KR100916716B1 - Novel 3-chloro-5-substituted-quinoxaline-2-amine derivatives and pharmaceutically acceptable salt thereof, method for preparation, therapeutic agent for antiinflammatory disease induced by spc activity containing 3-chloro-5-substituted-quinoxaline-2-amine derivatives as an effective ingredient - Google Patents

Abstract

The present invention is a 3-chloro-5-substituted-quinoxalin-2-amine derivative represented by the following formula (1) and a pharmaceutically acceptable salt thereof, a method for preparing the same and inflammation caused by SPC receptor activity containing the same as an active ingredient And its use useful for related diseases.

The 3-chloro-5-substituted-quinoxalin-2-amine derivative of the present invention was found to have anti-inflammatory effects through animal experiments using dermal cells and mice derived from humans. Thus, by containing the 3-chloro-5-substituted-quinoxalin-2-amine derivative and a pharmaceutically acceptable salt thereof as an active ingredient, inflammation, pruritus or atopic dermatitis caused by SPC receptor activity, other diseases or It can be usefully used as a therapeutic agent for inflammation-related diseases effective for skin infections.

(Wherein R ¹ and R ² are as defined in the specification).

Description

3-Chloro-5-substituted-quinoxalin-2-amine derivatives and pharmaceutically acceptable salts thereof, preparation methods thereof, and agents for treating inflammation-related diseases caused by SPC receptor activity containing the same as active ingredients {NOVEL 3-CHLORO- 5-SUBSTITUTED-QUINOXALINE-2-AMINE DERIVATIVES AND PHARMACEUTICALLY ACCEPTABLE SALT THEREOF, METHOD FOR PREPARATION, THERAPEUTIC AGENT FOR ANTIINFLAMMATORY DISEASE INDUCED BY SPC ACTIVITY CONTAINING 3-CHLORO-5-SUBSTITUTED-QUINODER AQUITATE

The present invention relates to a 3-chloro-5-substituted-quinoxaline-2-amine derivative, a pharmaceutically acceptable salt, a method for preparing the same, and an agent for treating inflammatory diseases related to SPC receptor activity containing the same as an active ingredient, and more particularly. Preferably, as a novel compound showing SPC receptor inhibitory activity, 3-chloro-5-substituted-quinoxalin-2-amine derivatives and pharmaceutically acceptable salts are provided, and the derivatives and pharmaceutically acceptable salts thereof are effective. It relates to a therapeutic agent for inflammation-related diseases containing as a component.

Spinosylphosphorylcholine (SPC) is a lysophosphatidic acid (LPA) with structurally similar spinphosine-1-phosphate (S1P), lysophosphatidic acid (LPA), etc. It is classified as a (lysophospholipid) family of substances, and these substances act as important signaling intermediates for immune functions such as cell proliferation, migration, and inflammatory responses.

SPC is produced by the action of spingomyelin deacylase from spingomyelin, a constituent of the cell membrane [Higuchi K, Biochem J. , 2000 , 350 , 747-56]. In addition, SPC can be used for growth and proliferation of various cells [Desai, Biochem. Biophys. Res. Commun ., 1991 , 181 , 361-366, angiogenesis [Boguslawski, Biochem. Biophys. Res. Commun ., 2000 , 272 , 603-609, and cell death [Jeon ES, Biochim Biophys Acta ., 2005 , 1734 (1) ; 25-33] is known to be deeply involved.

Representative examples of SPC-related diseases are atopic dermatitis. Atopic dermatitis is due to a decrease in the lipid content of the stratum corneum, the antimicrobial activity is reduced and the barrier function is weakened, so that the defense against external stimulants is reduced, the inflammatory reaction occurs and itching appears accordingly. Itching also causes a secondary infection, resulting in a vicious cycle of hyperimmune reactions.

In particular, SPC is known to be rarely present or detected in very low concentrations in normal people, but it is known to increase thousands of times in the epidermis of atopic dermatitis patients. Higuchi K, Biochem. J. , 2000 , 350 , 747-756 and Reiko Okamoto, Journal of Lipid Research, 2003 , 44 , 93-102. This is caused by the lack of intercellular lipids in the stratum corneum of atopic dermatitis patients [Junko Hara, J. invest. Dermatol ., 2000 , 115 , 406-413. In addition, SPC plays an important role in abnormal keratinization, a characteristic characteristic of atopic diseases (Higuchi, J. Lipid Res ., 2001, 42, 1562-1570). These findings suggest that SPC is not only a direct cause of skin barrier dysfunction, which is one of the major symptoms of atopic dermatitis, but also a secondary inflammatory response. Based on this, if the production of SPC is regulated, it can be used as a new treatment for dermatitis.

