KR20200142470A - Novel 1-tetralone chalcone and composition for preventing or treating inflammatory disease comprising the same as active ingredient - Google Patents

Abstract

The present invention relates to a novel 1-tetrarone chalcone-based compound, and a composition for preventing or treating inflammatory diseases and containing the same as an active ingredient. A 6-amino-1-tetralone chalcone derivative compound according to the present invention is represented by chemical formula 1. In the chemical formula 1, R is selected from halogen, C1-C4 trifluoroalkyl, and C1-C4 trifluoroalkoxy. According to the present invention, a composition for preventing, alleviating, or treating inflammatory diseases can be provided.

Description

A novel 1-tetralone chalcone compound and composition for preventing or treating inflammatory disease containing the same as an active ingredient {Novel 1-tetralone chalcone and composition for preventing or treating inflammatory disease comprising the same as active ingredient}

The present invention is a novel 1-tetralone chalcone derivatives, especially 6-amino-1-tetralone chalcone derivatives, and containing them as an active ingredient. It relates to a composition for preventing or treating inflammatory diseases.

Inflammation is an important part of the body's immune response to infection, injury, pain, toxins or irritation. In the wound healing process, these reactions are very important in clearing the infection. However, if it lasts longer than necessary, it can cause harmful effects, which can lead to chronic inflammation. This persistent, chronic inflammatory response, which has a detrimental effect on the body and tissues, is known to be mediated through reactive oxygen species (ROS), which are known to be mediated by rheumatoid arthritis, atherosclerosis, Alzheimer's, inflammatory bowel disease and cancer. It is related to disease. ROS is a product of oxygen molecules and tends to decompose oxygen to produce ROS even during breathing. It is naturally highly reactive and can oxidize biologically important molecules such as DNA, proteins and lipids. ROS plays a dual role by acting as a signaling molecule under physiological conditions and a mediator of the inflammatory process at high concentrations. Our body is constantly under attack from ROS, and in this case, there is a system of antioxidant defense mechanisms in our body to regulate and maintain it. When the balance between ROS and defense mechanisms is broken, oxidative stress occurs, which plays an important role in the progression of many physiological disorders. Therefore, regulating the overproduction of ROS and balancing oxidative stress can be considered as one of the important strategies of inflammation treatment.

Macrophages, a part of the immune system, play an important role in defending our body against pathogenic stimuli through an entulfing process called phagocytosis. During phagocytosis, many inflammatory mediators, such as cytokines, chemokines, and nitric oxide (NO), work together with the inflammatory response, helping to rapidly decompose and restore normal tissue structures. Lipopolysaccharide (LPS), acting as a powerful toxin, activates macrophages even at very low concentrations. Stimulating macrophages with LPS induces rapid production of ROS, which induces the production of a variety of inflammatory mediators, including tumor necrosis factor-α, interferons and interleukins. In addition, ROS can interact with NO, which forms reactive nitrogen species (RNS), which in turn increases oxidative and nitrifying stress responses. Thus, pharmacological interventions including regulation and balance of ROS production in macrophages will be a promising strategy in the treatment of inflammatory diseases.

In addition, Crohn's Disease (CD) and ulcerative colitis (UC) are representative inflammatory bowel diseases (IBD) characterized by chronic and recurrent inflammatory conditions of the gastrointestinal tract and multifactorial immune disorders. . The etiology of IBD is unknown, but it is related to abnormal regulation of pro-inflammatory cytokines such as Tumor necrosis factor (TNF-α) and Interleukin 6 (IL-6). The unregulated inflammatory response is additionally triggered by environmental and genetic factors. TNF-α is a cell adhesion molecule, Vascular cell adhesion protein 1 (VCAM-1), Intercellular Adhesion Molecule 1 (ICM-1), and Interleukin 8 (IL-). It plays an important role in leukocyte recruitment through expression of chemokines such as 8) and mast cell protease 1 (MCP-1). Overexpression of pro-inflammatory molecules such as reactive oxygen species (ROS) and cytokines trigger an inflammatory chain reaction in the process of colitis. Most anti-TNF treatments, such as Infliximbab and adalimumab, have been successful in treating patients with IBD.

S100 protein is a family consisting of 25 members of EF-hand (a helix-loop-a helix) calcium-binding protein, cancer, heart failure, psoriasis, vasculitis, arteriosclerosis, glomerulonephritis, skin-/poly-myositis, brain Injury, Alzheimer's disease, Down syndrome, schizophrenia, depression, keratosis erythematosus, hair loss, Tourette's syndrome, multiple sclerosis, hyperzincemia, systemic inflammation, etc. are associated with various diseases. The S100 protein is provided as damage associated molecular patterns (DAMPs). Serum and mucosal members S100A8, S100A9, and S100A12 are recognized as essential biomarkers in colitis, and play an important role in the pathogenesis of IBD. S100A8 and S100A9 are Ca ^{2 +} binding pro-inflammatory proteins that are frequently found in neutrophils and monocytes. Due to their role in monocyte activation and leukocyte recruitment, S100A8/S100A9 is considered as a characteristic for the onset conditions of chronic inflammation and autoimmune diseases such as rheumatoid arthritis, systemic lupus erythematosus and IBD. They have been implicated in both NF-κB-related intracellular effects on the development of colon inflammation, as well as monocyte and TH1 cell-related extracellular effects. They are greatly upregulated during the inflammatory process induced by inflammatory cytokines such as TNF-α, IL-6 and ROS in colon epithelial cells and macrophages.

The S100 protein was shown to activate NADPH oxidase in macrophages through the interaction of cytoplasmic factors p67phox and RAC2, not p47phox. The S100A8 and S100A9 proteins exogenously enhanced colon cell migration and proliferation during culture. When added exogenously, S100A8 and S100A8/A9 not only promoted the expression of adhesion molecules such as ICAM-1 and VCAM-1, but were also involved in the adhesion of monocytes to endothelium. S100A8 is involved in pro-inflammatory activity through the receptor for advanced glycation end products (RAGE). However, little is known about the S100A8-mediated mutual interference between monocytes and colon epithelial cells. Therefore, S100A8 can be an effective new therapeutic target for colon inflammation.

Chalcone and its various derivatives are naturally occurring compounds widely found in various plants, fruits, spices, vegetables, and the like, and have valuable properties such as anti-inflammatory, anti-cancer, anti-fungal and antioxidant (FIGS. 1A and 1B). The use of chalcone as a medicine dates back to ancient times and is still in use. Since the structure is simple while having numerous pharmaceutical activities, interest in the synthesis of new chalcone derivatives has increased tremendously. In the design and development of various new drugs, the two aryl rings present in the chalcone structure help the flexibility of structural modification. Recently, many chalcone-based compounds have been commercially available, such as metochalcone (an otitis agent) and sofalcone (ulcer treatment agent and mucoprotective agent). Recently, many studies showing the role of chalcone in the inflammatory response have been reported. Compounds with polymethoxychalcone have shown potential properties as anti-inflammatory agents, inhibitors of inflammatory cytokine expression and inhibitors of prostaglandin production. In the RAW 264.7 macrophage line, the dimethylaminophenyl chalcone derivative was reported as a potential inhibitor of NO and prostaglandin E ₂ production (FIG. 1C ). In addition, the present inventors continued to study various modifications in the chalcone skeleton, and 3-benzylidenechroman-4-one, a hard chalcone derivative having moderate to strong inhibitory activity against ROS production in RAW 264.7 macrophages ( 3-benzylidenechroman-4-ones; FIG. 1d) and 2-benzylidene-1-indanones; FIG. 1e) were synthesized.

On the other hand, Barlow et al studied by the 4-amino-3,4-dihydro -2 H - naphthalen-1-one (4-amino-3,4-dihydro -2 H -napthalen-1-one;. FIG. The novel amine derivative of 1f) exhibited significant mast cell stabilization properties, and through this, it was found that the 1-tetralone nucleus having an aminocyclopentyl group is required to exhibit an anti-inflammatory effect.

Korean Patent Registration No. 10-1669759 (registered on October 20, 2016)

Accordingly, in the present invention, novel 1-tetralone chalcone derivatives, particularly 6-amino-1-tetralone chalcone derivatives, and these as active ingredients An object thereof is to provide a composition for preventing, improving or treating inflammatory diseases containing.

In order to solve the above problems, the present invention provides a 6-amino-1-tetralone chalcone derivative compound, an isomer thereof, or a pharmaceutically acceptable salt thereof.

In addition, the present invention is a 6-amino-1-tetralone chalcone (6-amino-1-tetralone chalcone) derivative compound or 1-tetralone chalcone (1-tetralone chalcone) derivative compound; It provides a pharmaceutical composition for preventing or treating inflammatory diseases containing isomers thereof or a pharmaceutically acceptable salt thereof as an active ingredient.

In addition, the present invention is a 6-amino-1-tetralone chalcone (6-amino-1-tetralone chalcone) derivative compound or 1-tetralone chalcone (1-tetralone chalcone) derivative compound; It provides a health functional food composition for preventing or improving inflammatory diseases containing their isomers or their pharmaceutically acceptable salts as an active ingredient.

The present invention is a novel 1-tetralone chalcone derivatives, especially 6-amino-1-tetralone chalcone derivatives, and containing them as an active ingredient. Regarding a composition for preventing or treating inflammatory diseases, the present inventors systematically designed 32 halogenated 1-tetralone and 6-amino-1-tetralone chalcone derivatives and synthesized them through a Claisen- Schmidt condensation reaction. The synthesized compounds were measured for the inhibitory effect of LPS-stimulated ROS production in RAW 264.7 macrophages. Among them, compound 18 with a 6-amino-1-tetralone chalcone moiety showed the most potent inhibitory effect of LPS-stimulated ROS production in RAW 264.7 macrophages. As a result of the structure-activity relationship (SAR) analysis, the chalcone derivative with the 1-tetralone skeleton is the chalcone derived from the 1-indanone or 4-chromanone skeleton previously reported. It showed stronger ROS inhibitory effect than the derivative. In addition, it was confirmed that the substitution of the amino group at the 6 ^th position of the 1-tetrarone skeleton greatly increased the inhibitory effect of LPS-stimulated ROS production in RAW 264.7 macrophages.

