KR20110136960A - Composition for preventing and treating of allergic diseases comprising 4-chlorotetrazolo[1,5-a]quinoxaline and derivatives - Google Patents

Abstract

PURPOSE: A composition containing 4-chlorotetrazolo[1,5-a]quinoxaline and derivative thereof for anti-allergic effect is provided to suppress generation of TNF-alpha and IL-4 and to prevent and treat allergic diseases. CONSTITUTION: A pharmaceutical composition for preventing and treating allergic diseases contains 4-chlorotetrazolo[1,5-a]quinoxaline and derivative thereof as an active ingredient. The allergic diseases are caused by mast cells. The 4-chlorotetrazolo[1,5-a]quinoxaline and derivative thereof prevents and treats allergic diseases by suppressing Src-family kinase activity. A functional cosmetic composition for treating allergic diseases contains 4-chlorotetrazolo[1,5-a]quinoxaline and derivative thereof as an active ingredient.

Description

COMPOSITION FOR PREVENTING AND TREATING OF ALLERGIC DISEASES COMPRISING 4-CHLOROTETRAZOLO [1,5-a] QUINOXALINE AND DERIVATIVES}

The present invention relates to a composition containing 4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives as an active ingredient represented by the following formula (1), and more specifically, allergy due to secretion of granules from mast cells. Allergic disease caused by mast cells by inhibiting the secretion of triggers and inhibiting the production of tumor necrosis factor alpha (TNF-α) and interleukin-4 (IL-4) involved in allergic reactions The present invention relates to a pharmaceutical composition and a functional cosmetic composition comprising 4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives as an active ingredient, which are used for the prophylaxis and treatment thereof.

[Formula 1]

In general, allergies are one of several classes of immune responses called hypersensitivity reactions. Allergies are caused by the body's immune system reacting to foreign substances that act as antigens, allergens. This requires repeated contact with the allergens, causing an excessive immune response by the memory cells. The main cause of allergic reactions is an abnormal increase in IgE-specific antibodies caused by various stimulatory factors, which are attached to the surface of mast cells. Several stored chemical mediators (histamine, heparin, ECF-A, NCF, TNF-α) are released. As a result of the biological action of these chemical mediators, hypersensitivity, inflammation and asthma occur.

IgE binds to FcεRI of tissue mast cells to induce degranulation of mast cells, which is secreted by histamine, heparin, rucotriene, prostaglandin and other hypersensitivity factors. These secreted immune activators activate the immune system and cause inflammation and allergic reactions.

When mast cells are activated through the combination of IgE and allergens, they secrete a variety of chemical mediators, which contribute to the initial allergic reaction by causing mucosal edema, bronchial smooth muscle contraction, and increased mucus secretion. However, mast cells are not only limited to these initial reactions but have been found to be deeply involved in late and chronic allergic reactions.

That is, TNF-α produced and secreted from mast cells increases the expression of adhesion molecules of vascular endothelial cells, thereby promoting the influx of eosinophils and T lymphocytes into target organs. In addition, it secretes tryptase, PAF, PGD2, IL-5, GM-CSF to help chemotaxis and proliferation of eosinophils. IL-4, IL-13, IL-6 and the like produced in mast cells contribute to chronic allergic reactions by increasing the response of Th2 cells and eventually increasing the production of IgE.

Bronchial hypersensitivity in asthma has been reported to be proportional to the number of mast cells infiltrating into smooth muscle as well as eosinophils (SJ Galli et al, Nature, 454 (24): 445-454, 2008). In addition, TGF-β, which is secreted from mast cells, seems to be involved in remodeling of the airways by increasing the influx and activity of fibroblasts and promoting collagen synthesis.

Mast cells play an important role in allergic inflammation, starting with IgE, FcεRI and allergens.

Allergic reactions include asthma, rhinitis, urticaria and anaphylaxis (SJ Galli et al, Nature, 454 (24): 445-454, 2008). Some allergic reactions are associated with genetic binding to specific allergens. Allergy caused by asthma and eczema. These allergies are generally characterized by harmless substances (such as pollen, mold, animal hair and dandruff, dust, ticks, and peanuts).