Meanwhile, among the symptoms of atopic dermatitis, itching, which causes pain and poor quality of life, has been reported that LPA (lysophosphatidic acid) structurally similar to SPC causes itching [Hashimoto, Pharmacology , 2004 , 72 , 51-56]. Therefore, it can be inferred that SPC can also cause itching in the body, and in recent years, it has been demonstrated that intradermal injection of SPC can directly cause itching [WO 06/049451].

Therefore, the present inventors have searched for a novel compound that can be applied as a therapeutic agent for inflammatory diseases, and designed and synthesized the 3-chloro-5-substituted-quinoxalin-2-amine derivative compound, and the derivatives inhibited SPC receptor inhibitory activity. In view of the fact that it has not been reported yet, the prepared 3-chloro-5-substituted-quinoxaline-2-amine derivatives were conducted in animal experiments using human-derived dermal cells and mice, showing excellent anti-inflammatory effects. By confirming the effect, the present invention was completed.

It is an object of the present invention to provide 3-chloro-5-substituted-quinoxalin-2-amine derivatives and pharmaceutically acceptable salts thereof and methods for their preparation using organic synthesis techniques.

It is another object of the present invention to provide a use thereof as a therapeutic agent for inflammatory diseases related to SPC receptor activity, containing the 3-chloro-5-substituted-quinoxaline-2-amine derivative and a pharmaceutically acceptable salt thereof as an active ingredient. It is.

In order to achieve the above object, the present invention provides a 3-chloro-5-substituted-quinoxalin-2-amine derivative represented by the following formula (1) and a pharmaceutically acceptable salt thereof.

Formula 1

(Wherein R ¹ is hydrogen, a C ₁ -C ₁₀ straight or crushed alkyl group or C ₁ -C ₁₀ alkoxy group, or halogen, R ² is hydrogen, 1-4 C ₁ -C ₁₀ straight chain) Or a pulverized alkyl group, a C _{1 to} C ₁₀ alkoxy group, a hydroxy group, a halogen group, an amine group, or a substituted amine group, an amide group, a carbamate group, or a urea group, wherein the substituted group is C _{1 to} C ₁₀ Any of the substituents selected from the group consisting of linear or pulverized or cyclic alkyl groups, or halogen atoms, nitro groups, C ₁ -C ₁₀ alkyl groups, C ₁ -C ₁₀ alkoxy groups and C ₁ -C ₁₀ haloalkyl groups Represents 1 to 4 substituted phenyl groups.)

More preferably, the 3-chloro-5-substituted-2-amine derivatives of quinoxaline advantage, R ¹ is a group a linear or branched alkyl group or alkoxy group, or a fluoroalkyl group of C ₁ ~C _5, R ² is 1 to 2 is to use any one of C ₁ ~C ₅ straight or branched chain alkyl groups selected from an alkoxy group, a nitro group, a halogen group, or the group consisting of C ₁ ~C ₅ of.

The present invention provides a process for the preparation of 3-chloro-5-substituted-quinoxalin-2-amine derivatives, carried out using organic synthesis techniques.

In addition, the present invention provides a therapeutic agent for inflammatory diseases caused by SPC receptor activity containing the 3-chloro-5-substituted-quinoxaline-2-amine derivative represented by Formula 1 and a pharmaceutically acceptable salt thereof as an active ingredient. to provide.

Furthermore, the present invention contains a 3-chloro-5-substituted-quinoxaline-2-amine derivative represented by the formula (1) and a pharmaceutically acceptable salt thereof as an active ingredient, for inhibiting scar formation after trauma and promoting wound healing. Provided for use as a therapeutic agent for dermatitis diseases.

The present invention also provides a 3-chloro-5-substituted-quinoxaline-2-amine derivative represented by the formula (1) and a pharmaceutically acceptable salt thereof as an active ingredient, as a chemotactic migration-mediated symptom modulator of cells. To provide.