In addition, in an in vitro study, the 6-amino-1-tetrarone chalcone derivative showed the activity of inhibiting the adhesion of monocytes to TNF-α-induced epithelial cells, of which 6-amino-1-tetra Compound 18 with a lone chalcone moiety showed the strongest efficacy. Moreover, Compound 18 attenuated TNF-α-induced NADPH oxidase activity and NOX2 activation. In addition, the 6-amino-1-tetralone chalcone derivative significantly recovered weight loss and colon weight/length in DSS-induced colitis mice. Additionally, the present inventors found that the above compounds inhibit the expression of pro-inflammatory cytokines (TNF-α, IL-6, IL-1β) and cell adhesion molecules (ICAM-1, MCP-1) in DSS-treated colon tissue. Confirmed. These results support that NADPH oxidase-dependent ROS is associated with mononuclear and epithelial cell adhesion inducing colitis. Therefore, the present invention can be usefully utilized in the development of novel anti-inflammatory agents and therapeutic agents for inflammatory bowel disease.

Figure 1 is a) chalcone (chalcone), b) diphenyl substituted chalcones (diphenyl substituted chalcones), c) dimethylaminophenyl chalcone derivatives (dimethylaminophenyl chalcone derivatives), d) 3-benzylidenechroman-4-one derivative ( 3-benzylidenechroman-4-one derivatives), e) 2-benzylidene-1-indanone substituted derivatives and f) 4-amino-3,4-dihydro-2 It shows the structure of H -naphthalen-1-one derivatives (4-amino-3,4-dihydro-2 H- napthalen-1-one derivatives).
Figure 2 is a halogenated 2-benzylidene-3,4-dihydro-naphthalene -1 (2 H) - one (2-benzylidene-3,4-dihydronaphthalen -1 (2 H) -one) and 6-amino -2 - it represents a whole (6-amino-2-benzylidene -3,4-dihydronaphthalen-1 (2 H) -one) design strategy derivative-benzylidene-3,4-dihydro-naphthalene -1 (2 H).
3 shows the structure of a halogenated 1-tetralone substituted chalcone derivative 1-16 synthesized in the present invention.
4 shows the structure of a halogenated 6-amino-1-tetralone substituted chalcone derivative 17-32 synthesized in the present invention.
5 is a comparison of the effect of inhibiting ROS production through LPS stimulation in RAW 264.7 macrophages in 1-tetralone, 6-amino-1-tetralone chalcone derivatives and previously reported hard structural compounds.
6 shows the effect of inhibiting the adhesion of monocytes to TNF-α-induced colonic epithelial cells of a 6-amino-1-tetralone chalcone derivative.
7 shows the effect of 6-amino-1-tetralone chalcone derivatives in mouse DSS-induced colitis.

Thus, the present inventors 2-benzylidene (2-benzylidene) portion of ortho (ortho), meth (meta) or p (para) - with a functional group on the halogenated positions, 32 kinds of 1-tetralone (1-tetralone of ) Or 6-amino-1-tetralone (6-amino-1-tetralone) was designed and synthesized, and the inhibitory effect of these compounds on ROS production in LPS-stimulated 264.7 macrophages and TNF-α-induced epithelium The activity of inhibiting monocyte adhesion to cells was evaluated (Fig. 2).

As a result, 6-amino-1-tetralone chalcone derivatives were shown to inhibit LPS-induced ROS production in RAW 264.7 macrophages, and in particular, different derivatives in the basic skeleton of 6-amino-1-tetralone chalcone compounds The four derivatives of compounds 17, 18, 28, and 31, which are compounds to which are introduced, exhibited excellent ROS production inhibitory effects. The present inventors investigated the inhibition of adhesion of monocytes to the in vitro TNF-α-induced HT-29 colonic epithelial cells of the four 6-amino-1-tetralone chalcone derivatives and DSS-induced mice Colitis inhibitory effect was confirmed. In addition, the present inventors confirmed that the effects of the compounds are mediated through inhibition of NADPH activity, and completed the present invention.

The present invention provides a 6-amino-1-tetralone chalcone derivative compound represented by the following formula (1), an isomer thereof, or a pharmaceutically acceptable salt thereof.

[Formula 1]

In Formula 1, R is selected from halogen, C1-C4 trifluoroalkyl and C1-C4 trifluoroalkoxy.

Preferably, the 6-amino-1-tetralone chalcone derivative compound represented by Formula 1 may be any one selected from the following compounds 18 to 32, but is limited thereto. It is not.

The pharmaceutically acceptable salts are hydrochloride, bromate, sulfate, phosphate, nitrate, citrate, acetate, lactate, tartrate, maleate, gluconate, succinate, formate, trifluoroacetate, oxalate, fumaric acid. Salt, methanesulfonate, benzenesulfonate, paratoluenesulfonate, camphorsulfonate, sodium salt, potassium salt, lithium salt, calcium salt and magnesium salt may be selected from the group consisting of, but is not limited thereto.

In addition, the present invention is a 6-amino-1-tetralone chalcone derivative compound represented by the formula (1) or 1-tetralone chalcone (1-tetralone chalcone) derivative represented by the formula (2) compound; It provides a pharmaceutical composition for preventing or treating inflammatory diseases containing isomers thereof or a pharmaceutically acceptable salt thereof as an active ingredient.

[Formula 1]

[Formula 2]

In Formula 1 or Formula 2, R is selected from halogen, C1-C4 trifluoroalkyl and C1-C4 trifluoroalkoxy.

Preferably, the 6-amino-1-tetralone chalcone derivative compound represented by Formula 1 may be any one selected from the following compounds 17 to 32, but is limited thereto. It is not.

Preferably, the 1-tetralone chalcone derivative compound represented by Formula 2 may be any one selected from compounds 1 to 16 below, but is not limited thereto.

Preferably, the inflammatory disease is dermatitis, conjunctivitis, periodontitis, rhinitis, otitis media, sore throat, tonsillitis, pneumonia, gastric ulcer, gastritis, inflammatory bowel disease, Crohn's disease, colitis, hemorrhoids, gout, ankylosing spondylitis, lupus, fibromyalgia , Psoriatic arthritis, osteoarthritis, rheumatoid arthritis, peri-shoulder joint infection, tendinitis, tendonitis, tendonitis, myositis, hepatitis, cystitis, nephritis, sjogren's syndrome, multiple sclerosis, and an inflammatory disease caused by angiogenesis, but is limited thereto. It is not.

When the composition of the present invention is a pharmaceutical composition, it may be prepared using a pharmaceutically suitable and physiologically acceptable adjuvant in addition to the active ingredient, and the adjuvants include excipients, disintegrants, sweeteners, binders, coating agents, expanding agents, Solubilizing agents such as lubricants, lubricants, or flavoring agents may be used. The pharmaceutical composition of the present invention can be preferably formulated as a pharmaceutical composition, including at least one pharmaceutically acceptable carrier in addition to the active ingredient for administration. As acceptable pharmaceutical carriers for compositions formulated as liquid solutions, sterilization and biocompatible, saline, sterile water, Ringer's solution, buffered saline, albumin injection solution, dextrose solution, maltodextrin solution, glycerol, ethanol, and One or more of these components may be mixed and used, and other conventional additives such as antioxidants, buffers, and bacteriostatic agents may be added as necessary. In addition, diluents, dispersants, surfactants, binders, and lubricants may be additionally added to prepare injectable formulations such as aqueous solutions, suspensions, emulsions, etc., pills, capsules, granules, or tablets.

The pharmaceutical formulation forms of the pharmaceutical composition of the present invention may be granules, powders, coated tablets, tablets, capsules, suppositories, syrups, juices, suspensions, emulsions, drops or injectable solutions, and sustained-release formulations of active compounds. I can. The pharmaceutical composition of the present invention is a conventional method through intravenous, intraarterial, intraperitoneal, intramuscular, intraarterial, intraperitoneal, intrasternal, transdermal, nasal, inhalation, topical, rectal, oral, intraocular or intradermal routes. Can be administered. The effective amount of the active ingredient of the pharmaceutical composition of the present invention means an amount required for prevention or treatment of a disease. Therefore, the type of disease, the severity of the disease, the type and content of the active ingredient and other ingredients contained in the composition, the type of formulation and the patient's age, weight, general health condition, sex and diet, administration time, administration route and composition It can be adjusted according to various factors including the rate of secretion, duration of treatment, and drugs used simultaneously.

In addition, the present invention is a 6-amino-1-tetralone chalcone derivative compound represented by the formula (1) or 1-tetralone chalcone (1-tetralone chalcone) derivative represented by the formula (2) compound; It provides a health functional food composition for preventing or improving inflammatory diseases containing their isomers or their pharmaceutically acceptable salts as an active ingredient.

[Formula 1]

[Formula 2]

In Formula 1 or Formula 2, R is selected from halogen, C1-C4 trifluoroalkyl and C1-C4 trifluoroalkoxy.

When the composition of the present invention is a health functional food composition, the health functional food composition may be provided in the form of powder, granules, tablets, capsules, syrup or beverage, and the health functional food composition is a food or food additive other than the active ingredient It is used together with, and may be appropriately used according to a conventional method. The mixing amount of the active ingredient may be appropriately determined according to the purpose of use, for example, prevention, health or therapeutic treatment.

The effective dose of the active ingredient contained in the health functional food composition can be used in accordance with the effective dose of the pharmaceutical composition, but in the case of long-term intake for the purpose of health and hygiene or health control, it should be less than the above range. It is clear that the active ingredient can be used in an amount beyond the above range because there is no problem in terms of safety.

There is no particular limitation on the kind of health food, for example, meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, tea, Drinks, alcoholic beverages, and vitamin complexes.

Hereinafter, examples will be described in detail to aid understanding of the present invention. However, the following examples are for illustrative purposes only, and the scope of the present invention is not limited to the following examples. The embodiments of the present invention are provided to more completely explain the present invention to those of ordinary skill in the art.

<Experimental Example>

The following experimental examples are intended to provide experimental examples commonly applied to each of the examples according to the present invention.