Currently, allergic disease drugs are mainly antagonists of receptors such as histamine and leukotriene secreted by allergens in mast cells. However, since these drugs are resistant to patients in the short term after administration, most of them are used to improve symptoms, and they are not enough to block the cause of the disease, and side effects caused by drugs are also serious (Rabe KF, et al ., Eur Respir J Suppl. , 34: 34s-40s, 2001).

In addition, as a treatment for the treatment of allergic diseases, there is a method to identify allergens for allergies suffered by allergic patients and then gradually reduce the allergies by administering them in small amounts for several years. However, these treatments have the disadvantage that the treatment takes several years, there is a problem that can cause anaphylactic shock. Other methods include the use of DNA vaccines, therapies that block IgE from binding to mast cell receptors, and antibodies to IL-4, an allergic cytokine. The disadvantage is that the effect is not identified.

Accordingly, the present inventors have made diligent efforts to secure a substance capable of treating various allergic diseases in a fundamental manner. As a result, 4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives of the present invention induce allergy from mast cells. It was confirmed that the secretion of substances is fundamentally inhibited, and the 4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives provide a use as a therapeutic agent for allergic diseases.

The present invention has been made in view of the above problems and the need for the above, an object of the present invention to provide a pharmaceutical composition and a functional cosmetic composition for the treatment and prevention of allergic diseases.

In order to achieve the above object, the present invention is 4-chlorotetrazolo [1,5-a] quinoxaline (4-chlorotetrazolo [1,5-a] quinoxaline) and its derivatives represented by the formula (1) as an active ingredient Provided are pharmaceutical compositions for the prevention and treatment of allergic diseases.

[Formula 1]

In a preferred embodiment of the present invention, the allergic disease is characterized by an allergic disease generated through the mast cell, the allergic disease is allergic rhinitis, allergic conjunctivitis, allergic asthma, allergic dermatitis , Autoimmune hepatitis, allergic bronchopulmonary aspergillosis or allergic stomatitis, and the like, but are not limited thereto.

In addition, the 4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives of the present invention are characterized by preventing the activity of Src-family kinase to prevent or treat allergic diseases, preferably Syk (spleen). Inhibition of tyrosine kinase) inhibits the activity of Src-family kinase to prevent or treat allergic diseases.

The present invention also provides a functional cosmetic composition for improving allergic diseases containing 4-chlorotetrazolo [1,5-a] quinoxaline and derivatives thereof as an active ingredient.

4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives according to the present invention inhibit the secretion of allergens from mast cells and at the same time the tumor necrosis factor-α (TNF-α) involved in allergic reactions. By inhibiting the production of) and interleukin-4 (IL-4), not only enables the prevention and treatment of the underlying allergic disease, but also has a reversible inhibitory effect on the degranulation of mast cells, and has no side effects, allergic rhinitis and allergy. It can be usefully used as an effective and safe drug for the prevention and treatment of allergic diseases caused by mast cells such as asthma and allergic dermatitis or as a composition of functional cosmetics for the prevention and improvement of allergic diseases.

1 is a photograph showing the concentration-dependent inhibitory effect of 4-chlorotetrazolo [1,5-a] quinoxaline on mast cells of the present invention, Figure 1A is 4-chlorotetrazolo to RBL-2H3 mast cells Degranulation inhibitory effect of [1,5-a] quinoxaline, Figure 1B shows the degranulation inhibitory effect of 4-chlorotetrazolo [1,5-a] quinoxaline on bone marrow-derived mast cells, Figure 1C Shows the reversible effect of mast cell degranulation inhibition.
Figure 2 is a photograph showing the allergic inhibitory effect of 4-chlorotetrazolo [1,5-a] quinoxaline of the present invention on a local allergy animal model.
3 is 4-chlorotetrazolo [1,5-a] quinoxaline of the present invention for the production of inflammatory cytokines, tumor necrosis factor (TNF-α) and interleukin-4 (IL-4), in activated mast cells It is a photograph showing the inhibitory effect.
Figure 4 is a photograph showing the results of Western blotting (Western Blotting) for the pharmacologic mechanism of 4-chlorotetrazolo [1,5-a] quinoxaline of the present invention.
5 is a photograph showing the Syk kinase specific activity inhibitory effect of 4-chlorotetrazolo [1,5-a] quinoxaline of the present invention.

Hereinafter, the present invention will be described in detail.