The present invention provides 3-chloro-5-substituted-quinoxalin-2-amine derivatives and pharmaceutically acceptable salts thereof having excellent inhibitory activity against SPC receptors, and wherein the derivatives are derived from human dermal cells and The anti-inflammatory effect was searched through animal experiments using mice to identify excellent inhibitory activity on SPC receptor, thereby providing a use as a therapeutic agent for inflammatory diseases related to SPC receptor activity.

Hereinafter, the present invention will be described in detail.

The present invention provides 3-chloro-5-substituted-quinoxalin-2-amine derivatives represented by the following Chemical Formula 1 and pharmaceutically acceptable salts thereof.

Formula 1

(Wherein R ¹ and R ² are as defined above).

More preferably, the 3-chloro-5-substituted-2-amine derivatives of quinoxaline advantage, R ¹ is a group a linear or branched alkyl group or alkoxy group, or a fluoroalkyl group of C ₁ ~C _5, R ² is 1 to 2 is to use any one of C ₁ ~C ₅ straight or branched chain alkyl groups selected from an alkoxy group, a nitro group, a halogen group, or the group consisting of C ₁ ~C ₅ of.

The present invention can provide a method for preparing a 3-chloro-5-substituted-quinoxalin-2-amine derivative represented by Formula 1, which can be obtained by the following Scheme 1.

(Wherein R ¹ and R ² are as defined above).

Hereinafter, a method for preparing the 3-chloro-5-substituted-quinoxalin-2-amine derivative represented by Chemical Formula 1 of the present invention will be described in detail. A 1,2-diamino compound substituted with R ¹ represented by Chemical Formula 2 will be described. Reacting with oxalic acid to synthesize quinoxaline-2,3-dione represented by Formula 3 wherein various R ¹ are introduced;

A second step of synthesizing 2,3-dicloquinoxaline represented by Chemical Formula 4 by reacting the compound represented by Chemical Formula 3 with thionyl chloride; And

In the 2,3-dichloro substituent of the compound represented by Formula 4, 2-chloro substituent and 4-amino-1-arylmethylpiperidine substituted with R ² represented by Formula 5 are reacted with Formula 1 Comprising a third step of synthesizing a 3-chloro-5-substituted-quinoxalin-2-amine derivative represented by.

Referring to the reaction process according to the invention, the composition of the solvent system and the selection range of the reaction conditions in detail.

In the first step, water (H ₂ O), dioxane, or tetrahydrofuran (THF) is used as a solvent, and water is preferably used. As reactants, 1,2-diamine compounds having R ¹ substituents as defined above are used. Oxalic acid used in the reaction is preferably used in about 1.5 equivalents, preferably in the range of about 1 equivalent is excellent in economic efficiency. At this time, hydrochloric acid (HCl) or sulfuric acid (H ₂ SO ₄ ) and the like may be used as the acid, and it is most preferable to use 3N hydrochloric acid.

In the second step, chloroform (CHCl ₃ ) or 1,2-dichloroethane (DCE) is used as a solvent, and chloroform is preferably used. In this reaction, it is preferable to use about 5 equivalents of thionyl chloride, and it is preferable to use it in the range of about 3 equivalents preferably. At this time, the reaction is further effective by adding a catalytic amount of dimethylformamide (DMF).

In the third step reaction, dimethyl sulfoxide (DMSO), DMF, chloroform, or dichloromethane (CH ₂ Cl ₂ ) is used as a solvent, and DMSO is preferably used. In this reaction, it is preferable to use about 1.2 equivalents of 4-amino-1-arylmethylpiperidine substituted with R ² represented by the formula (5), and preferably, in an amount of about 1.0 equivalents, it is excellent in economy. Wherein R ² used is substituted 4-amino-1-arylmethylpiperidine as defined above.

In addition, in the process of producing the 3-chloro-5-substituted-quinoxalin-2-amine derivative represented by the formula (1) according to the invention the reaction progress was confirmed by the TLC method for each intermediate reaction, the formulas 3 and 4 Reaction intermediates represented by were separated and purified, respectively, and analyzed and confirmed the structure by NMR or Mass spectrum.

