example 2. ( R )-1-(3,4-bis((4-fluorobenzyl)oxy)phenoxy)-3-((2-hydroxyethyl)amino)propan-2-ol (Compound B) Preparation

[Scheme 2]

Step 1) Synthesis of 3,4-bis((4-fluorobenzyl)oxy)phenol (B-2)

Dichloromethane (15 ml) was dissolved in 3,4-bis((4-fluorobenzyl)oxy)benzaldehyde ( B-1 , 1.00 g, 3.0 mmol, 1 equivalent), and then mCPBA (0.78 g, 4.5 mmol, 1.5 equivalent) was added to the reaction and stirred at room temperature for 4 hours. The reaction was diluted with ethyl acetate, washed with saturated aqueous sodium carbonate solution, and the organic layer was separated. The organic layer was washed with an aqueous sodium chloride solution, dehydrated with anhydrous sodium sulfate, and filtered under reduced pressure. After concentrating the filtered solution, it was dissolved again in methanol (10ml), then 6N NaOH was added and stirred at room temperature for 30 minutes. After adding 4N HCl solution to the reaction, it was stirred for an additional 30 minutes. The reaction was diluted with ethyl acetate (50 ml), washed with salt water, and then dehydrated with anhydrous sodium sulfate and filtered under reduced pressure. The filtered solution was concentrated and purified by column chromatography (hexane/ethyl acetate ratio = 7/3) to produce 3,4-bis((4-fluorobenzyl)oxy)phenol ( B-2 , 0.87) as a white solid. g, yield 90%) was obtained.

¹ H NMR (DMSO-d ₆ , 400 MHz) δ (ppm) 7.49-7.42 (m, 4H), 7.24-7.15 (m, 4H), 6.82 (d, 1H), 6.49 (s, 1H), 6.27- 3.25 (m, 1H), 5.04 (s, 2H), 4.95 (s, 2H).

Step 2) ( R )-2-((3,4-bis((4-fluorobenzyl)oxy)phenoxy)methyl)oxirane (B-3) synthesis

3,4-bis((4-fluorobenzyl)oxy)phenol ( B-2 , 306 mg, 1.0 mmol) was diluted in ethanol (10 ml), then mixed with KOH aqueous solution (KOH 66 mg, 1.2 mmol, 1 ml) and ( R )-2-(chloromethyl)oxirane (410ul, 5.0 mmol) was sequentially added. After the reaction was stirred at room temperature for 5 hours, the organic solvent was removed under reduced pressure. The concentrated reaction product was diluted again with ethyl acetate, washed with water and then with salt water, and the extracted organic layer was dehydrated with anhydrous sodium sulfate and then filtered under reduced pressure. The filtered organic layer was concentrated and purified by column chromatography to produce ( R )-2-((3,4-bis((4-fluorobenzyl)oxy)phenoxy)methyl)oxirane ( B-3 , 297mg, yield 82%) was obtained.

¹ H NMR (DMSO-d ₆ , 400 MHz) δ (ppm) 7.51-7.43 (m, 4H), 7.25-7.17 (m, 4H), 6.94 (d, 1H), 6.73 (s, 1H), 6.47- 6.44 (m, 1H), 5.11 (s, 1H), 5.00 (s, 1H), 4.26-4.23 (m, 1H), 3.78-3.73 (m, 1H), 3.31-3.28 (m, 1H), 2.84- 2.82(m, 1H), 2.70-2.67 (m, 1H).