1. Experimental material

All commercially available starting materials and reagents are available from Sigma-Aldrich Chemical Co. (Darmstadt, Germany), TCI Chemicals (Tokyo, Japan), Junsei (Tokyo, Japan) and Alfa-Aesar (Heysham, England), and used without further purification. HPLC grade acetonitrile (ACN) and methanol were purchased from Budrick and Jackson (Shanghai, China). Thin layer chromatography (TLC) was performed by kieselgel 60 F _254. (Merck, Darmstadt, Germany). Since the synthetic compounds all contain aromatic rings, they were visualized and detected with UV ultraviolet (short wavelength, long wavelength, or both) on TLC plates. NMR spectra were recorded on Bruker AMX 250 (250 MHz, FT) for ¹ H NMR and 63 MHz for ¹³ C NMR, and chemical shifts were calculated according to TMS. The chemical shift ( δ ) was recorded in ppm and the binding constant ( J ) was recorded in hertz (Hz). The melting point was measured in an open capillary tube on an electrothermal 1A 9100 digital melting point apparatus, and no correction was made.

HPLC analysis was performed by Shimadzu Scientific Instruments (Kyoto, Japan) consisting of a pump (LC-20AD), an autoinjector (SIL-20-A), a UV-visible detector (SPD-20A) and a communications bus module (CBM-20A). This was done using an HPLC system.

95% ACN: Re-distilled water at 95:5 in a solvent gradient system for 20 minutes at a flow rate of 0.5 mL/min, UV detection at 254 nm, A Waters COSMOSIL 5C18-MS-II column (5 μm, 4.6 x 250 mm ) Was used. The purity of the compound was expressed in percentage (%), and the residence time was expressed in minutes (min). The mass spectrum was measured in a positive electrospray ionization (ESI) mode on an LCMS-2020 system of Shimadzu Scientific Instruments (Tokyo, Japan).

DMEM High Glucose, RPMI1640, and fetal bovine serum (FBS) were purchased from GE Healthcare Life Sciences (Logan, UT, USA). Penicillin/Streptomycin, Trypsin/EDTA were purchased from Thermo Fisher Scientific (Waltham, MA, USA). Recombinant human TNF-α was purchased from R & D system Inc (Minneapolis, MN, USA), and BCECF/AM (acetoxymethyl ester) was purchased from Molecular probes (Eugene, Oregon, USA). Salfasalazine and VAS2870 were supplied from Sigma-Aldrich (St. Louis, MO, USA). Antibodies against TNF-α, IL-1β, NOX2, NOX1, ICAM-1 and S100A8 were purchased from Abcam (Cambridge, MA, USA). The p-p47 phox, p47 phox rabbit and p67 phox antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Antibodies against IL-6 and IL-8 were purchased from Abbiotec (San Diego, CA, USA). Antibodies against MCP-1 were purchased from R & D system Inc. Anti-rabbit Alexa Fluor 647 was purchased from Abcam, and Anti-mouse Alexa Fluor 488 was purchased from Invitrogen Life Technologies (Carlsbad, CA, USA).

2. General Synthesis Method of Compound 1-32

The general synthesis method of compound 1-32 was carried out as previously reported (Eur. J. Med. Chem. 137 (2017) 575-597, Bioorg. Chem. 62 (2015) 30-40). The detailed properties of the compounds are mentioned below.

1) 2-benzylidene-3,4-dihydronaphthalen-1(2H)-one [2-benzylidene-3,4-dihydronaphthalen-1(2H)-one] (1)

mp: 109.8-110.7 °C (ref: 106.0-107.3 °C (ethanol)), HPLC: Retention time: 7.74 min, purity: 98.2%.

2) 2-(2-fluorobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [2-(2-fluorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one] ( 2)

mp: 60.1-61.1 °C (ref: 56.0-58.0 °C), HPLC: Retention time: 8.51 min, purity: 97.2%, ESI LC/MS: m/z calcd for C ₁₇ H ₁₃ F0 [MH] ⁺ 253.09; found 253.70.

3) 2-(3-fluorobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [2-(3-fluorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one] ( 3)

mp: 87.9-88.7 °C (ref: 89.0-91.0 °C), HPLC: Retention time: 8.41 min, purity: 95.2%, ESI LC/MS: m/z calcd for C ₁₇ H ₁₃ F0 [MH] ⁺ 253.09; found 253.70.

4) 2-(4-fluorobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [2-(4-fluorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one] ( 4)

mp: 113.1-114.1 °C (ref: 112.0-116.0 °C), HPLC: Retention time: 8.50 min, purity: 95.5%, ESI LC/MS: m/z calcd for C ₁₇ H ₁₃ F0 [MH] ⁺ 253.09; found 253.70.

5) 2-(2-chlorobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [2-(2-chlorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one] (5 )

mp: 72.4-73.2 °C (ref: 109.0-112.0 °C), HPLC: Retention time: 9.19 min, purity: 95.2%, ESI LC/MS: m/z calcd for C ₁₇ H ₁₃ ClO [MH] ⁺ 269.06; found 269.65.

6) 2-(3-chlorobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [2-(3-chlorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one] (6 )

mp: 111.4-112.1 ℃ (ref: 106.0-108.0° C.), HPLC: Retention time: 9.57 min, purity: 96.2%, ESI LC/MS: m/z calcd for C ₁₇ H ₁₃ ClO [MH] ⁺ 269.06; found 269.65.

7) 2-(4-chlorobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [2-(4-chlorobenzylidene)-3,4dihydronaphthalen-1(2H)-one] (7)

mp: 139.0-139.7 ℃ (ref: 134.0-136.0 °C), HPLC: Retention time: 9.59 min, purity: 95.2%.

8) 2-(2-bromobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [2-(2-bromobenzylidene)-3,4-dihydronaphthalen-1(2H)-one] ( 8)

Yield: 86.6%, white solid; TLC (ethyl acetate/ hexanes = 1:27 v/v) R _f : 0.26, mp: 77.9-78.5 ℃, (ref: 78.0-80.0° C.), HPLC: Retention time: 9.52 min, purity: 97.2%, ESI LC/MS: m/z calcd for C ₁₇ H ₁₃ BrO [MH] ⁺ 313.01; found 313.55.

¹ H NMR (250 MHz, CDCl ₃ ) δ 8.15 (d, J = 7.8 Hz, 1H), 7.82 (s, 1H), 7.63 (d, J = 7.9 Hz, 1H), 7.47 (dd, J = 10.5, 4.4 Hz, 1H), 7.32 (dt, J = 8.3, 6.6 Hz, 3H), 7.25-7.15 (m, 2H), 2.93 (s, 4H).

¹³ C NMR (63 MHz, CDCl ₃ ) δ 187.57, 143.42, 136.89, 136.34, 135.58, 133.43, 133.27, 132.93, 130.38, 129.67, 128.30, 128.27, 127.05, 126.97, 124.87, 29.01, 27.27.

9) 2-(3-bromobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [2-(3-bromobenzylidene)-3,4-dihydronaphthalen-1(2H)-one] ( 9)

mp: 110.9-111.4 °C (ref: 109.0-110.0 °C), HPLC: Retention time: 9.99 min, purity: 98.7%, ESI LC/MS: m/z calcd for C ₁₇ H ₁₃ BrO [MH] ⁺ 313.01; found 313.55.

10) 2-(4-bromobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [2-(4-bromobenzylidene)-3,4-dihydronaphthalen-1(2H)-one] ( 10)

mp: 153.5-154.2 °C (ref: 139.0-141.0 °C), HPLC: Retention time: 8.84 min, purity: 97.8%, ESI LC/MS: m/z calcd for C ₁₇ H ₁₃ BrO [MH] ⁺ 313.01; found 313.55.

11) 2-(2-(trifluoromethyl)benzylidene)-3,4-dihydronaphthalene-1(2H)-one [2-(2-(trifluoromethyl)benzylidene)-3,4-dihydronaphthalen-1 (2H)-one] (11)

Yield: 92.0%, white solid; TLC (ethyl acetate/ hexanes = 1:27 v/v) R _f : 0.21, mp: 77.4-78.4 ℃, HPLC: Retention time: 8.62 min, purity: 98.5%, ESI LC/MS: m/z calcd for C ₁₈ H ₁₃ F ₃ O [MH] ⁺ 303.09; found 303.60.

¹ H NMR (250 MHz, CDCl ₃ ) δ 8.15 (dd, J = 7.8, 1.3 Hz, 1H), 7.96 (s, 1H), 7.72 (d, J = 7.7 Hz, 1H), 7.56 (t, J = 7.4 Hz, 1H), 7.52-7.39 (m, 2H), 7.39-7.27 (m, 2H), 7.23 (d, J = 7.5 Hz, 1H), 2.92 (dd, J = 9.2, 4.0 Hz, 2H), 2.87-2.78 (m, 2H).

¹³ C NMR (63 MHz, CDCl ₃ ) δ 187.39, 143.50, 138.05, 134.88 (d, J = 1.9 Hz), 133.50, 133.15, 132.94, 131.47, 130.34, 129.21 (d, J = 30.2 Hz), 128.35, 128.30 , 128.04, 127.07, 126.07 (q, J = 5.3 Hz), 121.75, 29.03, 27.36.

12) 2-(3-(trifluoromethyl)benzylidene)-3,4-dihydronaphthalene-1(2H)-one[2-(3-(trifluoromethyl)benzylidene)-3,4-dihydronaphthalen-1 (2H)-one] (12)

Yield: 71.4%, off-white solid; TLC (ethyl acetate/ hexanes = 1:29 v/v) R _f : 0.30, mp: 106.5-107.5 ℃, HPLC: Retention time: 8.86 min, purity: 97.6%, ESI LC/MS: m/z calcd for C ₁₈ H ₁₃ F ₃ O [MH] ⁺ 303.09; found 303.60.

¹ H NMR (250 MHz, DMSO) δ 7.96 (d, J = 7.8 Hz, 1H), 7.81 (d, J = 8.1 Hz, 2H), 7.78-7.64 (m, 3H), 7.58 (td, J = 7.4 , 1.3 Hz, 1H), 7.39 (dd, J = 13.1, 7.5 Hz, 2H), 3.11-3.00 (m, 2H), 3.00-2.89 (m, 2H).

¹³ C NMR (63 MHz, DMSO) δ 186.73, 143.65, 137.34, 136.51, 133.96, 133.86, 133.68, 132.83, 129.81, 129.60 (d, J = 31.7 Hz), 128.75, 127.59, 127.21, 126.39 (q, J = 3.9 Hz), 125.25 (q, J = 3.6 Hz), 122.04, 27.99, 26.73.