The present invention provides a therapeutic agent for inflammatory diseases caused by SPC receptor activity, which contains the 3-chloro-5-substituted-quinoxalin-2-amine derivative represented by Chemical Formula 1 and a pharmaceutically acceptable salt thereof as an active ingredient. .

More preferably, the inflammatory disease treatment agent is N- (1-benzylpiperidin-4-yl, which is a compound of 1-1, in the 3-chloro-5-substituted-quinoxalin-2-amine derivative represented by the formula (1). ) -3-chloro-5-methoxyquinoxalin-2-amine or a compound of 1-13 3-chloro-N- [1- (3,5-diphenoxybenzyl) piperidin-4-yl] -5-methoxyquinoxalin-2-amine as an active ingredient.

Cell proliferation response to the 3-chloro-5-substituted-quinoxalin-2-amine derivative of the present invention was observed to confirm the antagonistic action on the selective cell proliferation response induced by SPC [ Table 2 ] is effective for dermatitis such as atopic dermatitis induced by excessive cell division proliferative response by SPC. In addition, excessive cell division proliferation causes scarring due to an inflammatory response that occurs when the wound is wound, but the 3-chloro-5-substituted-quinoxalin-2-amine derivative of the present invention is responsible for cell division proliferation. It can be used to prevent unwanted scar formation. At the same time, it can be used to promote wound healing after trauma.

In addition, the 3-chloro-5-substituted-quinoxalin-2-amine derivative compound of the present invention, as a result of the inflammatory reaction test induced by TPA, both ear edema and MPO activity was suppressed, and is commonly used as an anti-inflammatory drug. A comparable effect is shown for hydrocortisone [ Table 4 ].

Accordingly, the 3-chloro-5-substituted-quinoxalin-2-amine derivative of the present invention is effective for atopic dermatitis, inflammation, pruritus, skin infections, etc., which are shown in other diseases, and for preventing scar formation after trauma and promoting wound healing. It can be usefully used as a therapeutic agent for inflammatory diseases.

Furthermore, the present invention provides the use of a 3-chloro-5-substituted-quinoxalin-2-amine derivative represented by the formula (1) and a pharmaceutically acceptable salt thereof as a potent agent for chemotactic migration-mediated symptoms of cells. to provide.

The chemotactic migration of the cells is a phenomenon in which endothelial cells or immune cells are moved by a specific cytokine or chemokine, etc. in vivo, and immune cells are moved to an inflammation site or angiogenesis by endothelial cell migration ( angiogenesis) takes place.

Accordingly, the 3-chloro-5-substituted-quinoxalin-2-amine derivative represented by Chemical Formula 1 of the present invention was measured for chemotactic migration of cells induced by SPC, and thus, endothelial cells or immunity in vivo. The result of strongly inhibiting the cell migration reaction can be obtained [ Table 3 ]. Therefore, the chemotactic migration-mediated symptom modulator of cells containing the 3-chloro-5-substituted-quinoxalin-2-amine derivative represented by the general formula (1) of the present invention and a pharmaceutically acceptable salt thereof as an active ingredient is endothelial cells. Angiogenesis caused by migration has been suppressed, and the process of amplifying immune responses against externally invading antigens can be controlled.

Especially preferably, the chemotactic migration-mediated symptom modulator of the cells of the present invention is N- (1-benzylpiperidine-4 in the 3-chloro-5-substituted-quinoxalin-2-amine derivative represented by Formula 1 above. -Yl) -3-chloro-5-methoxyquinoxalin-2-amine as an active ingredient.

In the above, specific examples of the chemotactic migration-mediated symptoms of the cells in the chemotactic migration-mediated symptom modulator of cells of the present invention include atopic dermatitis, inflammation, pruritus and skin infections occurring in other diseases.

Pharmaceutically acceptable salts in the present invention can be prepared by conventional methods in the art, for example, salts with inorganic acids such as hydrochloric acid, hydrogen bromide, sulfuric acid, sodium hydrogen sulfate, phosphoric acid, carbonic acid, and the like. Or pharmaceutically acceptable organic compounds, such as formic acid, acetic acid, oxalic acid, benzoic acid, citric acid, tartaric acid, gluconic acid, gestyic acid, fumaric acid, lactobionic acid, salicylic acid, or acetylsalicylic acid (aspirin) It is also possible to form salts of acids, or to react with alkali metal ions such as sodium and potassium to form their metal salts, or to react with ammonium ions to form another form of a pharmaceutically acceptable salt.