Step 3) ( R )-1-(3,4-bis((4-fluorobenzyl)oxy)phenoxy)-3-((2-hydroxyethyl)amino)propan-2-ol (Compound B) Synthesis

( R )-2-((3,4-bis((4-fluorobenzyl)oxy)phenoxy)methyl)oxirane ( B-3 , 9 mg, 25 nmol) was diluted in absolute ethanol (1 ml). Then, 2-aminoethane-1-ol (7.6 μL, 125 nmol) was added and stirred at room temperature for 4 hours. After confirming the reaction by TLC, the reaction solvent was concentrated under reduced pressure, water was added to the concentrated reaction product, and extracted with dichloromethane ((3Υ 5 mL). The extracted organic layer was dehydrated with anhydrous sodium sulfate and then filtered under reduced pressure. The filtered organic layer Concentrated and purified by column chromatography (dichloromethane:methanol=19:1) ( R )-1-(3,4-(bis((4-fluorobenzyl)oxy)phenoxy)-3-( (2-Hydroxyethyl)amino)propan-2-ol ( Compound B , 9.0 mg, yield 85%) was obtained.

¹ H NMR (CDCl ₃ , 400 MHz) δ (ppm) 7.40-7.33 (m, 4H), 7.07-7.00 (m, 4H), 6.84 (d, 1H), 6.58 (s, 1H), 6.41-6.38 ( m, 1H), 5.05 (s, 2H), 5.00 (s, 2H), 4.06 (br s, 1H), 3.91 (m, 2H), 3.69 (m, 2H), 2.89-2.76 (m, 4H); LC-MS Calcd.: 459.4, MS Found: 460.9 [MS+1].

Example 3. Preparation of 2-(2-((3,4-bis(benzyloxy)benzyl)amino)ethoxy)ethan-1-ol (Compound C)

[Scheme 3]

Step 1) Synthesis of 3,4-bis(benzyloxy)benzaldehyde (E-1)

After dissolving 3,4-dihydroxybenzaldehyde (0.50 g, 3.62 mmol) in anhydrous DMF (5 mL), K ₂ CO ₃ (1.50 g, 10.86 mmol) was added, followed by benzyl bromide (0.92 mL, 7.96 mmol). ) was slowly added to the reaction and stirred at 60°C for 4 hours. When the reaction was completed, the reactant was cooled to room temperature, diluted in purified water, and extracted twice with diethyl ether (50 mL). The organic layer was washed twice with purified water (50 ml) and then again with saturated aqueous sodium chloride solution (50 ml). Next, anhydrous sodium sulfate was added to the organic layer, stirred, and then filtered under reduced pressure. The filtered solution was concentrated and purified through column chromatography to obtain 3,4-bis(benzyloxy)benzaldehyde ( E-1 , 1.04 g, yield 90%).

¹ H NMR (CDCl ₃ , 300 MHz) δ 9.81 (s, 1 H), 7.49-7.31 (m, 12 H), 7.04 (d, J = 8.3 Hz, 1 H), 5.27 (s, 2 H), 5.22 (s, 2H); ESIMS m/z: 319.33 [M+H] ⁺ .

Step 2) Synthesis of 2-(2-((3,4-bis(benzyloxy)benzyl)amino)ethoxy)ethan-1-ol (Compound C)

3,4-bis(benzyloxy)benzaldehyde ( E-1 , 33 mg, 0.1 mmol) was dissolved in absolute ethanol (5 ml), then 2-(2-aminoethoxy)ethan-1-ol (50 μL) , 0.5 mmol) was added and stirred under reflux for 4 hours. After cooling to room temperature, the production of imine was confirmed by TLC, then NaBH ₄ (7.6 mg, 0.2 mmol) was added, and the mixture was stirred for an additional 4 hours. The reaction solvent was concentrated under reduced pressure, water was added, and the mixture was extracted with ethyl acetate. The separated organic layer was washed once more with salt water, separated, dehydrated with anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The concentrated mixture was purified through column chromatography to produce 2-(2-((3,4-bis(benzyloxy)benzyl)amino)ethoxy)ethoxy)ethan-1-ol ( Compound 3 , yield 78%) as a white solid. got it

¹ H NMR (400 MHz, CDCl ₃ ) δ (ppm) 7.48 (d, 2H), 7.27-7.41 (m, 9H), 6.99-7.02 (m, 1H), 6.84 (d, 1H), 5.25 (s, 2H), 5.09 (s, 2H), 4.06 (s, 2H), 3.65-3.70 (m, 4H), 3.46-3.48 (m, 2H), 2.87 (t, 2H); Mass Calcd.:407.5; MS Found: 408.9 [MS+1].

