KR102575347B1 - Novel diarylpropandione derivative compound, preparation method thereof, and pharmaceutical composition for preventing or treating inflammatory or allergic disease comprising the same - Google Patents

Abstract

The present invention relates to a novel diarylpropanedinone derivative compound, a method for producing the same, and a pharmaceutical composition containing the same for preventing or treating inflammatory or allergic diseases, and more specifically, to the novel diarylpropanedinone derivative of the present invention. The compound has the activity of suppressing the release of NLRP3 inflammasome, a mediator of inflammatory diseases, or β-hexosaminidase secreted by RBL-2H3 cells, a mediator of allergic diseases, thereby providing excellent inflammation or allergy suppression. . In addition, it has excellent stability as it does not show cytotoxicity in weakened cells induced by inflammation or allergies, and furthermore, it can be usefully used as a composition for treating, preventing or improving inflammatory or allergic diseases.

Description

Novel diarylpropandione derivative compound, preparation method thereof, and pharmaceutical composition for preventing or treating inflammatory or allergic disease comprising the same the same}

The present invention relates to a novel diarylpropanedinone derivative compound, a method for producing the same, and a pharmaceutical composition containing the same for preventing or treating inflammatory or allergic diseases.

Currently, there are countless different types of diseases in the world, and many people are exposed to the risk of these diseases. Among these, inflammatory diseases are so common that their risk is easily overlooked, but they are very dangerous because chronic diseases such as cancer are caused by inflammatory diseases.

Inflammation is generally caused by factors such as stress, physical injury, and excessive intake of high-calorie foods, and has been reported to be closely related to chronic diseases such as neurological diseases, metabolic diseases, and cancer. Many aspects of the reaction process and the induction process between inflammatory reactions and chronic diseases have not yet been clarified.

According to research results to date, inflammation can be broadly divided into acute inflammation and chronic inflammation, and acute inflammation is the initial stage of inflammation that repairs damage caused by invasion that causes any organic changes to cells or tissues of the living body. It acts as a biological defense reaction process for regeneration, and can be said to be a normal body reaction induced against external invading bacteria. On the other hand, chronic inflammation is a dangerous disease in which an excessive inflammatory response is induced, abnormal free radicals are generated, tissues are damaged or worsened, and this reaction process causes chronic diseases such as heart disease and cancer. In particular, chronic inflammation is limited to preventing the initial inflammation stage, so research into new anti-inflammatory substances is urgently needed.

Meanwhile, allergic diseases have increased significantly over the past few decades. It is known that various factors such as environment, genetics, hygiene, and food affect the development of allergic diseases such as atopy, asthma, and rhinitis. Several effective treatment methods, such as corticosteroids and antihistamines, are being developed, but the causative mechanism or complete treatment method is still not well known.

For these inflammatory or allergic diseases, existing treatment methods do not prevent the initial inflammation or provide complete treatment, so research on new substances is necessary.

Korean Patent No. 10-1808950

In order to solve the above problems, the purpose of the present invention is to provide a diarylpropanedinone derivative compound represented by Formula 1, or a pharmaceutically acceptable salt thereof.

Additionally, the present invention provides a pharmaceutical composition for preventing or treating inflammatory or allergic diseases, comprising the compound according to the present invention or a pharmaceutically acceptable salt thereof as an active ingredient.

In addition, the present invention provides a health functional food composition for preventing or improving inflammatory or allergic diseases, comprising the compound according to the present invention or a pharmaceutically acceptable salt thereof as an active ingredient.

Additionally, the present invention provides a cosmetic composition for preventing or improving inflammatory or allergic diseases, comprising the compound according to the present invention or a pharmaceutically acceptable salt thereof as an active ingredient.

Additionally, the present invention provides a method for producing a diarylpropanedinone derivative compound represented by Formula 1.

The object of the present invention is not limited to the objects mentioned above. The object of the present invention will become clearer from the following description and may be realized by means and combinations thereof as set forth in the claims.

The present invention provides a diarylpropanedinone derivative compound represented by the following formula (1), or a pharmaceutically acceptable salt thereof.

[Formula 1]

(In Formula 1, R ₁ to R ₃ are the same or different from each other, and are each independently H, OH, OCH ₃ , NRR' (where R and R' are each independently H, a C ₁ to C ₁₀ alkyl group) , and C ₃ to C ₁₀ allyl groups), C ₁ to C ₆₀ alkyl groups, and OCH ₂ O.)

Additionally, the present invention provides a pharmaceutical composition for preventing or treating inflammatory or allergic diseases, comprising the compound according to the present invention, or a pharmaceutically acceptable salt thereof, as an active ingredient.