In addition, the pharmaceutical composition of the present invention is a conventional non-toxic pharmaceutically acceptable carrier, adjuvant and excipient, etc. to the 3-chloro-5-substituted-quinoxalin-2-amine derivatives and pharmaceutically acceptable salts thereof. It can be formulated into a conventional formulation in the pharmaceutical field, for example, oral administration such as tablets, capsules, troches, solutions, suspensions or parenteral administration.

In addition, the dosage of the compound according to the present invention to the human body may vary depending on the age, weight, sex, dosage form, health condition and degree of disease of the patient, and generally based on an adult patient having a weight of 70 Kg. It is 0.01-1000 mg / day, and can be dividedly administered once a day to several times at regular time intervals according to the judgment of a doctor or a pharmacist.

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

The following examples are merely illustrative of the present invention, but the scope of the present invention is not limited to the following examples.

Example 1 Synthesis of 3-chloro-5-substituted-quinoxalin-2-amine derivative (Formula 1-1)

Step 1: Reaction of 1,2-Diamino Compound (2) with Oxalic Acid

3-methoxybenzene-1,2-diamine compound represented by Formula 2-1 (1.86 g, 13.5 mmol) and oxalic acid (1.23 g, 13.6 mmol) were added to 10 ml of 3N hydrochloric acid, and stirred at room temperature for 10 minutes. The reaction was carried out at reflux for 3 days. After completion of the reaction, the mixture was cooled to room temperature and filtered. The filtered compound was washed repeatedly with cold water to obtain a quinoxaline-2,3-dione compound (2.26 g, 87%) represented by Chemical Formula 3-1.

¹ H NMR (500 MHz, DMSO-d ₆ ) δ 3.86 (s, 3H), 6.75 (d, J = 8.1 Hz, 1H), 6.80 (d, J = 8.0 Hz, 1H), 7.05 (t, J = 8.2 Hz, 1H), 11.24 (br s, 1 H), 11.91 (br s, 1 H); m / z ([M + 1] ⁺ ) 139.

Step 2: 2,3-dichloroquinoxalination reaction of quinoxaline-2,3-dione (3-1)

After dissolving 5-methoxyquinoxaline-2,3 (1H, 4H) -dione compound (2.26 g, 11.7 mmol) represented by Chemical Formula 3-1 in 10 ml of chloroform and 0.3 ml of catalytic amount of DMF, the mixture was dried at room temperature. Onyl chloride (SOCl ₂ ; 2.60 mL, 35.1 mmol) was added. After reacting under reflux for 3 days, the mixture was cooled to room temperature, distilled under reduced pressure, and purified by silica gel column chromatography to obtain a 2,3-dichloroquinoxaline compound represented by Chemical Formula 4-1 (1.74 g, 65%). .

¹ H NMR (500 MHz, CDCl ₃ ) δ 4.13 (s, 3H), 7.19 (d, J = 7.9 Hz, 1H), 7.64 (d, J = 8.1 Hz, 1H), 7.77 (t, J = 8.2 Hz , 1H); m / z ([M + l] ⁺ ) 228.

Step 3: reaction of adding amine (5-1) of 2,3-dichloroquinoxaline (4-1)

2,3-dichloro-5-methoxyquinoxaline compound represented by Formula 4-1 (60 mg, 0.26 mmol) and 1-benzylpiperidin-4-amine compound represented by Formula 5-1 (54 mg, 0.28 mmol ) Was added to a 3 ml solution of DMSO and stirred at room temperature for 10 minutes, and then reacted at 60 ° C. for 2 days. After completion of the reaction, the reaction mixture was dissolved in 20 ml of EtOAc and washed with 20 ml of brine, the organic layer was dried over Na ₂ SO ₄ and concentrated after filtration. The concentrated product was purified by silica gel column chromatography (hexane: EtOAc = 1: 1) to obtain 3-chloro-5-methoxyquinolin-2-amine (1-1, 41 mg, 41%) as a target compound.