The present invention provides a pharmaceutical composition for the prevention and treatment of allergic diseases containing 4-chlorotetrazolo [1,5-a] quinoxaline and derivatives thereof as an active ingredient.

In the present invention, the 4-chlorotetrazolo [1,5-a] quinoxaline is represented by the following Chemical Formula 1, and its derivative is 4-hydrazino [1,5-a] quinoxaline, 4-substituted phenylaminotetra Solo [1,5-a] quinoxaline, piperazinyltetrazolo [1,5-a] quinoxaline, 4-ethoxycarbonylhydrazinotetrazolo [1,5-a] quinoxaline, 6- Hydroxy-1,2,4-triazolo [3,4-c] tetrazolo [1,5-a] quinoxaline, 4- [2- (N-substituted thiocarbamoyl) hydrazino] -tetra Solo [1,5-a] quinoxaline, 4- [N- (5-methoxycarbonylmethylene-3-substituted-4-oxo-2-thioxoimidazolidin-1-yl) amino] tetra Solo [1,5-a] quinoxaline, 4-substituted benzylidenehydrazinotetrazolo [1,5-a] quinoxaline, 4-substituted heteromethylenehydrazinotetrazolo [1,5-a] quinox Salin, 4- (5-amino-4-alkoxycarbonylpyrazol-1-yl) tetrazolo [1,5-a] quinoxaline, but the scope of the present invention is limited thereto It is not.

[Formula 1]

In the present invention, 4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives can be used as compounds isolated from nature, chemically synthesized compounds or commercially available ones.

4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives of the present invention have the effect of preventing and treating allergic diseases. The allergic diseases are diseases caused by mast cells, specifically, allergic rhinitis, allergic conjunctivitis, allergic asthma, allergic dermatitis, autoimmune hepatitis, allergic bronchial aspergillosis and allergic disease Stomatitis.

In particular, the anti-allergic effect of 4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives may be characterized by the inhibition of allergen-induced secretion of mast cells, the anti-allergic effect in allergic animal models and the mast cells. Inhibition of cytokine production and the activation of various signaling pathways that cause allergens in mast cells were significantly inhibited at low concentrations. It was found to be excellently inhibited by more than%. In addition, it has been shown to significantly and concentration-dependently inhibit the production of tumor necrosis factor-α (TNF-α) and interleukin-4 (IL-4), which are well known as cytokines secreted from mast cells. It inhibited the phosphorylation of Syk (Spleen tyrosine kinase) and LAT (Linker for activation of T cells), which transmit signals by phosphorylation among various signaling pathways that cause allergy-induced allergy-induced mast cells. Mitogen-activated protein (MAP) kinase, which plays an important role in production, has been shown to be significant and concentration-dependently inhibited.

Thus, 4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives of the present invention, unlike the conventional therapeutic agent for symptomatic recovery-oriented allergic diseases, are fundamentally responsible for the secretion of allergens in mast cells. It can be used as a treatment and prevention of allergic diseases that can be blocked.

In addition, the inhibitory effect of the 4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives on mast cell degranulation is reversible, and such reversible mast cell degranulation inhibition prevents side effects that may result from irreversible properties. Has an advantage.

Compositions comprising a compound of the present invention may further comprise suitable carriers, excipients and diluents commonly used in the manufacture of pharmaceutical compositions, and may be prepared in a pharmaceutically acceptable formulation, for example powder, It may be prepared in the form of tablets, granules, capsules or injections.

The carrier or excipient includes water, dextrin, calcium carbonate, lactose, propylene glycol, liquid, paraffin, physiological saline, dextrose, sucrose, sorbitol, mannitol, ziitol, erythritol, maltitol, starch, gelatin, calcium phosphate, calcium Silicates, cellulose, methyl cellulose, polyvinylpyrrolidone, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and minerals, which may be used one or more, but are not limited thereto. Both carriers and excipients can be used. In addition, when formulating the composition, it may further include conventional fillers, extenders, binders, disintegrants, surfactants, anti-coagulants, lubricants, wetting agents, flavors, emulsifiers or preservatives.

When the composition containing the compound according to the present invention is formulated into tablets by a conventional method, it is preferable to use lactose, microcrystalline cellulose, magnesium stearate, and the like, which are used as bases, and the compound is used in a ratio of 1: 1. Do.