In addition, the present invention provides a health functional food composition for preventing or improving inflammatory or allergic diseases containing the compound according to the present invention, or a pharmaceutically acceptable salt thereof, as an active ingredient.

Additionally, the present invention provides a cosmetic composition for preventing or improving inflammatory or allergic diseases, comprising the compound according to the present invention, or a pharmaceutically acceptable salt thereof, as an active ingredient.

In addition, the present invention includes the steps of preparing compound 3 by mixing compound 1 and compound 2 in a first organic solvent, as shown in Scheme 1 below; Washing and drying the compound 3; Preparing a reaction mixture by mixing Compound 3 and Compound 4 in a second organic solvent; and refluxing the reaction mixture and purifying it to prepare a diarylpropanedinone derivative compound represented by Formula 1 according to the present invention. A method for producing a diarylpropanedinone derivative compound represented by Formula 1, comprising: to provide.

[Scheme 1]

(In Formula 1, R ₁ to R ₃ are the same or different from each other, and are each independently H, OH, OCH ₃ , NRR' (where R and R' are each independently H, a C ₁ to C ₁₀ alkyl group) , and C ₃ to C ₁₀ allyl groups), C ₁ to C ₆₀ alkyl groups, and OCH ₂ O.)

The novel diarylpropanedinone derivative compound of the present invention has the activity of inhibiting the release of NLRP3 inflammasome, a mediator of inflammatory diseases, or β-hexosaminidase secreted from RBL-2H3 cells, a mediator of allergic diseases. By having it, you can excellently suppress inflammation or allergies. In addition, it has excellent stability as it does not show cytotoxicity in weakened cells induced by inflammation or allergies, and furthermore, it can be usefully used as a composition for treating, preventing or improving inflammatory or allergic diseases.

The effects of the present invention are not limited to the effects mentioned above. The effects of the present invention should be understood to include all effects that can be inferred from the following description.

Figure 1 shows (A) cell viability, (B) cytotoxicity analysis, and (C) inhibition of NLRP ₃ inflammasome activation induced by LPS+Nigericin for compound SPD 1 prepared in Example 1 according to the present invention. This is a graph showing the activity results.
Figure 2 shows (A) cell viability, (B) cytotoxicity analysis, and (C) inhibition of NLRP ₃ inflammasome activation induced by LPS+Nigericin for compound SPD 2 prepared in Example 2 according to the present invention. This is a graph showing the activity results.
Figure 3 shows (A) cell viability, (B) activity evaluation for IgE+DNP-induced β hexosaminidase release, and (C) for compounds SPD 1 and SPD 2 prepared in Examples 1 and 2 according to the present invention. This is a graph showing the activity evaluation results for A23187-induced β-hexosaminidase release.

Hereinafter, the present invention will be described in more detail by way of example.

As used herein, “including as an active ingredient” means including an amount sufficient to achieve the efficacy or activity of improving, treating or preventing inflammatory or allergic diseases according to the present invention.

As used herein, “prevention” may mean any act of suppressing, delaying, or alleviating inflammatory or allergic diseases by administering the composition according to the present invention to an individual.

As used herein, “treatment” may mean any action to improve or benefit the itching symptoms by administering the composition according to the present invention to a subject suspected of having an inflammatory or allergic disease.

The present invention relates to a diarylpropanedinone derivative compound, a method for producing the same, and a pharmaceutical composition containing the same for preventing or treating inflammatory or allergic diseases.

As previously explained, existing treatment methods for inflammatory or allergic diseases were only effective in preventing the initial stage, and research on the causative mechanism or complete treatment method was still insufficient.

Accordingly, in the present invention, a novel diarylpropanedinone derivative compound was synthesized, and the diarylpropanedinone derivative compound of the present invention was used in the NLRP3 inflammasome, a mediator of inflammatory diseases, or RBL-2H3 cells, a mediator of allergic diseases. By having the activity of suppressing the release of secreted β-hexosaminidase, it can excellently suppress inflammation or allergies. In addition, it has excellent stability as it does not show cytotoxicity in weakened cells induced by inflammation or allergies, and furthermore, it can be usefully used as a composition for treating, preventing or improving inflammatory or allergic diseases.

Specifically, the present invention provides a diarylpropanedinone derivative compound represented by the following formula (1), or a pharmaceutically acceptable salt thereof.

[Formula 1]

(In Formula 1, R ₁ to R ₃ are the same or different from each other, and are each independently H, OH, OCH ₃ , NRR' (where R and R' are each independently H, a C ₁ to C ₁₀ alkyl group) , and C ₃ to C ₁₀ allyl groups), C ₁ to C ₆₀ alkyl groups, and OCH ₂ O.)