¹ H NMR (500 MHz, CDCl ₃ ) δ 1.60-1.67 (m, 2H), 2.12-2.17 (m, 2H), 2.25-2.31 (m, 4H), 2.87-2.89 (m, 2H), 3.56 (s , 2H), 4.01 (s, 3H), 4.12-4.16 (m, 1H), 5.43-5.45 (m, 1H), 6.77-6.78 (m, 1H), 7.26-7.30 (m, 1H), 7.31-7.34 (m, 5H), 7.46-7.50 (m, 1H); m / z ([M + 1] ⁺ ) 383.

Examples 2 to 40 Synthesis of 3-chloro-5-substituted-quinoxalin-2-amine derivatives.

In the compound represented by Formula 1, 3-chloro-5-substituted-quinoxaline- was carried out in the same manner as in Example 1, except that R ¹ and R ² were carried out as described in Table 1 below. 2-amine derivatives were synthesized.

In addition, the analysis results of the synthesized 3-chloro-5-substituted-quinoxalin-2-amine derivatives are shown in Table 1.

Experimental Example 1 Control of Cell Line Division Proliferation Response

Treatment of spinosilphosphorylcholine (SPC) with cells induces an excessive cell division proliferative response, which may be a cause of pathological symptoms of skin diseases such as atopic dermatitis [Desai, Biochem. Biophys. Res. Commun. , 1991 , 181 , 361-366. Thus, for the compounds prepared in the above example, the effect on the division proliferation response of the cells induced by SPC was tested.

Primarily in NIH 3T3 cells (American Type Culture Collection, Manassas, VA, USA), 1 × 10 ⁵ pieces (usually 1 × 10 ⁴ ~10 ⁶ pieces) and then seeded in culture plates incubated, containing the bovine serum for 24 hours Serum was depleted (serum starvation) by culturing in RPMI medium. Herein, as the compound prepared in the above examples and the control compound, FTY720, which is an agonist of spingosine-1-phosphate (S1P), is 0.001 μM, 0.01 μM, 0.1 μM, and 1 μM. After incubation for 30 minutes, SPC (Biomol, Plymouth Meeting, PA, USA) was added to 7 μM concentration and incubated at 37 ℃ for 24 hours. Cell proliferation was measured using the [3H] -thymidine reaction method inserted into the DNA strand to be replicated at the time of cell division [Beales IL, Life Sci ., 2004, 75, 83-95]. It calculated by 1 and shows the result in Table 2 .

As shown in Table 2, the compounds prepared in the examples of the present invention showed an antagonistic action on the selective cell proliferative response induced by SPC to the cell proliferation response. Since excessive cell division proliferation due to inflammatory reactions occurring during the wounding process when wounded, scars form, a substance that inhibits cell division proliferation may be used to prevent the formation of unnecessary scars. At the same time, agents that enhance cell proliferation can be used to promote wound healing after trauma.

As a result, it was found that Examples 1-1 and 1-16 inhibit cell division proliferation response by SPC in a dose-dependent manner. It is noteworthy that this effect has a chemical structure similar to that of SPC, and FTY720, an S1P agonist that shares some membrane receptors, did not inhibit the SPC-induced cell proliferation. Therefore, the cell division proliferation inhibitory effect is due to the inherent structural activity of the compound of the present invention, and it is possible to suppress the phenomenon of excessive cell division proliferation due to the inflammatory response occurring during the wound healing process. I think you can.

Experimental Example 2 Inhibitory Effect on Chemotactic Cell Migration Response Induced by SPC

Recently, studies have shown that SPC plays an important role in chemotactic cell migration similarly to vascular endothelial growth factor (VEGF) (Boguslawski et al ., Biochem Biophys Res Commun ., 2000, 272, 603-609). The movement of cells by attracting certain cytokines or chemokines, etc. in vivo is a key step in the process of immune cells moving to inflammatory sites or angiogenesis by endothelial cell migration. In response to chemotactic migration of cells induced by SPC, the effects of the compounds prepared in the above examples were tested by a Boyden chamber assay.

First, a 25 × 80 mm polycarbonate membrane (Neuro Probe, Inc.) having an 8 μm pore was immersed in a 0.01% gelatin, 0.1% acetic acid solution and coated overnight, followed by natural drying at room temperature.