13) 2-(4-(trifluoromethyl)benzylidene)-3,4-dihydronaphthalene-1(2H)-one [2-(4-(trifluoromethyl)benzylidene)-3,4-dihydronaphthalen-1 (2H)-one] (13)

mp: 176.9-177.9 °C, HPLC: Retention time: 9.10 min, purity: 99.5%.

14) 2-(2-(trifluoromethoxy)benzylidene)-3,4-dihydronaphthalene-1(2H)-one [2-(2-(trifluoromethoxy)benzylidene)-3,4-dihydronaphthalen-1 (2H)-one] (14)

Yield: 88.9%, light yellowish liquid; TLC (ethyl acetate/ hexanes = 1:27 v/v) R _f : 0.28, HPLC: Retention time: 8.47 min, purity: 95.5%, ESI LC/MS: m/z calcd C ₁₈ H ₁₃ F ₃ O ₂ [ MH] ⁺ 319.08; found 319.60

¹ H NMR (250 MHz, CDCl ₃ ) δ 8.14 (dd, J = 7.8, 1.2 Hz, 1H), 7.83 (s, 1H), 7.48 (td, J = 7.4, 1.5 Hz, 1H), 7.43-7.27 ( m, 5H), 7.23 (d, J = 7.4 Hz, 1H), 2.93 (s, 4H).

¹³ C NMR (63 MHz, CDCl ₃ ) δ 187.36, 147.50, 143.43, 138.05, 133.46, 133.19, 130.78, 130.38, 129.76, 129.58, 128.30 (2C), 127.08, 126.54, 121.18 (d, J = 1.3 Hz), 120.49 (d, J = 258.2 Hz), 28.96, 27.55.

15) 2-(3-(trifluoromethoxy)benzylidene)-3,4-dihydronaphthalene-1(2H)-one [2-(3-(trifluoromethoxy)benzylidene)-3,4-dihydronaphthalen-1 (2H)-one] (15)

Yield: 91.2%, white solid; TLC (ethyl acetate/ hexanes = 1:18 v/v) R _f : 0.27, mp: 150.8-151.2 ℃, HPLC: Retention time: 9.06 min, purity: 97.5%, ESI LC/MS: m/z calcd C ₁₈ H ₁₃ F ₃ O ₂ [MH] ⁺ 319.08; found 319.55.

¹ H NMR (250 MHz, CDCl ₃ ) δ 8.12 (d, J = 7.8 Hz, 1H), 7.79 (s, 1H), 7.53-7.39 (m, 2H), 7.35 (t, J = 7.9 Hz, 2H) , 7.28-7.16 (m, 3H), 3.14-3.03 (m, 2H), 3.01-2.90 (m, 2H).

¹³ C NMR (63 MHz, CDCl ₃ ) δ 187.50, 149.26, 143.16, 137.85, 136.85, 134.68, 133.49, 133.26, 129.85, 128.29, 128.24, 128.15, 127.13, 121.98, 120.74, 120.47 (d, J = 257.5 Hz) , 28.75, 27.10.

16) 2-(4-(trifluoromethoxy)benzylidene)-3,4-dihydronaphthalen-1(2H)-one [2-(4-(trifluoromethoxy)benzylidene)-3,4-dihydronaphthalen-1 (2H)-one] (16)

Yield: 40.6%, white solid; TLC (ethyl acetate/ hexanes = 1:27 v/v) R _f : 0.25, mp: 136.8-137.8 °C, HPLC: Retention time: 8.58 min, purity: 98.1%, ESI LC/MS: m/z calcd C ₁₈ H ₁₃ F ₃ O ₂ [MH] ⁺ 319.08; found 319.55.

¹ H NMR (250 MHz, CDCl ₃ ) δ 8.12 (dd, J = 7.7, 1.0 Hz, 1H), 7.81 (s, 1H), 7.52-7.40 (m, 3H), 7.35 (t, J = 7.5 Hz, 1H), 7.24 (d, J = 8.1 Hz, 3H), 3.08 (dd, J = 8.9, 4.3 Hz, 2H), 2.94 (t, J = 6.1 Hz, 2H).

¹³ C NMR (63 MHz, CDCl ₃ ) δ 187.58, 149.00, 143.12, 136.19, 134.89, 134.47, 133.42, 133.32, 131.27 (2C), 128.27, 128.21, 127.11, 120.81, 120.43 (d, J = 257.6 Hz), 28.77, 27.11.

17) 6-amino-2-benzylidene-3,4-dihydronaphthalen-1(2H)-one [6-amino-2-benzylidene-3,4-dihydronaphthalen-1(2H)-one] (17)

mp: 175.3-176.2 °C, (ref: 139.0-141.0 °C (ethanol)), HPLC: Retention time: 7.02 min, purity: 98.9%.

18) 6-amino-2-(2-fluorobenzylidene)-3,4-dihydronaphthalene-1(2H)-one[6-amino-2-(2-fluorobenzylidene)-3,4-dihydronaphthalen- 1(2H)-one] (18)

Yield: 34.1%, orange solid, TLC (ethyl acetate/ hexanes = 1:6v/v) R _f : 0.21, mp: 136.4-137.1 ℃, HPLC: Retention time: 7.17 min, purity: 98.9%, ESI LC/MS : m/z calcd for C ₁₇ H ₁₄ FNO [MH] ⁺ 268.10; found 268.70.

¹ H NMR (250 MHz, DMSO) δ 7.71 (d, J = 8.6 Hz, 1H), 7.50 (s, 1H), 7.49-7.37 (m, 2H), 7.27 (dt, J = 8.4, 6.3 Hz, 2H ), 6.51 (dd, J = 8.6, 2.1 Hz, 1H), 6.34 (d, J = 1.9 Hz, 1H), 6.20 (s, 2H), 2.90-2.79 (m, 2H), 2.79-2.69 (m, 2H).

¹³ C NMR (63 MHz, DMSO) δ 183.88, 162.16, 154.31, 145.99, 139.01, 130.96 (d, J = 3.0 Hz), 130.59 (d, J = 8.5 Hz), 130.40, 125.56 (d, J = 3.2 Hz) ), 124.54 (d, J = 3.4 Hz), 123.65 (d, J = 14.0 Hz), 121.49, 115.81 (d, J = 21.8 Hz), 112.88, 110.79, 28.64, 27.25.

19) 6-amino-2-(3-fluorobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [6-amino-2-(3-fluorobenzylidene)-3,4-dihydronaphthalen- 1(2H)-one] (19)

Yield: 20.2%, dark yellow solid, TLC (ethyl acetate/ hexanes = 1:6 v/v) R _f : 0.21, mp: 150.5-151.5 ℃, HPLC: Retention time: 7.37 min, purity: 99.9%, ESI LC /MS: m/z calcd for C ₁₇ H ₁₄ FNO [MH] ⁺ 268.10; found 268.70.

¹ H NMR (250 MHz, CDCl ₃ ) δ 7.98 (d, J = 8.5 Hz, 1H), 7.71 (s, 1H), 7.34 (dd, J = 14.0, 7.8 Hz, 1H), 7.20-6.94 (m, 3H), 6.58 (dd, J = 8.5, 2.0 Hz, 1H), 6.40 (s, 1H), 4.18 (s, 2H), 3.02 (t, J = 5.9 Hz, 2H), 2.81 (t, J = 6.3 Hz, 2H).

¹³ C NMR (63 MHz, CDCl ₃ ) δ 185.99, 151.43, 145.82, 138.43 (d, J = 7.7 Hz), 137.12, 133.67 (d, J = 2.3 Hz), 130.98, 129.84 (d, J = 8.4 Hz) , 125.54 (d, J = 2.9 Hz), 124.47, 116.21 (d, J = 21.6 Hz), 114.96 (d, J = 21.3 Hz), 113.54, 112.02, 99.98, 29.07, 27.13.

20) 6-amino-2-(4-fluorobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [6-amino-2-(4-fluorobenzylidene)-3,4-dihydronaphthalen- 1(2H)-one] (20)

Yield: 26.9%, brown solid; TLC (ethyl acetate/ hexanes = 1:6 v/v) R _f : 0.20, mp: 215.4-216.3 ℃, HPLC: Retention time: 7.1 min, purity: 99.8%, ESI LC/MS: m/z calcd for C ₁₇ H ₁₄ FNO [MH] ⁺ 268.10; found 268.70.

¹ H NMR (250 MHz, DMSO) δ 7.83-7.68 (m, J = 8.5 Hz, 4H), 7.61-7.48 (m, 3H), 7.26 (t, J = 8.9 Hz, 2H), 6.75 (dd, J = 8.5, 1.9 Hz, 1H), 6.62 (s, 1H), 2.97 (t, J = 6.0 Hz, 2H), 2.78 (t, J = 6.3 Hz, 2H).

¹³ C NMR (63 MHz, DMSO) δ 184.68, 163.97, 145.55, 136.19 (d, J = 1.2 Hz), 133.05, 132.30, 132.24, 132.21, 132.07, 130.06, 124.43, 115.83, 115.49, 115.27, 113.97, 28.42, 26.77.

21) 6-amino-2-(2-chlorobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [6-amino-2-(2-chlorobenzylidene)-3,4-dihydronaphthalen-1 (2H)-one] (21)

Yield: 22.4%, orange solid; TLC (ethyl acetate/ hexanes = 2:5 v/v) R _f : 0.26, mp: 60.7-61.7 ℃, HPLC: Retention time: 7.58 min, purity: 95.1%, ESI LC/MS: m/z calcd for C ₁₇ H ₁₄ ClNO [MH] ⁺ 284.07; found 284.60.

¹ H NMR (250 MHz, CDCl ₃ ) δ 8.00 (d, J = 8.5 Hz, 1H), 7.80 (s, 1H), 7.41 (dd, J = 7.9, 4.0 Hz, 1H), 7.30-7.21 (m, 3H), 6.58 (dd, J = 8.5, 2.2 Hz, 1H), 6.39 (d, J = 2.1 Hz, 1H), 4.16 (s, 2H), 2.94-2.84 (m, 2H), 2.85-2.75 (m , 2H).

¹³ C NMR (63 MHz, CDCl ₃ ) δ 185.98, 151.40, 146.02, 137.78, 134.89, 134.71, 132.02, 130.98, 130.44, 129.65, 129.21, 126.26, 124.52, 113.55, 112.08, 29.30, 27.37.