[Experimental Example 1] Confirmation of anti-inflammatory effect in inflammatory disease mouse model

The following experiment was performed to confirm the effect of extending survival and suppressing inflammatory cytokine expression when treated with Examples 1 and 2 prepared in the above Preparation Example on an inflammatory disease mouse model.

[Experimental Example 1-1] Changes in survival rate in a mouse model of inflammatory disease induced by lipopolysaccharide (LPS)

LPS was administered to 8-week-old mice by intraperitoneal injection at 28 mg/kg, and Examples 1 and 2 were administered by intraperitoneal injection at 20 mg/kg each 2 hours before and 1 hour after LPS administration. Subsequently, the mouse survival rate was measured every 12 hours from the time of LPS administration until 72 hours, and the results are shown in Figures 1a and 1b. At this time, vehicle-administered mice were used as a control group instead of Examples 1 and 2.

As shown in Figures 1a and 1b, due to LPS administration, the survival rate of the control group decreased to 50 to 70% after 12 to 24 hours, and the survival rate decreased to 25% or less after 36 hours of administration. On the other hand, when administered in Example 2, the survival rate was maintained at more than 75% even 72 hours after LPS administration, and the survival rate in Example 1 also increased compared to the control group.

Through this, it can be seen that the composition according to one aspect of the present invention has an excellent effect of increasing survival rate in an LPS-induced inflammatory disease model.

[Experimental Example 1-2] Changes in survival rate in mouse model of inflammatory disease induced by cecal ligation puncture (CLP)

After an 8-week-old mouse was anesthetized with tribromoethanol, the cecum was exposed through a laparotomy, and the cecum was tightly tied using a silk suture at 1 cm from the appendix. After puncturing the patient twice using a 21-gauge needle, the excrement from the puncture site was removed and returned to the peritoneum. The laparotomy site was closed with silk sutures. 20 mg/kg of Example 2 was administered by intraperitoneal injection 2 hours before and 12 hours after suturing. Subsequently, the mouse survival rate was measured every 12 hours from the time of suturing until 72 hours, and the results are shown in Figure 2. At this time, vehicle-administered mice were used as a control group instead of Example 2.

As shown in Figure 2, due to CLP, the survival rate in the control group decreased to 50% after 36 hours, while the survival rate was extended to more than 75% even after 72 hours when administered in Example 2.

Through this, it can be seen that the composition according to one aspect of the present invention has an excellent effect of increasing survival rate in a CLP-induced inflammatory disease model.

[Experimental Example 1-3] Changes in inflammatory cytokine expression in a LPS-induced inflammatory disease mouse model

LPS was administered to 8-week-old mice by intraperitoneal injection at 14 mg/kg, and Example 2 at 20 mg/kg was administered by intraperitoneal injection 2 hours before and 1 hour after LPS administration. Subsequently, the lungs and spleens of the mice were removed 6 hours after administration of 20 mg/kg. The extracted lung and spleen were homogenized in 1 ml of phosphate-buffered saline and centrifuged to collect lung and spleen tissues. The mRNA of inflammatory cytokines (IL-1b, IL-6, and CXCL5) expressed from the collected tissues was measured by relative quantitation through real-time polymerase chain reaction, and the results of the experiment are shown in Figure 3. At this time, vehicle-administered mice were used as a control instead of Example 2.

As shown in Figure 3, when treated with the compound of Example 2, it was confirmed that the expression levels of inflammatory cytokines IL-1b and IL-6 in the LPS-induced inflammatory disease mouse model were reduced by about half compared to the control group. there was.

Through this, it was found that the composition according to one aspect of the present invention has an anti-inflammatory effect and is effective in preventing or treating inflammatory diseases such as sepsis.