Meanwhile, human umbilical vein endothelial cells (HUVEC) cultured in complete EBM-2 medium containing 2% fetal bovine serum were cultured in EBM-2 medium (Cambrex, catalog number CC-3121) containing no serum for 4 hours. The serum was depleted (serum starvation) and then harvested with trypsin / EDTA solution. HUVEC cells were suspended in EBM-2 medium containing 0.1% bovine serum albumin and then aliquoted into silicone coated Eppendorf tubes to yield 0, 0.1, 1 to 10 μg / ml of the compound of Example 1-1 as experimental drug. It was added in the amount of and treated for 30 minutes at 37 ℃. Dispense 27 μL of EBM-2 medium or 10 μM SPC or EBM-2 medium into each well of the lower chamber of the Boyden chamber and place the membrane prepared with gelatinized coating with the glossy side facing down. The gasket was put up and the upper chamber was assembled. Drug treated HUVEC cells were dispensed in 5 × 10 ⁴ (56 μl volume) of the upper chamber and incubated for 8 hours in a 37 ° C. CO ₂ incubator. The membrane was separated, dyed with Diff-Quik dye (Sysmex Corporation), washed with deionized water, and glued to the slide glass with the shiny side facing down. The cells adhered to the top of the membrane were carefully wiped with a Kim wiper or a cotton swab, and then photographed randomly 5 fields per well (200 times magnification) to measure the number of cells. The reaction inhibition rate was calculated according to the following Equation 2, and the results are shown in Table 3 .

As shown in Table 3, it was found that the compound of Example 1-1 of the present invention strongly inhibits the chemotactic migration reaction of cells by SPC. This suggests that by inhibiting the migration of endothelial cells or immune cells in vivo, it is possible to regulate processes such as angiogenesis inside tumors and amplification of immune responses against externally invading antigens.

Experimental Example 3 Inflammation Response Control in Mouse TPA-Induced Ear Inflammation Model

In order to confirm the inhibitory effect of the inflammatory response, the following experiment was performed using a Tetradecanoyl phorbol acetate (TPA) -induced inflammation model. TPA-induced inflammation models have been widely used to test the mechanism of action of inflammatory responses and the efficacy of inhibitors (De Young LM et al., Agents and Actions , 1989, 26, 335-341). TPA is a substance that causes an inflammatory response. When applied to the ear of a subject, erythema and edema occur, and this inflammatory response can be measured by increasing activity of MPO (myeloperoxidase), which is essential for bacterial attack of white blood cells.

40 male 6-week-old ICR mice were prepared, and 20 μl of TPA solution (Sigma Aldrich Korea) dissolved in acetone at 125 μg / ml was applied to the left ear. After 1 hour of TPA application, 20 μl of acetone, 0.3% Example Compounds dissolved in acetone, or 0.3% hydrocortisone (Sigma Aldrich Korea) dissolved in acetone was applied to the TPA application site, and the same site 6 hours after TPA application. 20 μl was reapplied. At 24 hours after TPA application, the mice were euthanized by cervical dislocation, and the left ear was collected at a constant width to measure weight and MPO activity. Reaction inhibition rate was computed according to following formula (3), and is shown in Table 4 .

As shown in Table 4, the compounds of Example 1-1 significantly inhibited the inflammatory response induced by TPA on both measures of ear edema and MPO activity, the potency of which was widely used as an anti-inflammatory drug. Comparable to hydrocortisone. The experimental results indicate that the skin anti-inflammatory efficacy of the compound of Example 1-1 was achieved by inhibiting the infiltration of neutrophils into the particularly inflamed ear region.

< Formulation example  1> Preparation of tablet (press type)

As an active ingredient, 5.0 mg of the compound represented by Chemical Formula 1 of the present invention was sieved, and then tableted by mixing and pressing 14.1 mg of lactose, 0.8 mg of crospovidone USNF and 0.1 mg of magnesium stearate.

Preparation Example 2 Preparation of Tablet (Wet Granulation)

As an active ingredient, 5.0 mg of the compound represented by the formula (1) of the present invention was sieved, and then 16.0 mg of lactose and 4.0 mg of starch were mixed. 0.3 mg of Polysorbate 80 was dissolved in pure water and then an appropriate amount of this solution was added and then atomized. After drying, the fine particles were sieved and mixed with 2.7 mg of colloidal silicon dioxide and 2.0 mg of magnesium stearate. The granules were pressed into tablets.