22) 6-amino-2-(3-chlorobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [6-amino-2-(3-chlorobenzylidene)-3,4-dihydronaphthalen-1 (2H)-one] (22)

Yield: 24.3%, yellow solid; TLC (ethyl acetate/ hexanes = 1:2v/v) R _f : 0.28, mp: 185.2-186.0 °C, HPLC: Retention time: 7.86 min, purity: 96.9%, ESI LC/MS: m/z calcd for C ₁₇ H ₁₄ ClNO [MH] ⁺ 284.07; found 284.65.

¹ H NMR (250 MHz, CDCl ₃ ) δ 7.98 (d, J = 8.5 Hz, 1H), 7.69 (s, 1H), 7.36 (s, 1H), 7.33-7.22 (m, 3H), 6.58 (dd, J = 8.5, 2.3 Hz, 1H), 6.40 (d, J = 2.1 Hz, 1H), 4.16 (s, 2H), 3.07-2.95 (m, 2H), 2.87-2.74 (m, 2H).

¹³ C NMR (63 MHz, CDCl ₃ ) δ 185.94, 151.41, 145.82, 138.10, 137.27, 134.25, 133.47, 131.00, 129.61, 129.39, 128.08, 127.88, 124.50, 113.56, 112.04, 29.10, 27.15.

23) 6-amino-2-(4-chlorobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [6-amino-2-(4-chlorobenzylidene)-3,4-dihydronaphthalen-1 (2H)-one] (23)

Yield: 24.6%, yellow solid; TLC (ethyl acetate/ hexanes = 1:2 v/v) R _f : 0.29, mp: 193.4-194.3 °C, HPLC: Retention time: 7.86 min, purity: 99.5%, ESI LC/MS: m/z calcd for C ₁₇ H ₁₄ ClNO [MH] ⁺ 284.07; found 284.65.

¹ H NMR (250 MHz, CDCl ₃ ) δ 7.97 (d, J = 8.5 Hz, 1H), 7.70 (s, 1H), 7.40-7.27 (m, 4H), 6.57 (dd, J = 8.5, 2.3 Hz, 1H), 6.39 (d, J = 2.2 Hz, 1H), 4.14 (s, 2H), 3.06-2.93 (m, 2H), 2.87-2.73 (m, 2H).

¹³ C NMR (63 MHz, CDCl ₃ ) δ 185.96, 151.40, 145.74, 136.68, 134.77, 134.01, 133.73, 130.98 (3C), 128.60 (2C), 124.64, 113.57, 112.05, 29.12, 27.17.

24) 6-amino-2-(2-bromobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [6-amino-2-(2-bromobenzylidene)-3,4-dihydronaphthalen- 1(2H)-one] (24)

Yield: 20.8%, a yellow solid; TLC (ethyl acetate/ hexanes = 2:5 v/v) R _f : 0.24, mp: 121.4-122.3 ℃, HPLC: Retention time: 8.05 min, purity: 99.7%, ESI LC/MS: m/z calcd for C ₁₇ H ₁₄ BrNO [MH] ⁺ 328.02; found 328.50.

¹ H NMR (250 MHz, CDCl ₃ ) δ 8.00 (d, J = 8.5 Hz, 1H), 7.73 (s, 1H), 7.61 (d, J = 7.9 Hz, 1H), 7.35-7.22 (m, 2H) , 7.21-7.11 (m, 1H), 6.58 (dd, J = 8.5, 2.2 Hz, 1H), 6.39 (d, J = 1.9 Hz, 1H), 4.17 (s, 2H), 2.91-2.83 (m, 2H) ), 2.83-2.76 (m, 2H).

¹³ C NMR (63 MHz, CDCl ₃ ) δ 186.03, 151.40, 146.05, 137.45, 136.72, 134.22, 132.82, 130.96, 130.46, 129.36, 126.88, 124.84, 124.48, 113.54, 112.08, 29.29, 27.28.

25) 6-amino-2-(3-bromobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [6-amino-2-(3-bromobenzylidene)-3,4-dihydronaphthalen- 1(2H)-one] (25)

Yield: 20.1%, yellow solid; TLC (ethyl acetate/ hexanes = 1:2 v/v) R _f : 0.24, mp: 177.2-178.2 ℃, HPLC: Retention time: 8.10 min, purity: 97.7%, ESI LC/MS: m/z calcd for C ₁₇ H ₁₄ BrNO [MH] ⁺ 328.02; found 328.45.

¹ H NMR (250 MHz, CDCl ₃ ) δ 8.00 (d, J = 8.5 Hz, 1H), 7.71 (s, 1H), 7.54 (s, 1H), 7.46 (d, J = 7.3 Hz, 1H), 7.36 -7.22 (m, 2H), 6.60 (dd, J = 8.5, 2.1 Hz, 1H), 6.43 (d, J = 1.7 Hz, 1H), 4.20 (s, 2H), 3.09-2.97 (m, J = 5.9 Hz, 2H), 2.90-2.78 (m, 2H).

¹³ C NMR (63 MHz, CDCl ₃ ) δ 185.90, 151.42, 145.81, 138.40, 137.31, 133.35, 132.27, 130.99, 130.98, 129.87, 128.31, 124.47, 122.40, 113.56, 112.03, 29.09, 27.12.

26) 6-amino-2-(4-bromobenzylidene)-3,4-dihydronaphthalene-1(2H)-one [6-amino-2-(4-bromobenzylidene)-3,4-dihydronaphthalen- 1(2H)-one] (26)

Yield: 28.3%, yellow solid; TLC (ethyl acetate/ hexanes = 1:2 v/v) R _f : 0.26, mp: 206.8- 207.7 ℃, HPLC: Retention time: 8.20 min, purity: 99.1%, ESI LC/MS: m/z calcd for C ₁₇ H ₁₄ BrNO [MH] ⁺ 328.02; found 328.45.

¹ H NMR (250 MHz, CDCl ₃ ) δ 7.99 (d, J = 8.5 Hz, 1H), 7.69 (s, 1H), 7.51 (dd, J = 8.3, 1.3 Hz, 2H), 7.30-7.24 (m, 2H), 6.58 (dd, J = 8.5, 1.8 Hz, 1H), 6.40 (s, 1H), 4.13 (s, 2H), 3.05-2.95 (m, J = 6.6 Hz, 2H), 2.86-2.75 (m , 2H).

¹³ C NMR (63 MHz, CDCl ₃ ) δ 185.95, 151.40, 145.76, 136.82, 135.27, 133.76, 131.59 (2C), 131.25 (2C), 131.01, 124.71, 122.24, 113.62, 112.09, 29.14, 27.20.

27) 6-amino-2-(2-(trifluoromethyl)benzylidene)-3,4-dihydronaphthalene-1(2H)-one [6-amino-2-(2-(trifluoromethyl)benzylidene) -3,4-dihydronaphthalen-1(2H)-one] (27)

Yield: 20.0%, a yellow solid; TLC (ethyl acetate/ hexanes = 1:2v/v) R _f : 0.22 mp: 144.3-145.2 °C, HPLC: Retention time: 7.43 min, purity: 99.12%, ESI LC/MS: m/z calcd for C ₁₈ H ₁₄ F ₃ NO [MH] ⁺ 318.10; found 318.55.

¹ H NMR (250 MHz, CDCl ₃ ) δ 8.00 (d, J = 8.5 Hz, 1H), 7.88 (s, 1H), 7.69 (d, J = 7.7 Hz, 1H), 7.53 (t, J = 7.4 Hz , 1H), 7.40 (t, J = 7.5 Hz, 1H), 7.28 (d, J = 7.6 Hz, 1H), 6.57 (dd, J = 8.5, 2.3 Hz, 1H), 6.38 (d, J = 2.2 Hz , 1H), 4.18 (s, 2H), 2.76 (s, 4H).

¹³ C NMR (63 MHz, CDCl ₃ ) δ 185.82, 151.51, 146.12, 138.62, 135.35, 131.51, 131.37, 130.98, 130.48, 129.14 (d, J = 30.1 Hz), 127.73, 125.95 (q, J = 5.4 Hz) , 124.35, 123.96 (d, J = 273.9 Hz), 113.55, 112.09, 29.31, 27.37.

28) 6-amino-2-(3-(trifluoromethyl)benzylidene)-3,4-dihydronaphthalene-1(2H)-one [6-amino-2-(3-(trifluoromethyl)benzylidene) -3,4-dihydronaphthalen-1(2H)-one] (28)

Yield: 20.7%, yellow solid; TLC (ethyl acetate/ hexanes = 1:3 v/v) R _f : 0.27 mp: 161.7-162.7 °C, HPLC: Retention time: 7.53 min, purity: 99.3% ESI LC/MS: m/z calcd for C ₁₈ H ₁₄ F ₃ NO [MH] ⁺ 318.10; found 318.55.

¹ H NMR (250 MHz, DMSO) δ 7.80-7.61 (m, 5H), 7.60 (s, 1H), 6.51 (dd, J = 8.5, 2.2 Hz, 1H), 6.34 (d, J = 1.9 Hz, 1H ), 6.20 (s, 2H), 2.95 (t, J = 5.8 Hz, 2H), 2.74 (t, J = 6.2 Hz, 2H).

¹³ C NMR (63 MHz, DMSO) δ 184.01, 154.30, 145.81, 138.43, 137.11, 133.57, 131.61, 130.37, 129.71, 128.83, 126.13 (d, J = 3.9 Hz), 124.72 (dd, J = 7.0, 3.0 Hz) ), 124.27 (d, J = 272.5 Hz), 121.54, 112.86, 110.73, 28.55, 26.86.

29) 6-amino-2-(4-(trifluoromethyl)benzylidene)-3,4-dihydronaphthalene-1(2H)-one [6-amino-2-(4-(trifluoromethyl)benzylidene) -3,4-dihydronaphthalen-1(2H)-one] (29)

Yield: 28.0%, yellow solid; TLC (ethyl acetate/ hexanes = 1:2 v/v) R _f : 0.22, mp: 196.4-197.4 °C, HPLC: Retention time: 7.58 min, purity: 99.8%, ESI LC/MS: m/z calcd for C ₁₈ H ₁₄ F ₃ NO [MH] ⁺ 318.10; found 318.60.