[Experimental Example 2] Confirmation of anti-inflammatory effect in intraperitoneal macrophages and bone marrow-derived macrophages of LPS-induced inflammatory disease mouse model

When Examples 1 and 2 prepared in the above Preparation Examples were treated with an LPS-induced inflammatory disease mouse model, the expression of inflammatory cytokines and chemokines was suppressed in intraperitoneal macrophages (PM) and bone marrow-derived macrophages (BMDM), respectively. To confirm the effect, the following experiment was performed.

[Experimental Example 2-1] Changes in expression levels of inflammatory cytokines and chemokines in intraperitoneal macrophages

To collect intraperitoneal macrophages (PM), 1 ml of phosphate-buffered saline containing 3% thioglycolate was administered to 8-week-old mice by intraperitoneal injection. Three days after administration, the mouse was sacrificed and peritoneal fluid was collected by intraperitoneally injecting 6 ml of pre-cooled phosphate-buffered saline containing 3% fetal bovine serum. Subsequently, the cell suspension was centrifuged, the supernatant was removed, and the obtained intraperitoneal macrophages (PM) were counted. Cell culture medium containing 10% fetal bovine serum and 1% streptomycin/penicillin was used, and 5% carbon dioxide and 37 Cultured for 1 day in an incubator maintained at °C. Afterwards, 2 hours before cell treatment, the cell culture medium was removed and replaced with cell culture medium containing 5% fetal calf serum, and Examples 1 and 2 were added to 2 × 10 ⁵ cells at 1, 5, and 10 μM for each concentration. After pretreatment for 1 hour and treatment with 10 ng/ml LPS for 6 hours, cells were collected. Real-time polymerase chain reaction was performed to measure the relative quantity of mRNA of inflammatory cytokines and chemokines expressed intracellularly from the collected cells, and the results of the experiment are shown in Figure 4. At this time, mice treated with LPS but not in Examples 1 and 2 were used as a control group (LPS in FIG. 4).

As shown in Figure 4, when treated in Example 1 or 2, among the intracellular inflammatory cytokines induced by LPS, tumor necrosis factor-alpha (TNF-α) and hypoxia-inducible factor 1-alpha (HIF1-α) There was no difference in the mRNA expression level compared to the control group, but the mRNA expression levels of IL-1b, IL-6, and CXCL10 were confirmed to be significantly decreased in a concentration-dependent manner compared to the control group.

[Experimental Example 2-2] Changes in inflammatory cytokine expression in bone marrow-derived macrophages

An 8-week-old mouse was sacrificed, and the bone marrow of the tibia and femur was collected using pre-cooled cell culture medium. The cell suspension was centrifuged, the supernatant was removed, the obtained bone marrow-derived macrophages (BMDM) were counted, and 10% fetal bovine serum was added. , 1% streptomycin/penicillin, and 25 ng/ml of macrophage colony-stimulating factor were used to culture the cells for 4 days in an incubator maintained at 5% carbon dioxide and 37°C. Afterwards, 2 hours before cell treatment, the cell culture medium was removed and replaced with cell culture medium containing 5% fetal bovine serum, and each of Examples 1 and 2 was used at 1, 5, and 10 μM for each concentration, 2 × 10 ⁵ cells. for 1 hour and LPS at 10 ng/ml for 6 hours, and then the cells were collected. Real-time polymerase chain reaction was performed to measure the relative quantity of inflammatory cytokine mRNA expressed intracellularly from the collected cells, and the results of the experiment are shown in Figure 5. At this time, mice treated with LPS but not in Examples 1 and 2 were used as a control group (LPS in Figure 5).

As shown in Figure 5, when treated in Example 1 or 2, among the intracellular inflammatory cytokines induced by LPS, tumor necrosis factor-alpha (TNF-α) and hypoxia-inducible factor 1-alpha (HIF1-α) There was no difference in the mRNA expression level compared to the control group, but the mRNA expression levels of IL-1b and IL-6 were confirmed to be significantly decreased in a concentration-dependent manner or in the highest concentration treatment group compared to the control group.