Preferably, in Formula 1, R ₁ to R ₃ are the same as or different from each other, and may each independently be selected from H, OH, OCH ₃ , and OCH ₂ O.

Most preferably, the diarylpropanedinone derivative compound represented by Formula 1 is 1-(benzo[d][1,3]dioxol-5-yl)-3-(4-meth) represented by Formula 2 below: Toxyphenyl)-2-(3,4,5-trimethoxybenzylidene)propane-1,3-dione, or 2-(benzo[d][1,3]dioxole-5 represented by the following formula 3 -ylmethylene)-1-(4-methoxyphenyl)-3-(3,4,5-trimethoxyphenyl)propane-1,3-dione.

[Formula 2]

[Formula 3]

The diarylpropanedinone derivative compound represented by Formula 1 is a NLRP3 (nucleotide-binding oligomerization domain (NOD)-like receptor family, pyrin domain containing 3) inflammasome, which is a mediator of inflammatory diseases, or a mediator of allergic diseases. It may have the activity of inhibiting the release of β-hexosaminidase secreted from RBL-2H3 cells. This may be useful for preventing or treating inflammatory or allergic diseases. In addition, the diarylpropanedinone derivative compound has the advantage of excellent stability as it does not show cytotoxicity in weakened cells induced by inflammation or allergy.

The inflammatory disease may be any one selected from the group consisting of inflammatory bowel disease, systemic lupus erythematosus, inflammatory collagen vascular disease, glomerulonephritis, inflammatory skin disease, sarcoidosis, retinitis, gastritis, hepatitis, enteritis, pancreatitis, and nephritis. However, it is not limited to this.

The allergic disease may be any one selected from the group consisting of hypersensitivity, allergic rhinitis, asthma, allergic conjunctivitis, allergic dermatitis, atopic dermatitis, contact dermatitis, and urticaria, but is not limited thereto.

Meanwhile, the present invention provides a pharmaceutical composition for preventing or treating inflammatory or allergic diseases comprising the compound according to the present invention, or a pharmaceutically acceptable salt thereof, as an active ingredient.

The compound represented by Formula 2 may be included at a concentration of 0.625 to 5 uM, preferably 0.625 to 1.25 uM, to inhibit the release of NLRP3 inflammasome, a mediator of inflammatory diseases.

In addition, the compound represented by Formula 2 is used in an amount of 0.625 to 5 uM, preferably 1.25 to 5 uM, and most preferably to inhibit the release of β-hexosaminidase secreted from RBL-2H3 cells, which is a mediator of allergic diseases. It may be included at a concentration of 2.5 to 5 uM.

The diarylpropanedinone derivative compound represented by Formula 3 may be included at a concentration of 0.625 to 5 uM, preferably 0.625 to 2.5 uM, to inhibit the release of NLRP3 inflammasome, a mediator of inflammatory diseases.

In addition, the diarylpropanedinone derivative compound represented by Formula 2 inhibits the release of β-hexosaminidase secreted from RBL-2H3 cells, which is a mediator of allergic diseases, in an amount of 1.25 to 5 uM, preferably 2.5 to 5 uM. It may be included in a concentration of uM.

The appropriate dosage of the pharmaceutical composition varies depending on factors such as formulation method, administration method, patient's age, weight, sex, pathological condition, food, administration time, administration route, excretion rate, and reaction sensitivity, and is usually A skilled physician can easily determine and prescribe an effective dosage for desired treatment or prevention. According to a preferred embodiment, the daily dosage of the pharmaceutical composition is 0.001 to 10 g/kg.

The pharmaceutical composition can be administered orally or parenterally, and in the case of parenteral administration, it can be administered by intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, transdermal administration, etc., and is preferably oral administration.

The pharmaceutical composition is prepared by adding at least one selected from the group consisting of pharmaceutically acceptable carriers, excipients and diluents according to a method that can be easily performed by a person skilled in the art to which the invention pertains. It can be manufactured in unit dosage form by formulating it, or it can be manufactured by placing it in a multi-dose container. At this time, the dosage form may be any one selected from the group consisting of granules, powders, tablets, coatings, capsules, solutions, syrups, juices, suspensions, emulsions, drops, and injection solutions.

The pharmaceutical compositions include medicaments for human and veterinary use. Specifically, the diarylpropanedinone derivative compound can be used in the production of various veterinary medicines. At this time, the veterinary medicine can be used as a veterinary medicine or supplement that can efficiently treat and prevent atopic dermatitis disease in animals, an auxiliary treatment that can enhance treatment efficacy, a feed additive, etc.