¹ H NMR (250 MHz, CDCl ₃ ) δ 7.99 (d, J = 8.5 Hz, 1H), 7.76 (s, 1H), 7.63 (d, J = 8.2 Hz, 2H), 7.47 (d, J = 8.1 Hz , 2H), 6.58 (dd, J = 8.5, 2.3 Hz, 1H), 6.40 (d, J = 2.1 Hz, 1H), 4.14 (s, 2H), 3.07-2.95 (m, 2H), 2.89-2.77 ( m, 2H).

¹³ C NMR (63 MHz, CDCl ₃ ) δ 185.80, 151.51, 145.82, 139.99, 138.08, 133.24, 131.09, 129.80 (4C), 125.29 (q, J = 3.8 Hz), 124.54, 124.08 (d, J = 272.1 Hz) ), 113.66, 112.08, 29.15, 27.18.

30) 6-amino-2-(2-(trifluoromethoxy)benzylidene)-3,4-dihydronaphthalene-1(2H)-one [6-amino-2-(2-(trifluoromethoxy)benzylidene) -3,4-dihydronaphthalen-1(2H)-one] (30)

Yield: 21.0%, yellow solid; TLC (ethyl acetate/ hexanes = 1:6 v/v) R _f : 0.22, mp: 155.7-155.6 °C, HPLC: Retention time: 7.47 min, purity: 97.9%, ESI LC/MS: m/z calcd for C ₁₈ H ₁₄ F ₃ NO ₂ [MH] ⁺ 334.09; found 334.55.

¹ H NMR (250 MHz, DMSO) δ 7.71 (d, J = 8.6 Hz, 1H), 7.57-7.41 (m, 5H), 6.52 (dd, J = 8.6, 2.0 Hz, 1H), 6.34 (d, J = 1.8 Hz, 1H), 6.23 (s, 2H), 2.87-2.76 (m, 2H), 2.77-2.68 (m, 2H).

¹³ C NMR (63 MHz, DMSO) δ 183.82, 154.40, 146.71, 145.98, 139.49, 131.24, 130.42, 130.27, 129.57, 127.67, 126.49, 121.58, 121.39, 120.30 (d, J = 257.0 Hz), 112.92, 110.78, 28.68, 27.13.

31) 6-amino-2-(3-(trifluoromethoxy)benzylidene)-3,4-dihydronaphthalene-1(2H)-one [6-amino-2-(3-(trifluoromethoxy)benzylidene) -3,4-dihydronaphthalen-1(2H)-one] (31)

Yield: 51.0%, yellow solid; TLC (ethyl acetate/ hexanes = 1:4 v/v) R _f : 0.23, mp: 152.5-153.5 °C, HPLC: Retention time: 7.60 min, purity: 99.7%, ESI LC/MS: m/z calcd for C ₁₈ H ₁₄ F ₃ NO ₂ [MH] ⁺ 334.09; found 334.55.

¹ H NMR (250 MHz, CDCl ₃ ) δ 7.98 (d, J = 8.5 Hz, 1H), 7.72 (s, 1H), 7.41 (t, J = 7.9 Hz, 1H), 7.30 (d, J = 7.7 Hz , 1H), 7.22 (s, 1H), 7.16 (d, J = 8.0 Hz, 1H), 6.58 (dd, J = 8.5, 2.3 Hz, 1H), 6.40 (d, J = 2.2 Hz, 1H), 4.17 (s, 2H), 3.07-2.96 (m, J = 9.2, 3.9 Hz, 2H), 2.87-2.77 (m, 2H).

¹³ C NMR (63 MHz, CDCl ₃ ) δ 185.86, 151.45, 149.20, 145.81, 138.29, 137.48, 133.26, 131.02, 129.72, 128.07, 124.46, 121.89, 120.47 (d, J = 257.4 Hz), 120.35, 113.58, 112.03 , 29.07, 27.08.

32) 6-amino-2-(4-(trifluoromethoxy)benzylidene)-3,4-dihydronaphthalene-1(2H)-one [6-amino-2-(4-(trifluoromethoxy)benzylidene) -3,4-dihydronaphthalen-1(2H)-one] (32)

Yield: 20.4%, yellow solid; TLC (ethyl acetate/ hexanes = 1:2v/v) R _f : 0.24, mp: 160.0-161.0 °C, HPLC: Retention time: 7.66 min, purity: 98.8%, ESI LC/MS: m/z calcd for C ₁₈ H ₁₄ F ₃ NO ₂ [MH] ⁺ 334.09; found 334.55.

¹ H NMR (250 MHz, CDCl ₃ ) δ 7.99 (d, J = 8.6 Hz, 1H), 7.75 (s, 1H), 7.42 (d, J = 8.4 Hz, 2H), 7.24 (d, J = 8.3 Hz , 2H), 6.59 (d, J = 7.4 Hz, 1H), 6.42 (s, 1H), 4.23 (s, 2H), 3.03 (t, J = 6.4 Hz, 2H), 2.88-2.77 (m, 2H) .

¹³ C NMR (63 MHz, CDCl ₃ ) δ 186.00, 151.50, 148.70, 145.78, 136.83, 134.88, 133.44, 131.11 (2C), 130.95, 124.41, 120.71 (2C), 120.42 (d, J = 257.4 Hz), 113.54 , 112.00, 29.06, 27.07.

3. Cell culture

RAW 264.7 rat macrophages were purchased from Korean cell line bank (Seoul, Korea), and 95 from Dulbecco's Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and 1% penicillin-streptomycin. % Oxygen and 5% CO _{2 It} was routinely cultured at 37°C with an incubator in a humid atmosphere. HT-29 (human colonic epithelial cell line), U937 (human pre-monocyte cell line) and CCD841 (human normal colonic epithelial cell line) were purchased from the American Type Culture Collection (Manassas, VA, USA). Cells were cultured in RPMI-1640 (HT-29 and U937) and DMEM high-glucose (CCD841) medium containing 10% FBS, 100 IU/mL penicillin and 100 μg/mL streptomycin, and 5% CO ₂ moist atmosphere Maintained at 37°C under.

4. Cell Viability Measurement (MTS analysis)

Cell viability was measured according to the manufacturer's instructions using CellTiter 96 Aqueous One Kit (Promega, Madison, WI, USA). Specifically, RAW 264.7 macrophages were inoculated into a 96-well plate at a density of 5 × 10 ⁴ cells/well. After incubating overnight, the cells were treated with the indicated compound (10 μM or more) for 24 hours, and 3-(4,5-dimethylhiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl ) 2H-tetrazolium (MTS) was further incubated for 2 hours at 37 ℃ with a solution. Cell viability was measured based on the amount of formazan production produced by metabolically activated cells, and absorbance was measured at 490 nm using a SPECTROstar Nano microplate reader (BMG Labtech Inc., Ortenberg, Germany).

5. Active Oxygen species (reactive oxygen species; ROS ) Generate measurement

ROS production was measured as previously reported. Specifically, RAW 264.7 macrophages were inoculated into a 96-well black plate at a density of 2 × 10 ⁵ /well. After overnight incubation, the cells were pretreated with compounds for 1 hour and stimulated with LPS for an additional 24 hours. Then, the cells were cultured in the dark for 30 minutes with 5-chloromethyl-2,7-dichlorodihydrofluorescein diacetate (CMH ₂ DCFDA). The residual dye was removed by washing with HBSS solution, and the fluorescence was measured using a FLUOstar OPTIMA fluorometer (BMG Labtech, Ortenberg, Germany) to confirm the total ROS generation. The excitation and emission wavelengths were set to 485 and 520 nm, respectively. The results were analyzed through FLUOstar OPTIMA software (BMG Labtech, Ortenberg, Germany).

6. Attachment of monocytes to colon epithelial cells

The analysis of monocyte adhesion to colon epithelial cells was 2',7'-bis (2-carboxyethyl)-5(6) carboxyl fluorescein acetoxymethyl ester [2',7'-bis (2-carboxyethyl)-5. (6) Carboxyl fluorescein acetoxymethyl ester] pre-labeled U937 pre-monocyte human cells were used. HT-29 cells (2 × 10 ⁵ cells/well) cultured in 48-well plates were pretreated with the compound for 1 hour. Thereafter, pre-labeled U937 cells were inoculated on a monolayer of HT-29 cells (5×10 ⁵ cells/well), and treated with TNF-α (10 ng/mL) at 37° C. for 3 hours. Non-adherent U937 cells were removed by washing 3 times with PBS. Cells were lysed at 25° C. for 30 minutes with 0.1% Triton X-100 dissolved in Tris (0.1M). Thereafter, fluorescence intensities of excitation and emission wavelengths were measured at 485 and 520 nm, respectively, using a Fluostar Optima microplate reader (BMG LABTECH GmbH, Germany).

7. Western Blotting

Cells scraped from the culture dish were centrifuged at 900 g for 5 minutes, and the obtained pellet was lysed in RIPA buffer containing protease and protease inhibitor. A BCA protein assay reagent (Pierce, Rockford, IL, USA) was used to measure the total protein concentration. Proteins were separated through SDS-PAGE, and transferred to 200 mA for 1 hour on Hybond ECL nitrocellulose membranes (Amersham Life Science, Buckinghamshire, UK). Non-specific binding was blocked for 1 hour using 5% bovine serum albumin (BSA) dissolved in Tris-buffered saline (TBS)-Tween 20 (TBS-T). The membrane was reacted overnight at 4°C with a primary antibody dissolved in BSA or skim milk-TBS, washed three times with TBS-T, and reacted with mustard peroxidase-conjugated secondary antibody at room temperature for 1 hour. . The membrane was washed, reacted with an ECL (Pierce) detection reagent, and exposed under a luminescent image analyzer, LAS-4000 mini (Fuji, Japan). β-actin was used as a loading control.