In addition, the dosage of the compound according to the present invention may be changed according to body absorption, weight, age, sex, health condition, diet, administration time, administration method, excretion rate, severity of disease, and the like. However, for the desired effect, the compound of the present invention is preferably administered at 0.001 to 150 mg, preferably 0.01 to 100 mg per kg of body weight per day. Administration may be administered once a day or may be divided several times. The dosage does not limit the scope of the invention in any aspect.

The compounds of the present invention can be administered to mammals such as mice, mice, livestock and the like by various routes. All modes of administration can be expected, for example by oral, rectal or intravenous, intramuscular, subcutaneous, intrauterine dural or intracerebroventricular injection.

The present invention also provides a functional cosmetic containing the 4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives as an active ingredient, which exhibits a prophylactic and therapeutic effect of allergic diseases.

Functional cosmetics of the present invention can be used to prevent, alleviate and treat allergic diseases by blocking the secretion of allergens from mast cells.

Hereinafter, the present invention will be described in more detail with reference to Examples and Experimental Examples.

However, the following examples are merely to illustrate the invention, but the content of the present invention is not limited to the following examples.

Example 1. Investigation of the inhibitory effect of 4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives on the secretion of allergens in mast cells

In order to detect the effect of inhibiting the secretion of allergens from allergen-inducing mast cells in the body of 4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives obtained from the above examples, the following experiments were carried out.

RBL-2H3 cells were cultured in minimal medium supplemented with glutamine, antibiotics and 15% bovine serum. Cells for secretion experiments were harvested by trypsin and 200,000 cells per well in a 25-well / ml DNP- in 24-well incubator. Incubated with specific IgE. The next day the cells were washed with PIPES buffer (25 mM PIPES, pH 7.2, 159 mM NaCl, 5 mM KCl, 0.4 mM MgCl ₂ , 1 mM CaCl ₂ , 5.6 mM glucose and 0.1% BSA) and then 30 minutes before antigen was added. Pre-incubated for minutes. After pre-culture, the antigen was stimulated to a final concentration of 25 ng / ml, and the secretion of allergen-inducing substance was determined by the activity of hexosaminidase, a marker of degranulation secreted in the medium, p -nitrophenyl-acetyl- β- It was determined by the amount of p -nitrophenyl of D-glucosaminide ( p- nitrophenyl-acetyl- β- D-glucosaminide). Mouse bone marrow-derived mast cells (BMMC) were divided into 4 ml round bottom tubes by 2.0 × 10 ⁶ cells each, followed by media containing 500 ng / ml DNP-specific IgE. After culturing for 24 hours to sensitize the cells, the supernatant was discarded by centrifugation at 1,000 rpm for 5 minutes and Tyrode buffer solution (135 mM NaCl, 5 mM KCl, 20 mM HEPES, 5.6 mM glucose, 1.8 mM CaCl ₂ , 1 mM MgCl). Cells were washed once with ₂ , 0.05% BSA, pH7.4) and then transferred to a 1.5 ml tube and incubated in a 37 ° C. thermostat for 10 minutes. The cells were then stimulated with 25 ng / ml DNP-BSA (antigen) for 15 minutes, then placed on ice for 5 minutes to stop the reaction and centrifuged at 1,000 rpm for 5 minutes to transfer the supernatant to an E-tube. Cells remaining in the tube were lysed with 0.1% Trypton X-100. The soluble supernatant and cell P 1 mM in a 96-well plate-nitrophenyl -N- acetyl - β -D- glucoside Sami Need (P- nitrophenyl- N -acetyl- β -D- glucosaminide) and 30 by respective ㎕ After mixing and incubating for 1 hour on a 37 ℃ constant temperature water bath, 0.1 M carbonate buffer (Carbonate buffer) was added, the absorbance was measured at a wavelength of 405 nm.

The results are shown in Figure 1, the secretion of allergens from RBL-2H3 mast cells (Fig. 1.A) and bone marrow mast cells (Fig. 1.B) by the antigen as shown in Figure 1 4-chlorotetra The low concentration (IC ₅₀ : about 3 μM) was significantly inhibited by zolo [1,5-a] quinoxaline, and allergens were induced at 10 μM of 4-chlorotetrazolo [1,5-a] quinoxaline. It was confirmed that secretion of more than 80%.