In addition, the present invention provides a health functional food composition for preventing or improving inflammatory or allergic diseases containing the compound according to the present invention, or a pharmaceutically acceptable salt thereof, as an active ingredient.

The health functional food composition is any one formulation selected from the group consisting of capsules, tablets, powders, granules, liquids, pills, flakes, pastes, syrups, gels, jellies, and bars, and when ingested, it has specific health effects. However, unlike regular drugs, it has the advantage of not having any side effects that may occur when taking the drug for a long time since it is made from food.

The health functional food composition may contain not only the diarylpropanedinone derivative compound of the present invention as an active ingredient, but also ingredients commonly added during food production, such as proteins, carbohydrates, fats, nutrients, and seasonings. It may include agents and flavoring agents. Examples of the above-mentioned carbohydrates include monosaccharides such as glucose, fructose, etc.; Disaccharides such as maltose, sucrose, oligosaccharides, etc.; and polysaccharides, such as common sugars such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol.

The flavoring agent may be a natural flavoring agent such as thaumatin, stevia extract, and synthetic flavoring agent. At this time, the stevia extract may be rebaudioside A, glycyrrhizin, etc., and the synthetic flavoring agent may be saccharin, aspartame, etc. For example, when the health functional food composition is manufactured as a drink or beverage, in addition to the diarylpropanedinone derivative compound of the present invention, citric acid, high fructose corn syrup, sugar, glucose, acetic acid, malic acid, fruit juice, and various plant extracts are added. Can be included.

Additionally, the present invention provides a cosmetic composition for preventing or improving inflammatory or allergic diseases, comprising the compound according to the present invention, or a pharmaceutically acceptable salt thereof, as an active ingredient.

Ingredients included in the cosmetic composition include ingredients commonly used in cosmetic compositions in addition to the diarylpropanedinone derivative compound as an active ingredient, for example, antioxidants, stabilizers, solubilizers, vitamins, and fragrances. It may further include phosphorus adjuvants and detergents.

The cosmetic composition includes suspension, emulsion, paste, gel, cream, soap for atopic dermatitis, cleansing foam, cleansing cream, cleansing water, bath agent, skin softener, skin toner, lotion, cream, essence, astringent, emulsion, gel, lipstick, It may be any one formulation selected from the group consisting of spray, shampoo, rinse, treatment, body cleanser, pack, massage agent, face powder, compact, foundation, two-way cake, and makeup base.

If the formulation of the cosmetic composition is a paste, cream, or gel, it may further include vegetable oil, wax, starch, cellulose, silicone, bentonite, silica, zinc oxide, etc. as a carrier component, and if the formulation is a powder, lactose as a carrier component. , may further include powders such as silica, aluminum hydroxide, and calcium silicate. When the formulation of the cosmetic composition is an emulsion, it may further include a solvent, solubilizing agent, or emulsifying agent as a carrier component, such as water, ethanol, glycerin, ethyl carbonate, ethyl acetate, propylene glycol, butylene glycol. , 1,2-hexanediol, glycerol aliphatic ester, and sorbitan fatty acid ester.

When the formulation of the cosmetic composition is a suspension, the carrier ingredients include water, liquid diluents such as ethanol and propylene glycol, and suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester, and polyoxyethylene sorbitan ester. , microcrystalline cellulose, aluminum metahydroxide, bentonite, agar, etc. may be further included.

In addition, the present invention includes the steps of preparing compound 3 by mixing compound 1 and compound 2 in a first organic solvent, as shown in Scheme 1 below; Washing and drying the compound 3; Preparing a reaction mixture by mixing Compound 3 and Compound 4 in a second organic solvent; and refluxing the reaction mixture and purifying it to prepare a diarylpropanedinone derivative compound represented by Formula 1 according to the present invention. A method for producing a diarylpropanedinone derivative compound represented by Formula 1, comprising: to provide.

[Scheme 1]

(In Formula 1, R ₁ to R ₃ are the same or different from each other, and are each independently H, OH, OCH ₃ , NRR' (where R and R' are each independently H, a C ₁ to C ₁₀ alkyl group) , and C ₃ to C ₁₀ allyl groups), C ₁ to C ₆₀ alkyl groups, and OCH ₂ O.)

Preferably, in Formula 1, R ₁ to R ₃ are the same as or different from each other, and may each independently be selected from H, OH, OCH ₃ , and OCH ₂ O.

Most preferably, the diarylpropanedinone derivative compound represented by Formula 1 is 1-(benzo[d][1,3]dioxol-5-yl)-3-(4-meth) represented by Formula 2 below: Toxyphenyl)-2-(3,4,5-trimethoxybenzylidene)propane-1,3-dione, or 2-(benzo[d][1,3]dioxole-5 represented by the following formula 3 -ylmethylene)-1-(4-methoxyphenyl)-3-(3,4,5-trimethoxyphenyl)propane-1,3-dione.