8. DSS-induced colitis

7-8 weeks old female C57BL/6 mice (n = 5) were used in the present invention. Mice were given 2% (w/v) DSS (MW 36-50 kDa; MP Biomedicals, Solon, OH, USA) as drinking water for 7 days. DSS water was changed every 3 days. Control mice were provided with water at will. From day 6 to day 14, OSI-10-1 (10 mg/kg), MYI-15-1 (10 mg/kg), MYI-18-1 (10 mg/kg), OSI-11-1 (3 And 10 mg/kg) of the drug solution was administered by the peritoneal route, and OSI-11-1 (10 and 30 mg/kg) and SSZ (300 mg/kg) were administered orally. During DSS and drug treatment, animals were examined daily for body weight, diarrhea and bleeding. On the 15th day, the animals were sacrificed and the entire colon was amputated. The large intestine was excised, some portions were stored in formalin, and the remaining portions were frozen for Western blot and MPO activity measurements.

9. NADPH Oxidase activity assay

Using Krebs-HEPES buffer (pH 7.4) containing protease and phosphatase inhibitor cocktail (Thermo Fisher Scientific, Waltham, MA, USA), HT-29 cells were harvested, homogenized with Dounce homogenizer, and 10,000×g Centrifuged for 15 minutes. Protein concentration was measured using the BCA protein assay kit (Thermo Fisher Scientific). Equal amounts of protein extract were transferred to the wells of a 96-well plate with a white coated bottom. Immediately after drug treatment, NADPH (100 μM) was added. Lucigenin (400 μM) was added, and chemiluminescence was measured using a FLUROstar Omega microplate reader (BMG Labtech GmbH, Offenburg, Germany).

< Example 1> compound synthesis

First, a halogenated 1-tetralone or 6-amino-1-tetralone derivative (1-32) was synthesized, 1-tetralone / 6-amino- It was synthesized through a condensation reaction of 1-tetrarone and halogenated aryl aldehyde (R ¹ = ap) (Scheme 1 and Scheme 2). A solution of 50% aqueous NaOH or KOH (0.5 mL) in ethanol/6-amino-1-tetralone (1.0 mmol) and halogenated aryl aldehyde (R ¹ = ap) (1.2 mmol) in ethanol (4 mL), a base-catalyzed Claisen- Schmidt condensation reaction was performed, and a pure expected chalcone derivative in a yield of 20.0-96.3% was obtained.

[Scheme 1]

[Scheme 2]

The structure of the synthesized target compound (1-32) is shown in FIGS. 3 and 4. First, of the 16 synthetic 1-tetralone chalcones, compound 1 contains a phenyl group in 1-tetralone, and compounds 2-4, 5-7, 8-10, 11-13 and 14-16 are 1- the tetralone ortho (ortho), meth (meta) or p (para) includes a phenyl group, respectively (Fig. 3) - phenyl and trifluoromethoxy phenyl, chlorophenyl, bromophenyl, trifluoroacetic acid. Similarly, among the following synthesized 6-amino-1-tetralone chalcone, compound 17 contains a phenyl group in 6-amino-1-tetralone, and compounds 18-20, 21-23, 24-26, 27-29 and 30-32 are 6-amino-1-tetralone in the ortho (ortho), meth (meta) or p (para) - phenyl, chlorophenyl, bromophenyl, tri-fluoro-phenyl and trifluoromethyl Each of the lomethoxyphenyl groups is included (Fig. 4). Depending on the different positions of the fluoro, chloro, bromo, trifluoromethyl or trifluoromethoxy group and the presence or absence of the amino group of the 1-tetralone or 6-amino-1-tetralone chalcone derivative, the structure-activity relationship (Structure -activity relationship (SAR) was determined.

< Example 2> in RAW 264.7 macrophages LPS Through stimulation ROS Production inhibitory activity

For compound 1-32, ROS production inhibitory activity was measured in RAW 264.7 macrophages through LPS stimulation, which are shown in Table 1. Most of the compounds showed strong ROS inhibitory activity, especially in 6-amino-1-tetralone substituted with a chalcone derivative. First, the 16 compound is ortho 1-16 (ortho) of fluorine, chlorine, bromine, methyl, or trifluoromethoxy-benzyl-2-ethoxy substituent in the ring chalkon fluoride in trifluoroacetic basic skeleton of 1-tetralone synthesized , meth (meta) or p (para) - there is included in the position, of the compound 1, 5 to 10 and 12 to 16 is considerably suppressed was the LPS- stimulated ROS production in macrophages RAW 264.7 for (1.27 to 6.24 μM of IC ₅₀ ). Compared to 1-tetralone (compound 1) unsubstituted at the 2-benzylidene moiety of chalcone, o -chloro, p -chloro, p -bromo, m -trifluoromethyl and p -trifluoromethoxy benzyl Compounds 5, 7, 10, 12 and 16 each having a leadene moiety significantly increased the inhibitory effect, which is o -chloro, p -chloro, p -bromo, m -trifluoromethyl and This indicates that substitution of p -trifluoromethoxy is important to strongly inhibit LPS-stimulated ROS production in RAW 264.7 macrophages. Similarly, the following synthesized compound 17-32 has a fluorine, chlorine, bromine, trifluoromethyl or trifluoromethoxy substituent in the basic skeleton of 6-amino-1-tetralone of the 2-benzylidene ring of chalcone. ortho (ortho), meth (meta) or p (para) - there is included in the location, compounds 17-32 all showed a strong inhibitory activity, most of the compounds inhibit LPS- stimulated ROS generation in macrophages RAW 264.7 for The activity was significantly increased (0.25 to 8.77 μM of IC ₅₀ ). Compounds each having o -fluoro, p -fluoro and m -trifluoromethyl benzylidene moieties compared to unsubstituted 6-amino-1-tetralone (compound 17) at the 2-benzylidene moiety of chalcone 18, 20, and 28 significantly increased the inhibitory effect, which indicated that substitution of o -fluoro, p -fluoro and m -trifluoromethyl in the 2-benzylidene site was LPS-stimulated ROS in RAW 264.7 macrophages. It indicates that it is important to strongly inhibit production. It is an unexpected result that the inhibitory effect according to the functional group substituted at the 2-benzylidene moiety of 1-tetralone and 6-amino-1-tetrarone is not consistent. Of the 32 synthesized 1-tetralone / 6-amino-1-tetralone substituted chalcone compounds (1-32), there were no compounds showing toxicity in RAW 264.7 macrophages as a result of performing 3 independent repeated experiments. .

Structure-activity relationship (SAR) analysis of 16 compounds having 1-tetrarone as the basic skeleton showed that, for LPS-stimulated ROS production in RAW 264.7 macrophages, the p -bromophenyl moiety was The compound with showed the highest inhibitory activity (1.27±0.29 μM of IC ₅₀ ), while the compound with fluoro and o -trifluoromethylphenyl moieties showed the lowest inhibitory activity (>10 μM of IC ₅₀ ). Comparing the inhibitory activity with other halogenated functional groups, p -bromophenyl substitution (1.27±0.29 μM of IC ₅₀ ) showed the highest ROS inhibition, followed by p -chlorophenyl (1.55±0.64 μM of IC _50). ), p -trifluoromethoxyphenyl (1.90±0.23 μM of IC ₅₀ ), m -trifluoromethylphenyl (3.02±0.72 μM of IC ₅₀ ) and phenyl (5.43±1.80 μM of IC ₅₀ ) in the order of substitution, respectively. . Ortho (ortho), meth (meta), and p (para) SAR results between the substitution of a halogen functional group, typically para (para) substituted This showed the highest ROS inhibition, and then the metadata (meta) and ortho (ortho) substituted Appeared in order.

Compounds ROS Inhibition
(IC ₅₀ , ^a μM) Toxicity ^b Relative ROS Inhibitory Potency of ( 1-16 ) with respect to malvidin One 5.43±1.80 NT 1.7 2 >10 NT <0.9 3 >10 NT <0.9 4 >10 NT <0.9 5 3.11±1.11 NT 2.9 6 6.24±0.99 NT 1.4 7 1.55±0.64 NT 5.8 8 5.35±0.23 NT 1.7 9 4.31±0.67 NT 2.1 10 1.27±0.29 NT 7.1 11 >10 NT <0.9 12 3.02±0.72 NT 3.0 13 4.18±1.40 NT 2.2 14 5.35±0.75 NT 1.7 15 3.99±0.51 NT 2.3 16 1.90±0.23 NT 4.7 Compounds ROS Inhibition
(IC ₅₀ , ^a μM) Toxicity ^b Relative ROS Inhibitory Potency of ( 17-32 ) with respect to malvidin 17 0.76±0.11 NT 11.8 18 0.25±0.13 NT 36.0 19 0.76±0.09 NT 11.8 20 0.40±0.15 NT 22.5 21 1.04±0.02 NT 8.7 22 1.12±0.06 NT 8.0 23 2.93±0.49 NT 3.1 24 2.88±0.81 NT 3.1 25 0.80±0.11 NT 11.3 26 1.29±0.31 NT 7.0 27 8.77±1.31 NT 1.0 28 0.41±0.04 NT 22.0 29 4.51±1.39 NT 2.0 30 3.75±1.38 NT 2.4 31 0.84±0.12 NT 10.7 32 1.06±0.29 NT 8.5 Malvidin 9.0±0.8

NT: non-toxic

^a Each data was expressed as mean±SD by performing three independent repeated experiments.

^b Relative ROS inhibition ability: IC ₅₀ (μM) of Malvidin / IC ₅₀ (μM) of synthesized compounds (1-32)

As a result of the analysis of the inhibitory effect of a compound having 6-amino-1-tetrarone as a basic skeleton (Compound 17-32), the compound with o -fluorophenyl moiety was the most for LPS-stimulated ROS production in RAW 264.7 macrophages. While exhibiting a high inhibitory effect (0.25±0.13 μM of IC ₅₀ ), the compound with o -trifluoromethyl moiety showed the lowest inhibitory effect (8.77±1.31 μM of IC ₅₀ ). 6-amino-1-tetralone of ortho halogen functional group of the compounds 17-32 having a portion (ortho), meth (meta), and p (para) and meta excluding a functional group to the SAR results, in general, according to the fluoro-substituted ( meta ) substitution showed higher inhibitory activity. There was no specific association between the different functional groups of the compound, so it was difficult to clearly measure SAR.