In order to confirm the reversible effect of inhibiting mast cell activation of 4-chlorotetrazolo [1,5-a] quinoxaline, 4-chlorotetrazolo [1,5-a] quinoxaline was incubated 30 minutes before stimulation with antigen. When the antigen was treated after washing with the medium about 5 times, it was confirmed that degranulation inhibition by 4-chlorotetrazolo [1,5-a] quinoxaline was recovered as shown in FIG. 1C. These results indicate that the suppression of allergen-induced secretion from mast cells by antigen of 4-chlorotetrazolo [1,5-a] quinoxaline is reversible. Such reversible mast cell degranulation inhibition has the advantage of preventing side effects that can come from irreversible properties.

Example 2 Investigation of Antiallergic Effects in Allergic Animal Models

In order to investigate the anti-allergic effect in the allergic animal model of 4-chlorotetrazolo [1,5-a] quinoxaline of the present invention, the following experiments were carried out.

0.5 g of IgE was injected intramuscularly into one ear of an ICR mouse, followed by sensitization for 12 hours, followed by dose of 4-chlorotetrazolo [1,5-a] quinoxaline suspended in 5% arabic gum. One hour after oral administration, 250 μl of antigen solution (1 mg antigen + 5 mg Evansblue / ml PBS solution) was injected through the tail vein. 30 minutes after the injection, the ears of the mouse were removed and extracted for 12 hours or more in a formamide solution of 700 ° C. at 63 ° C., and the extracted Evansblue was measured for absorbance at 620 nm to confirm an antiallergic effect.

The results are shown in FIG. 2, and as shown in FIG. 2, the 4-chlorotetrazolo [1,5-a] quinoxaline of the present invention inhibited allergic reactions due to antigen by 42% in the 10 mg / kg administration group. In the 50 mg / kg administration group, 60% was inhibited. Therefore, it was confirmed that 4-chlorotetrazolo [1,5-a] quinoxaline of the present invention has an excellent anti-allergic effect even in an allergic animal model induced by the antigen.

Example 3. Investigation of the inhibitory effect of 4-chlorotetrazolo [1,5-a] quinoxaline on cytokine production on mast cells

In order to examine the cytokine production inhibitory effect of 4-chlorotetrazolo [1,5-a] quinoxaline on mast cells of the present invention, the following experiment was performed.

Inhibition of 4-chlorotetrazolo [1,5-a] quinoxaline against tumor necrosis factor-α (TNF-α) and interleukin-4 (IL-4), well known cytokines secreted from allergic mast cells In order to examine the effect, RT-PCR was performed using primers specific for each cytokine. First, stimulated RBL-2H3 cells in 6-well plates were lysed with trizol reagent (Invitrogen), and then the lysed samples were transferred to 1.5 ml E-tubes, mixed with 200 µl of chloroform and centrifuged at 12,000 g for 20 minutes. To give a supernatant. After mixing the supernatant and isopropanol in a 1: 1 ratio, the RNA was precipitated at -20 ° C. for at least one hour, centrifuged at 12,000 g for 20 minutes to completely precipitate the RNA, and then washed once with ethanol, and then washed with 0.1. The total RNA was obtained by dissolving in 50 µl of% DEPC-water. PCR (polymerase chain reaction) was repeated 30 times 45 seconds at 94 ℃, 45 seconds at 55 ℃, 60 seconds at 72 ℃, primers, rat TNF-α is forward 5'-CAC CAC GCT CTT CTG TCT ACT GAA C -3 '(SEQ ID NO: 1), reverse is 5'-CCG GAC TCC GTG ATG TCT AAG TAC T-3' (SEQ ID NO: 2), IL-4 is forward 5'-CCG ATT ATG GTG TAA TTT CCT ATG CTG- 3 '(SEQ ID NO: 3), reverse is 5'-GGC CAA TCA GCA CCT CTC TTC CAG-3' (SEQ ID NO: 4), and rat GAPDH forward is 5'-GTG GAG TCT ACT GGC GTC TTC-3 '(SEQ ID NO: 5), reverse was 5'-CCA AGG CTG TGG GCA AGG TCA-3 '(SEQ ID NO: 6).