[Formula 2]

[Formula 3]

The first organic solvent or the second organic solvent may be the same or different from each other, and may be tetrahydrofuran (THF), glacial acetic acid, piperidine, 1,4-dioxane, acetonitrile (MeCN), ethanol (EtOH), diethyl Ether, diphenyl ether, diisopropyl ether (DIPE), dimethylformamide (DMF), dimethylacetamide (DMA), dichloromethane (DCM), chlorobenzene, benzene, carbon tetrachloride (CCl ₄ ), It may be one or more selected from the group consisting of acetone and chloroform (CHCl ₃ ).

Preferably, the first organic solvent or the second organic solvent are different from each other, the first organic solvent may be tetrahydrofuran (THF), and the second organic solvent may be a mixture of glacial acetic acid and piperidine. there is.

Hereinafter, the present invention will be described in more detail based on examples, but the present invention is not limited to the following examples.

Example 1: Preparation of SPD 1

[Scheme 2]

Referring to Scheme 2 above, SPD1 was prepared in the following manner.

NaH (60%) was added to dry THF (4 mL) in a 25 mL one-neck round bottom flask equipped with a magnetic stir bar at room temperature under N ₂ atmosphere. Then, methyl 4-methoxybenzoate (1, 242 mg, 1.46 mmol) and 3', 4'-(methylenedioxy)acetophenone (2, 200 mg, 1.22 mmol) were added to dry THF mixed with NaH. After addition, the mixture was prepared by stirring. The mixture was placed in an ice bath and stirred under reflux for 10 hours. The mixture was then cooled and filtered through a Celite pad to obtain the reaction product. The obtained reaction was dissolved in Et ₂ O (20 mL) and aq. After mixing and extraction with HCl (1M, 20 mL), the organic layer was obtained, and the aqueous layer was extracted twice with Et ₂ O to obtain an extract. The obtained extract was washed three times with brine and dried. The extract was then evaporated and the residue was purified by flash column chromatography on silica to give compound 3, 1-(1,3-benzodioxol-5-yl)-3-(4-methoxyphenyl)-1, 3-Propanedione (160 mg, 45%) was obtained.

Then, 3,4 was added to 1-(1,3-benzodioxol-5-yl)-3-(4-methoxyphenyl)-1,3-propanedione (3, 100 mg, 36 mmol) of compound 3. , 5-Trimethoxybenzaldehyde (79 mg, 40 mmol) was added and stirred at room temperature for 30 minutes. Dry ethanol (5 mL), piperidine (0.1 mL, 87 mmol), and glacial acetic acid (0.5 mL) were added to the stirred reaction mixture. After refluxing for 24 hours, the reaction mixture was concentrated in vacuo and the residue was purified by flash column chromatography on silica to give SPD 1(1-(benzo[d][1,3]dioxol-5-yl) -3-(4-methoxyphenyl)-2-(3,4,5-trimethoxybenzylidene)propane-1,3-dione) (80 mg, 50%) was obtained as a yellow solid.

The ¹ H-NMR data of SPD 1 is as follows.

^1H NMR (600 MHz, CDCl ₃ ) δ 7.97 (dt, J = 9.0, 1.9 Hz, 1H), 7.93 - 7.88 (m, 1H), 7.62 - 7.56 (m, 1H), 7.51 - 7.46 (m, 1H) ), 7.34 (d, J = 1.4 Hz, 1H), 7.00 - 6.94 (m, 1H), 6.94 - 6.82 (m, 2H), 6.59 (s, 2H), 6.11 - 6.06 (m, 1H), 6.03 ( d, J = 0.9 Hz, 1H), 3.92 - 3.68 (m, 12H); ¹³ C NMR (150 MHz, CDCl ₃ ) δ 195.46, 195.31, 193.25, 192.90, 164.26, 163.37, 153.05, 153.02, 152.63, 151.68, 148.48, 148.09, 141.8 5, 141.73, 139.71, 139.13, 139.11, 132.12, 131.87, 131.56 , 130.08, 129.69, 128.60, 128.57, 126.77, 126.25, 114.21, 113.81, 109.57, 108.48, 108.42, 108.35, 107.92, 107.48, 107.40, 102.04, 101.92, 60.91, 55.98, 55.94, 55.54; HRMS (ES) caculated for C ₂₇ H ₂₄ O ₈ , [M+H] ⁺ = 476.1471 found: 476.1472

Example 2: Preparation of SPD 2

[Scheme 3]

Referring to Scheme 3 above, SPD2 was prepared in the following manner.