However, as a result of SAR analysis, it was found that the substitution of the amino group at the 6 ^th position of the 1-tetralone ring significantly improved ROS inhibition, which indicates that the 6-amino-1-tetralone moiety inhibits ROS than the 1-tetralone moiety Shows that it greatly contributes to the activity (Table 1). Comparing the inhibitory activity between 1-tetralone and 6-amino-1-tetrarone substituted with the chalcone derivative, all of the 6-amino-1-tetralone chalcone derivatives 17-32 except for compounds 23 and 29 were 1-tetralone. The level of ROS inhibition was significantly increased compared to the lonchalcon derivatives 1-16. In addition, among all halogenated functional groups, fluorine-containing 1-tetralone chalcone derivative 2-4 did not exhibit any ROS inhibitory activity, whereas 6-amino-1-tetralone chalcone derivative 18-20 each had an ROS inhibitory effect. Significantly improved (0.25 to 0.76 μM of IC ₅₀ ). In addition, compounds 18 and 20 showed the highest ROS inhibitory effect compared to all synthesized 1-tetralone and 6-amino-1-tetralone chalcone derivatives. In addition, the relative ROS inhibitory effect of 6-amino-1-tetralone chalcone (17-32) and 1-tetralone chalcone (1-16) compared to malvidin was found in 6-amino substituted compounds 17, 18, It was found that 19, 20, 25, 28, and 31 increased by 11.8, 36.0, 11.8, 22.5, 11.3, 22.0 and 10.7 times, respectively, compared to malvidin, which is the positive control. On the other hand, the non-amino substituted compounds 1, 2, 3, 4, 9, 12 and 15 were 1.7, <0.9, <0.9, <0.9, 2.1, 3.0 and 2.3 times level, respectively (Table 1). In other words, it was found that when an amino group was introduced at the 6 ^th position of 1-tetrarone chalcone, ROS inhibition in RAW 264.7 macrophages was significantly improved.

Finally, SAR analysis was performed by comparing several hard chalcone derivatives previously reported by the present inventors with 1-tetralone and 6-amino-1-tetralone substituted chalcone derivatives. Among the various hard structures, 6-amino-1-tetralone derivatives showed the greatest ROS inhibitory effect, whereas 4-chromanone and 1-indanone had moderate and weak ROS inhibitory effects, respectively. Was shown (Fig. 5).

< Example 3> 6-amino-1- Tetraron Chalcone Derivative of TNF -α-induced inhibition of monocyte adhesion to colon epithelial cells

TNF-α is one of the major pro-inflammatory mediators and acts as a major characteristic molecule in the pathogenesis of IBD. In order to confirm the inhibitory activity of compounds 17, 18, 28, and 31, which are 6-amino-1-tetrarone chalcone derivatives, against intestinal inflammation, the present inventors first described TNF-α-induced monocytes into human colon epithelial cells. The inhibitory activity on adhesion was measured. As a result, it was confirmed that when HT-29 cells were treated with TNF-α (10 ng/ml) for 3 hours, monocyte adhesion was significantly increased, which was inhibited by the 6-amino-1-tetralone chalcone derivative. Confirmed. Compound 18 showed the strongest inhibitory effect, followed by compound 28 (Fig. 6). IC ₅₀ values indicating the effect of inhibiting adhesion of HT-29 and U937 of Compound 18 and Compound 28 were 0.40 ± 0.11 μM and 0.72 ± 0.09 μM, respectively (Table 2). In CCD-841, a normal human colon epithelial cell line, treatment with compound 18 (48 h) did not show any cytotoxicity up to a concentration of 10 μM.

Compound 18 and Compound 28 inhibited TNF-α-induced NOX activity in HT-29 cells according to a concentration-dependent manner, and Compound 18 showed a much stronger effect than Compound 28. The IC ₅₀ values of Compound 18 and Compound 28 for TNF-α-induced NOX activity were 0.38 ± 0.08 μM and 0.83 ± 0.15 μM, respectively (Table 2). In addition, it was found that the inhibitory effect of Compound 18 was stronger than that of VAS-2870, a NOX2/4 inhibitor, as a positive control compound.

compound IC ₅₀ (μM) ^* Attach NADPH activity 18 0.40 ± 0.11 0.38 ± 0.08 28 0.72 ± 0.09 0.83 ± 0.15

^* Each data was expressed as mean±SD by performing 3 independent repeated experiments.

< Example 4> DSS-induced mouse colitis Improving 6-amino-1- Tetraron Chalcone derivative

The present inventors tested the association between the in vitro anti-inflammatory activity of the four compounds and the in vivo effect in the DSS-induced mouse colitis model. Administration of 2% DSS induced significant weight loss, indicating intestinal inflammation. After 5 days of DSS treatment, mice were given the drug and their therapeutic effects were compared. As a positive control, 300 mg/kg SSZ (orally administrable IBD drug) was administered orally. The three compounds were administered peritoneally (intraperitoneal; ip) for 15 consecutive days at a dose of 10 mg/kg. In the case of compound 18, the effect according to the dose dependence (3 and 10 mg/kg) in DSS-induced colitis was measured, and DSS-induced weight loss was significantly recovered in 4 compounds, and compound 18 was found to be the most effective. Administration of compound 18 via ip showed a dose-dependent recovery effect. When compound 18 was administered orally 3 times at the ip dose, it was found that the recovery effect was similar to that of the ip dose, and when 30 mg/kg of compound 18 was administered by the oral route, 300 mg/kg of sulfasalazine (SSZ) was administered orally. It was found to be more effective than administration. In the gross form test, DSS induced swelling and shortening of the colon tissue, which was recovered by administration of the compound. DSS-induced reduction in colon weight/unit length was recovered in a pattern similar to body weight recovery by compound administration. In addition, DSS significantly increased MPO activity, indicating neutrophil invasion, a biochemical indicator of colon inflammation. Neutrophil invasion was significantly reduced by these compounds, and in particular, compound 18 showed the strongest effect (FIG. 7). In colon tissue, DSS significantly increased the expression of pro-inflammatory cytokines such as TNF-α, IL-6, and IL-1β and chemokines such as IL-8 and MCP-1 protein expression, but the administration of these compounds Changes in kine and chemokine expression were significantly restored. In addition, in the DSS-treated mouse colon, the expression level of NOX2 and its regulatory subunit (p-p47/p47 phox) protein was also significantly increased compared to the control mice.

In conclusion, the present inventors systematically designed 32 halogenated 1-tetralone and 6-amino-1-tetralone chalcone derivatives and synthesized them through the Claisen- Schmidt condensation reaction. The synthesized compounds were measured for the inhibitory effect of LPS-stimulated ROS production in RAW 264.7 macrophages. Among them, compound 18 with a 6-amino-1-tetralone chalcone moiety showed the most potent inhibitory effect of LPS-stimulated ROS production in RAW 264.7 macrophages. As a result of SAR analysis, the chalcone derivatives having a 1-tetralone skeleton showed stronger ROS inhibitory effects than the previously reported chalcone derivatives derived from 1-indanone or 4-chromanone skeletons. In addition, it was confirmed that the substitution of the amino group at the 6 ^th position of the 1-tetrarone skeleton greatly increased the inhibitory effect of LPS-stimulated ROS production in RAW 264.7 macrophages. In addition, the above results demonstrate that administration of the chalcone derivative inhibits TNF-α-induced NOX2 activity and improves mouse colitis by inhibiting pro-inflammatory cytokines and cell adhesion. Inhibition of cell adhesion by compound 18 occurs by inhibition of S100A8 mediated NOX2 activation. In other words, Compound 18 could be a potent therapeutic drug for IBD. The above results can provide a new perspective for developing new anti-inflammatory agents and therapeutic agents for inflammatory bowel disease.

As described above, specific parts of the present invention have been described in detail, and for those of ordinary skill in the art, it is obvious that these specific techniques are only preferred embodiments, and the scope of the present invention is not limited thereby. something to do. Therefore, it will be said that the practical scope of the present invention is defined by the appended claims and their equivalents.

Claims (7)

A 6-amino-1-tetralone chalcone derivative compound represented by Formula 1 below, an isomer thereof, or a pharmaceutically acceptable salt thereof.
[Formula 1]

In Formula 1, R is selected from halogen, C1-C4 trifluoroalkyl and C1-C4 trifluoroalkoxy. The method of claim 1, wherein the 6-amino-1-tetralone chalcone derivative compound represented by Formula 1 is any one selected from compounds of the following compounds 18 to 32. A compound, an isomer thereof, or a pharmaceutically acceptable salt thereof.

A 6-amino-1-tetralone chalcone derivative compound represented by Formula 1 or 1-tetralone chalcone derivative compound represented by Formula 2; A pharmaceutical composition for preventing or treating inflammatory diseases containing isomers thereof or a pharmaceutically acceptable salt thereof as an active ingredient.
[Formula 1]

[Formula 2]

In Formula 1 or Formula 2, R is selected from halogen, C1-C4 trifluoroalkyl and C1-C4 trifluoroalkoxy. The method of claim 3, wherein the 6-amino-1-tetralone chalcone derivative compound represented by Formula 1 is any one selected from compounds of the following compounds 17 to 32. Pharmaceutical composition for preventing or treating inflammatory diseases.

The pharmaceutical composition for preventing or treating inflammatory diseases according to claim 3, wherein the 1-tetralone chalcone derivative compound represented by Formula 2 is any one selected from the following compounds 1 to 16 .

The method of any one of claims 3 to 5, wherein the inflammatory disease is dermatitis, conjunctivitis, periodontitis, rhinitis, otitis media, pharyngitis, tonsillitis, pneumonia, gastric ulcer, gastritis, inflammatory bowel disease, Crohn's disease, colitis, hemorrhoids, gout. , Ankylosing spondylitis, lupus, fibromyalgia, psoriatic arthritis, osteoarthritis, rheumatoid arthritis, peri-shoulderitis, tendonitis, tendonitis, peritonitis, myositis, hepatitis, cystitis, nephritis, sjogren's syndrome, multiple sclerosis and angiogenesis. A pharmaceutical composition for preventing or treating inflammatory diseases, characterized in that selected from the group consisting of inflammatory diseases caused by. A 6-amino-1-tetralone chalcone derivative compound represented by Formula 1 or 1-tetralone chalcone derivative compound represented by Formula 2; Health functional food composition for preventing or improving inflammatory diseases containing their isomers or their pharmaceutically acceptable salts as an active ingredient.
[Formula 1]

[Formula 2]

In Formula 1 or Formula 2, R is selected from halogen, C1-C4 trifluoroalkyl and C1-C4 trifluoroalkoxy.

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