Through this, it was found that the composition according to one aspect of the present invention has an anti-inflammatory effect of suppressing inflammatory cytokines and chemokines, thereby preventing or treating inflammatory diseases such as sepsis.

[Experimental Example 3] Measurement of inflammatory cytokine expression in a mouse model of inflammatory disease induced by treatment with a toll-like receptor (TLR) ligand

To confirm the efficacy of Example 2 prepared in the above Preparation Example in suppressing inflammatory cytokines and chemokines, a real-time polymerase chain reaction experiment was performed as follows.

Specifically, in the same manner as in Experimental Examples 2-1 and 2-2 ^{, five} 2Х10 each were treated with Example 2 at 10 μM for 1 hour in each of intraperitoneal macrophages (PM) and bone marrow-derived macrophages (BMDM). 10 μl/ml of zymosan (Zym), a ligand for similar receptors, 20 μg/ml of polyinosine-polycytidylic acid (Poly I:C), and 100 ng/ml of Pam3CSK4 (P3C4) at 3 , cells were collected after treatment for 6 and 18 hours. Real-time polymerase chain reaction was performed to measure the relative quantity of mRNA of inflammatory cytokines and chemokines expressed within cells from the collected cells, and the results of the experiment are shown in Figure 6. At this time, instead of Example 2, mice treated only with toll-like receptor ligands were used as a control group (Zym, Poly I:C, P3C4 in Figure 6).

As shown in Figure 6, when treated in Example 2, the mRNA expression levels of IL-1b and IL-6 among the intracellular inflammatory cytokines induced by the ligand of the toll-like receptor were significantly higher compared to the control group. It was confirmed that there was a decrease.

Through this, it was found that Example 2 according to one aspect of the present invention exhibited an inflammatory response inhibition effect and was effective in preventing or treating inflammatory diseases such as sepsis.

[Experimental Example 4] Confirmation of effect in inflammatory disease model caused by bacterial infection

The following experiment was performed to confirm the effect of suppressing the expression of inflammatory cytokines and chemokines when treated with Example 3 prepared in the above Preparation Example on an inflammatory disease model caused by bacterial infection.

[Experimental Example 4-1] Typhoid fever ( Salmonella Typhimurium ) Changes in cytokine expression level in infected HeLa cell model

HeLa cells were cultured using DMEM medium containing 10% FBS and 1% streptomycin/penicillin in an incubator maintained at 5% carbon dioxide and 37°C. Cells were dispensed into a 6-well plate and cultured for more than 12 hours to ensure that the cells were completely attached to the plate surface. Afterwards, 5x10 ⁴ cell lines were infected with Salmonella Typhimurium at an MOI of 10 for 30 minutes, and then treated with the compound of Example 3 for 6 hours, and then the cells were collected. Quantitative changes in mRNA of intracellularly expressed cytokines from the collected cells were measured by RT-qPCR, and the results of the experiment are shown in Figure 7. At this time, a group not infected with Typhoid fever bacteria (UI) and a group infected with Typhoid fever bacteria but not treated with Example 3 (grey graph of Sal in FIG. 7) were used as control groups.

As shown in Figure 7, when treated with the compound of Example 3, the expression levels of cytokines IL-1b and IL-6 in HeLa cells infected with Typhoid fever decreased by about half compared to the control group.

[Experimental Example 4-2] Mycobacterium tuberculosis ( Mycobacterium tuberculosis ) Changes in inflammatory cytokine and chemokine expression levels in infected BMDM cell model