NaH (60%) was added to dry THF (4 mL) in a 25 mL one-neck round bottom flask equipped with a magnetic stir bar at room temperature under N ₂ atmosphere. Then, methyl 4-methoxybenzoate (1, 242 mg, 1.46 mmol) and 3,4,5-trimethoxybenzaldehyde (6, 200 mg, 0.95 mmol) were added to the dry THF mixed with NaH. A mixture was prepared by stirring. The mixture was placed in an ice bath and stirred under reflux for 12 hours. The mixture was then cooled and filtered through a Celite pad to obtain the reaction product. The obtained reaction was dissolved in Et ₂ O (20 mL) and aq. After mixing and extracting with HCl (1 M, 20 mL) to obtain an organic layer, the aqueous layer was extracted twice with Et ₂ O to obtain an extract. The obtained extract was washed three times with brine and dried. The extract was then evaporated and the residue was purified by flash column chromatography on silica to obtain compound 6 (120 mg, 43%).

Then, piperonal (7, 44 mg, 32 mmol) was added to compound 6 (100 mg, 29 mmol), and stirred at room temperature for 30 minutes. Dry ethanol (5 mL), piperidine (0.1 mL, 87 mmol), and glacial acetic acid (0.5 mL) were added to the stirred reaction mixture. After refluxing for 24 hours, the reaction mixture was concentrated in vacuo and the residue was purified by flash column chromatography on silica to obtain (2-(benzo[d][1,3]dioxol-5-yl) of SPD 2. Methylene)-1-(4-methoxyphenyl)-3-(3,4,5-trimethoxyphenyl)propane-1,3-dione) (72 mg, 52%) was obtained as an orange solid.

The ¹ H-NMR data of SPD 2 is as follows.

¹ H NMR (400 MHz, CDCl ₃ ) δ 7.96 - 7.88 (m, 2H), 7.49 (s, 1H), 7.25 (s, 1H), 7.09 (s, 1H), 6.96 - 6.82 (m, 4H), 6.71 (t, J = 8.0 Hz, 1H), 5.94 (d, J = 7.8 Hz, 2H), 3.91 - 3.87 (m, 4H), 3.83 (t, J = 4.2 Hz, 8H); ¹³ C NMR (100 MHz, CDCl ₃ ) δ 195.74, 193.79, 164.20, 153.20, 152.91, 142.78, 131.99, 131.93, 126.24, 114.16, 113.80, 109.26, 108.5 6, 106.96, 106.78, 101.88, 101.59, 60.93, 56.22, 56.17 , 55.51; HRMS (ESI) caculated for C ₂₇ H ₂₄ O ₈ , [M+H] ⁺ = 476.1471 found: 476.1472

Experimental Example 1: Evaluation of anti-inflammatory activity

In order to confirm the anti-inflammatory activity of compounds SPD 1 and SPD 2 prepared in Examples 1 and 2, compounds SPD 1 and SPD 2 were tested for cell viability, cytotoxicity, and NLRP3 inflammasome activation induced by LPS+Nigericin. The inhibitory activity was evaluated by the following method. At this time, the concentrations of the compounds SPD 1 and SPD 2 were 0.625, 1.25, 2.5, 5, 10, and 20 uM. The experimental results are shown in Figures 1 and 2.

(1) Evaluation of NLRP3 inflammasome stimulation

Specifically, to activate the NLRP3 inflammasome, bone marrow-derived macrophages (BMDM) were prepared with LPS (100 ng/ml) for 3 hours, and then the cells were washed with warm PBS before replacing the medium with Opti-MEM. The cells were cultured for 1 hour in the presence or absence of the sample containing the compound SPD 1 or SPD 2 prepared in Examples 1 and 2, and stimulated with KCL (150 mM) and Nigericin (5 uM) for 40 minutes. Next, SPD 1 and SPD 2 prepared in Examples 1 and 2 were analyzed for cell viability, cytotoxicity, and inhibitory activity on NLRP3 inflammasome activation induced by LPS+Nigericin. At this time, the concentrations of the compounds SPD 1 and SPD 2 were 0.625, 1.25, 2.5, 5, 10, and 20 uM.

(2) Lactate dehydrogenase (LDH) analysis

Supernatants of inflammasome-activated cells were collected to measure LDH release by cells. The EZ-LDH kit was used to measure LDH released from cells according to the instructions. OD was measured with a microplate reader (Multiskan GO, Thermo Scientific, Vantaa, Finland) at 450 nm.

(3) ELISA analysis

Cytokine IL-1β was performed using an ELISA kit. IL-1β levels were measured using Invitrogen's mouse IL-1 beta/IL-1F2 DuoSet ELISA kit. The resulting color was detected at 450 nm using a spectrophotometric microplate reader.