Inhibition of gastric cytokine secretion after the pretreatment as described above was confirmed by ELISA method after collection.

The results are shown in Figure 3, as shown in Figure 3 of the tumor necrosis factor (TNF-α) and interleukin-4 (IL-4) at 1 μM of 4-chlorotetrazolo [1,5-a] quinoxaline It was found that production began to decrease and was almost suppressed at 10 μM or less. Accordingly, it was confirmed that cytokine production was significantly and concentration-dependently inhibited by the 4-chlorotetrazolo [1,5-a] quinoxaline of the present invention.

Example 4 Investigation of Mechanism of Action of 4-Chlorotetrazolo [1,5-a] quinoxaline on Mast Cells

Investigation of 4-chlorotetrazolo [1,5-a] quinoxaline obtained from the above-described examples shows the effect of inhibiting activation of Syk-LAT in various signaling pathways that cause allergen induction in allergic mast cells. In order to do this, the experiment was carried out as follows.

After RBL-2H3 cells were dispensed at a well of 1.0 × 10 ⁶ per 6 well plate, the cells were sensitized by culturing for 12 hours in MEM medium containing 20 ng / ml DNP specific immunoglobulin E. The cells were pretreated with 4-chlorotetrazolo [1,5-a] quinoxaline for 30 minutes and the cells stimulated with antigen were washed twice with cold PBS, followed by Lysis buffer (20 mM HEES, pH 7.5). , 150 mM NaCl, 1% Nonidet p-40, 10% glycerol, 60 mM octyl- β -glucoside, 10 mM NaF, 1 mM Na ₃ VO ₄ , 1 mM phenylmethylsulfonyl fluoride, 2.5 mM nitrophenylphosphate, 0.7 μg / ml pepstatin And protein inhibition purification). Next, the mixture was centrifuged at 13,000 rpm for 5 minutes, and the supernatant was mixed with 2 × Laemmli sample buffer to proceed with protein denaturation at 100 ° C. for 5 minutes. The protein sample thus obtained was subjected to SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel) for 1 hour and 30 minutes at 120 V using a 10% or 12% poly acrylamide gel (polyacrylamide: bis acrylamide = 29: 1). After electrophoresis, the cells were transferred to a nitrocellulose membrane (nitrocellulose membrane, Schleicher & Schuell, BA85) at 300 mA for 1 hour 30 minutes. 5% skim milk or 5% BSA was made with TBS-T (10 mM Tris-HCl, pH 7.5, 150 mM NaCl, 0.05% Tween-20), blocked for 1 hour, and then each specific antibody And reacted for 1 hour. After reacting with monoclonal or polyclonal antibody with horseradish peroxidase (HRP) for 1 hour, the remaining nonspecific antibody was washed off with TBS-T and removed using X-ray using ECL detection kit (Amersham Pharmacia Biotech). It was photosensitive to the film.

The results are shown in FIG. 4, and as shown in FIG. 4, the phosphorylation level of LAT at 1 μM of 4-chlorotetrazolo [1,5-a] quinoxaline of the present invention began to decrease, and at 3 μM and 10 μM, respectively. It was found that phosphorylation of protein was almost suppressed. In addition, it was confirmed that MAP kinase, which plays an important role in the production of cytokines in the body, was also significantly and concentration-dependently inhibited by 4-chlorotetrazolo [1,5-a] quinoxaline.

Example 5 Investigation of the Inhibition of Src-family Kinase

Phosphorylation of Syk (spleen tyrosine kinase), a substrate of Src-family kinase (c-Src, Fgr, Fyn), an early signal transduction agent, was confirmed by immunoprecipitation.

The results are shown in FIG. 5, and since phosphorylation of Syk, a substrate of SFK, which is an initial substance for granular secretion signal transduction, is inhibited, 4-chlorotetrazolo [1,5-a] quinoxaline is used for It is anticipated that the secretion can be blocked at the source.