An 8-week-old mouse was sacrificed, and the bone marrow of the tibia and femur was collected using pre-cooled cell culture medium. The cell suspension was centrifuged, the supernatant was removed, the obtained bone marrow-derived macrophages (BMDM) were counted, and 10% fetal bovine serum was added. , 1% streptomycin/penicillin, and 25 ng/ml of macrophage colony-stimulating factor were used and cultured for 4 days in an incubator maintained at 5% carbon dioxide and 37°C. Afterwards, 2 hours before cell infection, the cell culture medium was removed and replaced with cell culture medium containing 5% fetal bovine serum, and 2 × 10 ⁵ cells were infected with Tubercle bacillus at an MOI of 5 for 2 hours. Subsequently, the cells were washed twice with preheated phosphate-buffered saline, exchanged with cell culture medium containing 5% fetal bovine serum, and treated with 10 μM of Example 3 for 6 hours, and then collected. Real-time polymerase chain reaction was performed to measure the relative quantity of mRNA of inflammatory cytokines and chemokines expressed within cells from the collected cells, and the results of the experiment are shown in Figure 8. At this time, a group not infected with Mycobacterium tuberculosis (UI) and a group infected with Mycobacterium tuberculosis but not treated with Example 3 (gray graph of Mtb in Figure 8) were used as control groups.

As shown in Figure 8, when treated in Example 3, the mRNA expression levels of IL-1b, IL-6, and CXCL5 among inflammatory cytokines and chemokines in macrophages infected with Mycobacterium tuberculosis were compared to the control group. Example 3 It was confirmed that there was a significant decrease in the treatment group.

[Experimental Example 4-3] Mycobacterium tuberculosis ( Mycobacterium tuberculosis ) Changes in inflammatory cytokine and chemokine expression levels in infected PM cell model

To collect intraperitoneal macrophages (PM), 1 ml of phosphate-buffered saline containing 3% thioglycolate was administered to 8-week-old mice by intraperitoneal injection. Three days after administration, the mouse was sacrificed and peritoneal fluid was collected by intraperitoneally injecting 6 ml of pre-cooled phosphate-buffered saline containing 3% fetal bovine serum. Subsequently, the cell suspension was centrifuged, the supernatant was removed, and the obtained intraperitoneal macrophages (PM) were counted. Cell culture medium containing 10% fetal bovine serum and 1% streptomycin/penicillin was used, and 5% carbon dioxide and 37 Cultured for 1 day in an incubator maintained at °C. Afterwards, 2 hours before cell infection, the cell culture medium was removed and replaced with cell culture medium containing 5% fetal calf serum, and 2 × 10 ⁵ cells were infected with Mycobacterium tuberculosis at an MOI of 5 for 2 hours. Next, the cells were washed twice using preheated phosphate-buffered saline, exchanged for cell culture medium containing 5% fetal bovine serum, and treated with 10 μM of Example 1 for 6 hours, and then collected. Real-time polymerase chain reaction was performed to measure the relative quantity of mRNA of inflammatory cytokines and chemokines expressed within cells from the collected cells, and the results of the experiment are shown in Figure 9. At this time, a group not infected with Mycobacterium tuberculosis (UI) and a group infected with Mycobacterium tuberculosis but not treated with Example 1 (gray graph of Mtb in Figure 9) were used as control groups.

As shown in Figure 9, when treated in Example 1, the mRNA expression levels of IL-1b, IL-6, and CXCL5 among inflammatory cytokines and chemokines in macrophages infected with Mycobacterium tuberculosis were compared to the control group. It was confirmed that there was a significant decrease in treatment group 1.

[Experimental Example 4-4] Changes in expression levels of inflammatory cytokines and chemokines in BCG (bacille de Calmette-Guerin) infection mouse model

After anesthetizing 8-week-old mice with tribromoethanol, they were infected with 25 μl of phosphate-buffered saline containing 1×10 ⁷ BCG bacteria by intranasal administration. From 3 days after infection, Example 3 was administered by intraperitoneal injection at 10 mg/kg every day. Seven days after infection, mice were sacrificed and lungs were removed. The extracted lung was homogenized in 1 ml of phosphate-buffered saline and centrifuged to collect lung tissue. The mRNA of inflammatory cytokines expressed from the collected tissues was measured by relative quantitation through real-time polymerase chain reaction, and the results of the experiment are shown in Figure 10. At this time, the group infected with BCG bacteria but not treated with Example 1 (Control in Figure 10) was used as a control group.