(4) Immunoblot analysis

Immunoblot analysis was used to determine proteins in cell supernatants and cell lysates. After mixing with sample buffer and boiling for 5 minutes, equal amounts of protein from the cell supernatant were loaded onto an SDS-Page gel for separation. Then, the proteins separated from the gel were transferred to a nitrocellulose membrane and later blocked with blocking buffer for 1 hour. Membranes were incubated with primary antibodies overnight at 4°C. The next day, the membrane was washed with washing buffer PBST and incubated for secondary antibodies at room temperature for 1 hour. After secondary antibody incubation, the membrane was washed and developed for protein detection by ECL kit.

(5) MTT analysis

To determine cell viability, after incubation with samples for 24 h, the supernatant was discarded, MTT (0.5 mg/mL) was replaced in each well, and then incubated at 37 °C for 2 h. After incubation, the supernatant was discarded and DMSO was added to each well to solubilize formazan. OD was measured at 450 nm for the LDH assay and 550 nm for the MTT assay, and values were calculated by comparison to control cells.

Figure 1 shows the results of (A) cell viability, (B) cytotoxicity analysis, and (C) inhibitory activity on NLRP ₃ inflammasome activation induced by LPS+Nigericin for compound SPD 1 prepared in Example 1. This is the graph shown. Referring to FIG. 1, the cell viability results show that compound SPD 1 prepared in Example 1 has an excellent inhibitory activity of more than 98% against the NLRP3 inflammasome at a concentration of 0.625 to 10 uM, and does not show cytotoxicity. confirmed. On the other hand, at a concentration of 20 uM of the compound SPD 1, cell viability was significantly low at 20%.

In addition, referring to (C) of FIG. 1, the results of evaluating NLRP ₃ inflammasome activation induced by LPS + Nigericin confirmed that the concentration of the compound SPD 1 exhibited a concentration-dependent inhibitory activity from 5 uM to 0.625 uM. did.

Figure 2 shows the results of (A) cell viability, (B) cytotoxicity analysis, and (C) inhibitory activity on NLRP ₃ inflammasome activation induced by LPS+Nigericin for compound SPD 2 prepared in Example 2. This is the graph shown. Referring to FIG. 2, the cell viability results show that compound SPD 2 prepared in Example 2 has an excellent inhibitory activity of more than 98% against the NLRP3 inflammasome at a concentration of 0.625 to 10 uM, and does not show cytotoxicity. confirmed. On the other hand, at a concentration of 20 uM, the compound SPD 2 showed low cell viability of 20% and cytotoxicity of 5%, and at 30 uM, cytotoxicity rapidly increased to 100%.

In addition, referring to (C) of FIG. 2, the results of evaluating NLRP ₃ inflammasome activation induced by LPS + Nigericin show that the concentration of the compound SPD 2 exhibits concentration-dependent inhibitory activity at the level of 5 uM to 0.625 uM. Confirmed.

Experimental Example 2: Evaluation of anti-allergic activity

To confirm the anti-allergy activity of the compounds SPD 1 and SPD 2 prepared in Examples 1 and 2, the anti-allergy activities of SPD 1 and SPD 2 were evaluated in RBL-2H3 cells. The experimental results are shown in Figure 3 and Tables 1 and 2.

Specifically, 150,000 RBL-2H3 cells were inoculated into a 24 well plate and cultured for 16 hours, and then the cells were sensitized with anti-DNP IgE for 2 hours. Thereafter, the cells were treated with the compounds SPD 1 or SPD 2 prepared in Examples 1 and 2 at levels of 0.625, 1.25, 2.5, and 5 uM, and then treated with DNP-HSA or A23187 to cross-link with IgE to degranulate the cells. After induction, β-hexosaminidase release was calculated 15 minutes later. The supernatant was then incubated with p-nitrophenyl-N-acetyl-β-d-glucosamine in sodium citrate (pH 4.5) at 37°C for 1 h, and then the reaction was stopped with sodium carbonate and sodium bicarbonate. Enzyme activity was assessed by measuring optical density at 405 nm. For comparison, test groups treated with no treatment (NT), pure IgE, pure DNP, and pure A23187 were also measured.

Figure 3 shows (A) cell viability, (B) activity evaluation for IgE+DNP-induced β hexosaminidase release, and (C) A23187-induced β for compounds SPD 1 and SPD 2 prepared in Examples 1 and 2. This is a graph showing the results of activity evaluation for hexosaminidase release.

Referring to (A) of FIG. 3, the compounds SPD 1 and SPD 2 showed a cell viability of more than 99% in the concentration range of 0.625 to 5 uM for RBL-2H3 cells, and no significant cytotoxicity was observed.