Preparation Example 3 Preparation of Powder and Capsule

As an active ingredient, 5.0 mg of the compound represented by Formula 1 of the present invention was sieved, and then mixed with 14.8 mg of lactose, 10.0 mg of polyvinyl pyrrolidone, and 0.2 mg of magnesium stearate. No. solid the mixture using a suitable device. Filled in 5 gelatin capsules.

Preparation Example 4 Preparation of Injection

As an active ingredient, 100 mg of the compound represented by the formula (1) of the present invention was contained, and in addition, 180 mg of mannitol, 26 mg of Na ₂ HPO ₄ .12H ₂ O, and 2,974 mg of distilled water were used to prepare an injection.

As described above, the present invention

First, 3-chloro-5-substituted-quinoxalin-2-amine derivatives and pharmaceutically acceptable salts thereof were provided using solid phase combinatorial chemical synthesis techniques,

Second, the anti-inflammatory effect of the 3-chloro-5-substituted-quinoxalin-2-amine derivatives in human dermal cells and mice was searched to find an excellent inhibitory activity against the SPC receptor.

Third, an inhibitor of the SPC receptor containing the 3-chloro-5-substituted-quinoxalin-2-amine derivative as an active ingredient, and furthermore, its use as a therapeutic agent for inflammatory diseases related to the SPC receptor activity.

Although the present invention has been described in detail only with respect to the embodiments described, it will be apparent to those skilled in the art that various modifications and variations are possible within the technical spirit of the present invention, and such modifications and variations belong to the appended claims. .

Claims (7)

3-Chloro-5-substituted-quinoxalin-2-amine derivatives having excellent inhibitory activity against the SPC receptor represented by the following formula (1) and pharmaceutically acceptable salts thereof. Formula 1 (Wherein R ¹ is hydrogen, a C ₁ -C ₁₀ straight or crushed alkyl group or C ₁ -C ₁₀ alkoxy group, or halogen, R ² is hydrogen, 1-4 C ₁ -C ₁₀ straight chain) Or a pulverized alkyl group, a C _{1 to} C ₁₀ alkoxy group, a hydroxy group, a halogen group, an amine group, or a substituted amine group, an amide group, a carbamate group, or a urea group, wherein the substituted group is C _{1 to} C ₁₀ Any of the substituents selected from the group consisting of linear or pulverized or cyclic alkyl groups, or halogen atoms, nitro groups, C ₁ -C ₁₀ alkyl groups, C ₁ -C ₁₀ alkoxy groups and C ₁ -C ₁₀ haloalkyl groups Represents 1 to 4 substituted phenyl groups.) According to claim 1, in the formula (1), R ¹ is a C ₁ ^~ C ₅ linear or crushed alkyl group or alkoxy group, or a fluoro group, R ² is 1 to 2 C ₁ ~ C ₅ linear or The 3-chloro-5-substituted-quinoxalin-2-amine derivatives and pharmaceuticals, which are selected from the group consisting of a ground alkyl group, a C _{1 to} C ₅ alkoxy group, a nitro group, or a halogen group Acceptable salts thereof. delete It contains a 3-chloro-5-substituted-quinoxalin-2-amine derivative represented by the formula (1) and a pharmaceutically acceptable salt thereof as an active ingredient, atopic dermatitis caused by SPC receptor activity, pruritus and Inflammatory disease therapeutic agent useful for any one inflammatory disease selected from the group consisting of skin infections. The method of claim 4, wherein the 3-chloro-5-substituted-quinoxalin-2-amine derivative represented by Formula 1 is N- (1-benzylpiperidin-4-yl) -3-chloro-5-meth Methoxyquinoxalin-2-amine or 3-chloro-N- [1- (3,5-dimethoxybenzyl) piperidin-4-yl] -5-methoxyquinoxalin-2-amine The inflammatory disease treatment. delete Inhibiting scar formation and promoting wound healing after trauma, comprising as an active ingredient the 3-chloro-5-substituted-quinoxaline-2-amine derivative represented by Formula 1 and a pharmaceutically acceptable salt thereof. For the treatment of dermatitis diseases.