As shown in Figure 10, when treated in Example 1, it was confirmed that the mRNA expression level of the inflammatory cytokine tumor necrosis factor alpha (TNF-α) in the BCG respiratory infection mouse model was reduced by about half compared to the control group. .

Through this, it was found that the composition according to one aspect of the present invention exhibits the effect of suppressing the inflammatory response during bacterial infection such as typhoid fever bacillus and Mycobacterium tuberculosis, and is effective in preventing or treating inflammatory diseases caused by bacterial infection such as sepsis and tuberculosis. .

Claims (11)

An anti-inflammatory composition comprising a compound represented by the following formula (1), a salt, stereoisomer, solvate or hydrate thereof as an active ingredient:
[Formula 1]

In Formula 1,
R ₁ and R ₂ are independently -(CH ₂ ) _n1 -R _a , n1 is an integer of 1 to 3, and R _a is unsubstituted or fluoro-substituted phenyl;
R ₃ is -(CH ₂ ) _n2 - or -O-CH ₂ -CH(OH)-CH ₂ -, and n2 is an integer of 1 to 3;
R ₄ is -CH ₂ -CH ₂ - or -CH ₂ -CH ₂ -O-CH ₂ -CH ₂ -. A composition for preventing, alleviating or treating inflammatory diseases, comprising as an active ingredient a compound represented by the following formula (1), a salt, stereoisomer, solvate or hydrate thereof:
[Formula 1]

In Formula 1,
R ₁ and R _a are independently -(CH ₂ ) _n1 -R _a , n1 is an integer of 1 to 3, and R _a is unsubstituted or fluoro-substituted phenyl;
R ₃ is -(CH ₂ ) _n2 - or -O-CH ₂ -CH(OH)-CH ₂ -, and n2 is an integer of 1 to 3;
R ₄ is -CH ₂ -CH ₂ - or -CH ₂ -CH ₂ -O-CH ₂ -CH ₂ -. According to claim 1 or 2,
The composition, wherein the inflammation is caused by bacterial infection. According to paragraph 3,
The bacteria include Salmonella species, Yersinia species, Escherichia species, Chlamydia species, Klebsiella species, Proteus species, Xanthomonas species, Erwinia species, Pseudomonas species, Ralstonia species, Mycobacterium species, Streptococcus species, Staphylococcus species A composition, which is at least one selected from the group consisting of Staphylococcus species, Enterococcus species, and Lactobacillus species. According to paragraph 2,
The inflammatory disease is a disease caused by bacterial infection,
Diseases caused by the above bacterial infections include sepsis, tuberculosis, pneumonia, food poisoning, dysentery, Pseudomonas aeruginosa infection, impetigo, purulent disease, acute dermatitis, cystitis, vaginitis, meningitis, typhoid fever, paratyphoid fever, tetanus, Lyme disease, Rocky Mountain spotted fever, cholera, A composition that is one or more selected from the group consisting of leprosy, brucellosis, ehrlichiosis, Q fever, scarlet fever, listeriosis, botulism, leptospirosis, plague, typhus, bacteremia, endocarditis and enteritis. According to paragraph 2,
A composition wherein the inflammatory disease is an autoimmune disease. According to clause 6,
The composition, wherein the autoimmune disease is a toll-like receptor (TLR) related disease. According to claim 1 or 2,
The composition inhibits the expression of inflammatory cytokines or chemokines. According to clause 8,
The inflammatory cytokine is one selected from the group consisting of interleukin-1b (IL-1b), interleukin-6 (IL-6), and tumor necrosis factor alpha (TNF-α). Lee Sang-in, composition. According to clause 8,
The composition, wherein the chemokine is at least one selected from the group consisting of CXC motif chemokine 5 (CXCL5) and CXC motif chemokine 10 (CXCL10). According to claim 1 or 2,
The composition is a pharmaceutical, food or cosmetic composition.