<110> Konkuk University Industrial Cooperation Corp. <120> Composition for Preventing and Treating of Allergic Diseases          Comprising 4-chlorotetrazolo [1,5-a] quinoxaline and Derivatives <130> P10-E143 <160> 6 <170> KopatentIn 1.71 <210> 1 <211> 25 <212> DNA <213> Artificial Sequence <220> <223> PCR primer for rat TNF-a forward <400> 1 caccacgctc ttctgtctac tgaac 25 <210> 2 <211> 25 <212> DNA <213> Artificial Sequence <220> <223> PCR primer for rat TNF-a reverse <400> 2 ccggactccg tgatgtctaa gtact 25 <210> 3 <211> 27 <212> DNA <213> Artificial Sequence <220> <223> PCR primer for IL-4 forward <400> 3 ccgattatgg tgtaatttcc tatgctg 27 <210> 4 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> PCR primer for IL-4 reverse <400> 4 ggccaatcag cacctctctt ccag 24 <210> 5 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> PCR primer for rat GAPDH forward <400> 5 gtggagtcta ctggcgtctt c 21 <210> 6 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> PCR primer for rat GAPDH reverse <400> 6 ccaaggctgt gggcaaggtc a 21

Claims (8)

A pharmaceutical composition for the prevention and treatment of allergic diseases containing 4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives as an active ingredient. The method of claim 1,
The derivatives include 4-substituted phenylaminotetrazolo [1,5-a] quinoxaline, piperazinyltetrazolo [1,5-a] quinoxaline, 4-ethoxycarbonylhydrazinotetrazolo [1, 5-a] quinoxaline, 6-hydroxy-1,2,4-triazolo [3,4-c] tetrazolo [1,5-a] quinoxaline, 4- [2- (N-substituted thio Carbamoyl) hydrazino] -tetrazolo [1,5-a] quinoxaline, 4- [N- (5-methoxycarbonylmethylene-3-substituted-4-oxo-2-thioxoimidazoli Din-1-yl) amino] tetrazolo [1,5-a] quinoxaline, 4-substituted benzylidenehydrazinotetrazolo [1,5-a] quinoxaline, 4-substituted heteromethylenehydrazinotetra It is either a solo [1,5-a] quinoxaline or 4- (5-amino-4-alkoxycarbonylpyrazol-1-yl) tetrazolo [1,5-a] quinoxaline. Pharmaceutical composition for the prevention and treatment of allergic diseases. The method of claim 1,
The allergic disease is a pharmaceutical composition for the prevention and treatment of allergic diseases, characterized in that allergic disease generated by the mediation of mast cells. The method according to claim 1 or 3,
The allergic disease may be allergic rhinitis, allergic conjunctivitis, allergic asthma, allergic dermatitis, autoimmune hepatitis, allergic bronchial aspergillosis or allergic stomatitis. Pharmaceutical composition for. The method of claim 1,
The 4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives inhibit the activity of Src-family kinase to prevent or treat allergic diseases, the pharmaceutical composition for the prevention and treatment of allergic diseases. . 6. The method of claim 5,
The 4-chlorotetrazolo [1,5-a] quinoxaline and its derivatives inhibit the phosphorylation of Syk (spleen tyrosine kinase) to prevent and treat allergic diseases characterized in that they inhibit the activity of Src-family kinase. Pharmaceutical composition for. Functional cosmetic composition for improving allergic diseases containing 4-chlorotetrazolo [1,5-a] quinoxaline and derivatives thereof as an active ingredient. The method of claim 7, wherein
The derivatives include 4-substituted phenylaminotetrazolo [1,5-a] quinoxaline, piperazinyltetrazolo [1,5-a] quinoxaline, 4-ethoxycarbonylhydrazinotetrazolo [1, 5-a] quinoxaline, 6-hydroxy-1,2,4-triazolo [3,4-c] tetrazolo [1,5-a] quinoxaline, 4- [2- (N-substituted thio Carbamoyl) hydrazino] -tetrazolo [1,5-a] quinoxaline, 4- [N- (5-methoxycarbonylmethylene-3-substituted-4-oxo-2-thioxoimidazoli Din-1-yl) amino] tetrazolo [1,5-a] quinoxaline, 4-substituted benzylidenehydrazinotetrazolo [1,5-a] quinoxaline, 4-substituted heteromethylenehydrazinotetra It is either a solo [1,5-a] quinoxaline or 4- (5-amino-4-alkoxycarbonylpyrazol-1-yl) tetrazolo [1,5-a] quinoxaline. Functional cosmetic composition for improvement of allergic diseases.

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