Table 1 below shows the results of the inhibitory effect on IgE-DNP-induced β-hexosaminidase release by SPD1 and 2 treatment.

Also, referring to (B) of FIG. 3 and Table 1, with respect to IgE+DNP-induced β-hexosaminidase release, except for the concentration of the compound SPD 1 in the range of 0.625 to 5 mM and SPD 2 of 0.625 uM. , it was confirmed that IgE+DNP-induced β-hexosaminidase release was suppressed by 15.6% to 72.3% as the concentration increased at each concentration. In particular, the compound SPD 1 inhibited β hexosaminidase release at the most significant level at a concentration of 1.25 to 5 uM, and the SPD 2 inhibited β hexosaminidase release at a significant level at concentrations of 12.5 and 5 uM. confirmed.

Table 2 below shows the results of the inhibition effect on A23187-induced β-hexosaminidase release by SPD1 and 2 treatment.

Also, referring to Figure 3 (C) and Table 2, with respect to A23187-induced β-hexosaminidase release, the compound SPD 1 was treated alone with A23187 at concentrations of 2.5 and 5 uM, except for concentrations of 0.625 and 1.25 uM. Compared to , as the concentration increased, the inhibitory effect was 24.5% and 41.5%, respectively, and SPD2 showed an inhibitory effect on β hexosaminidase release of 15 and 26.4% at concentrations of 2.5 and 5 uM, respectively.

Claims (12)

A diarylpropanedinone derivative compound represented by the following formula (1), or a pharmaceutically acceptable salt thereof,
The diarylpropanedinone derivative compound represented by Formula 1 is a diarylpropanedinone derivative compound represented by the following Formula 2 or Formula 3, or a pharmaceutically acceptable salt thereof:
[Formula 1]

(In Formula 1, R ₁ to R ₃ are the same or different from each other, and are each independently H, OH, OCH ₃ , NRR' (where R and R' are each independently H, a C ₁ to C ₁₀ alkyl group) , and C ₃ to C ₁₀ allyl groups), C ₁ to C ₆₀ alkyl groups, and OCH ₂ O.)
[Formula 2]

[Formula 3]

delete According to paragraph 1,
The diarylpropanedinone derivative compound represented by Formula 1 has the activity of inhibiting the release of NLRP3 inflammasome, a mediator of inflammatory diseases, or the release of β-hexosaminidase secreted from RBL-2H3 cells, a mediator of allergic diseases. A compound having a, or a pharmaceutically acceptable salt thereof.
According to paragraph 3,
The inflammatory disease is any one selected from the group consisting of inflammatory bowel disease, systemic lupus erythematosus, inflammatory collagen vascular disease, glomerulonephritis, inflammatory skin disease, sarcoidosis, retinitis, gastritis, hepatitis, enteritis, pancreatitis, and nephritis. A phosphorus compound, or a pharmaceutically acceptable salt thereof.
According to paragraph 3,
The allergic disease is a compound selected from the group consisting of hypersensitivity, allergic rhinitis, asthma, allergic conjunctivitis, allergic dermatitis, atopic dermatitis, contact dermatitis, and urticaria, or a pharmaceutically acceptable drug thereof. Possible salt.
A pharmaceutical composition for preventing or treating inflammatory or allergic diseases, comprising the compound of claim 1 or a pharmaceutically acceptable salt thereof as an active ingredient.
According to clause 6,
The compound represented by Formula 2 is included at a concentration of 0.625 to 5 uM to inhibit the release of NLRP3 inflammasome, a mediator of inflammatory diseases,
A pharmaceutical composition for preventing or treating inflammatory or allergic diseases, which is contained at a concentration of 0.625 to 5 uM to inhibit the release of β-hexosaminidase secreted from RBL-2H3 cells, which is a mediator of allergic diseases.
According to clause 6,
The compound represented by Formula 3 is included at a concentration of 0.625 to 5 uM to inhibit the release of NLRP3 inflammasome, a mediator of inflammatory diseases,
A pharmaceutical composition for preventing or treating inflammatory or allergic diseases, which is contained at a concentration of 1.25 to 5 uM to inhibit the release of β-hexosaminidase secreted from RBL-2H3 cells, which is a mediator of allergic diseases.
A health functional food composition for preventing or improving inflammatory or allergic diseases, comprising the compound of claim 1 or a pharmaceutically acceptable salt thereof as an active ingredient.
A cosmetic composition for preventing or improving inflammatory or allergic diseases comprising the compound of claim 1 or a pharmaceutically acceptable salt thereof as an active ingredient.
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