KR20030055777A - Novel 1-pyridino-5-phenyl- pyrazole derivatives - Google Patents

Abstract

PURPOSE: 1-pyridino-5-phenyl-pyrazol derivatives having a selective inhibition effect on cyclooxygenase-2 and a preparation process thereof are provided, which compounds are useful for treatment of inflammatory diseases. CONSTITUTION: 1-pyridino-5-phenyl-pyrazol derivatives are characteristically represented by the formula(1), wherein R1 is hydrogen, methyl, or trifluoromethyl; R2 is halogen, C1-C3 alkyl, halogenomethyl, C1-C3 alkoxy, hydroxy, hydroxycarbonyl or nitro; and n is an integer of 1 to 5. A process for preparing the 1-pyridino-5-phenyl-pyrazol derivatives comprises reacting hydrazine derivatives of the formula(2) with a compound of the formula(3) in a solvent in the presence of acid catalyst.

Description

Novel 1-pyridino-5-phenyl-pyrazole derivatives}

The present invention relates to a 1-pyridino-5-phenyl-pyrazole derivative having excellent selectivity as a cyclooxygenase-2 inhibitor.

Most of the nonsteroidal anti-inflammatory drugs show their anti-inflammatory, analgesic, and antipyretic effects through inhibition of enzymes called cyclooxygenase or prostaglandin G / H synthase, and also inhibit hormonal contractions caused by hormones and Inhibits growth Initially only cyclooxygenase-1, a constitutive enzyme found in cattle, was known, and recently an inducible form of cyclooxygenase-2 has been identified. Cyclooxygenase-2 is clearly different from cyclooxygenase-1 and is easily induced by mitogen, endotoxin, hormones, growth factors and cytokines and the like.

Prostaglandins play a pathological and physiological role. Cyclooxygenase-1, a constitutive enzyme, is involved in the secretion of basic endogenous prostaglandins and plays an important role in physiological aspects such as maintaining gastrointestinal status and blood circulation in the kidneys. Cyclooxygenase-2, on the other hand, is caused by inflammatory factors, hormones, growth factors and cytokines and the like, and thus plays a major role in the pathological effects of prostaglandins. Therefore, inhibitors selective for cyclooxygenase-2 are expected to have no side effects due to mechanisms of action, and anti-inflammatory, analgesic, and antipyretic effects, and also hormone-induced uterine contraction, compared to conventional nonsteroidal anti-inflammatory drugs. Is expected to inhibit the growth of some types of cancer. In particular, side effects such as gastrointestinal toxicity and kidney toxicity are expected to be low. It is also expected to prevent the synthesis of contractile prostanoids and thus inhibit the contraction of smooth muscles induced by prostanoids, and thus are expected to be useful in diseases related to premature birth, menstrual irregularities, asthma and eosinophils. In addition, it is expected to be useful in the treatment of osteoporosis, glaucoma, colorectal cancer and dementia. John Vane, "Towards a better aspirin" in Nature , Vol, for the usefulness of inhibitors selective to cyclooxygenase-2 .367, pp 215-216, 1994; Bruno Battistini, Regina Botting and YS Bakhle, "COX-1 and COX-2: Toward the Development of More Selective NSAIDs" in Drug News and Perspectives , Vol. 7, pp501-512, 1994 David B. Reitz and Karen Seibert, "Selective Cyclooxygenase Inhibitors" in Annual Reports in Medicinal Chemistry , James A. Bristol, Editor, Vol. 30, pp179-188, 1995).

Existing drugs known as inhibitors that selectively act on cyclooxygenase-2 take a wide variety of forms in their structure. The structure most commonly studied and thus designed with the most candidates is the structure of the diaryl heterocycle, i.e., a tricyclic system, which is essentially a sulfonamide or methanesulfone group in one phenyl. The initial material of this structure is Dup697 (Bioorganic & Medicinal Chemistry Letters, Vol 5, No. 18, p2123, 1995), and then SC-58635 (Journal of Medicinal Chemistry, Vol 40, p1347) having a pyrazole structure as a derivative thereof. , 1997), MK-966 (WO 95/00501) having a furanon structure, and the like have been published.

Under these technical backgrounds, the present inventors conducted a continuous study to develop novel compounds having excellent selectivity as inhibitors of cyclooxygenase-2, and as a result, the characteristic structure of the conventional compounds, ie, the following formula containing sulfonamide groups The 1-pyridino-5-phenyl-pyrazole derivative of 1 meets this purpose and has been completed.

Accordingly, an object of the present invention is to provide a 1-pyridino-5-phenyl-pyrazole derivative of formula 1 or a pharmaceutically acceptable salt thereof.

It is another object of the present invention to provide a 1-pyridino-5-phenyl-pyrazole derivative of formula (1) or a pharmaceutically acceptable salt thereof.

Hereinafter, the present invention will be described in more detail.

The present invention relates to a 1-pyridino-5-phenyl-pyrazole derivative of the formula (1) or a pharmaceutically acceptable salt thereof.

[Formula 1]

In the above formula,

R ¹ represents hydrogen, methyl or trifluoromethyl,

R ² represents halogen, C ₁ -C ₃ -alkyl, halogenomethyl, C ₁ -C ₃ -alkoxy, hydroxy, hydroxycarbonyl or nitro,

n represents the number of 0-5.

In the above substituent definition according to the invention halogen represents fluorine, chlorine, bromine and the like, alkyl represents methyl, ethyl and the like, alkoxy represents methoxy, ethoxy and the like.

The compounds according to the invention may also exist in the form of pharmaceutically acceptable salts. The pharmaceutically acceptable salts herein mean pharmaceutically acceptable non-toxic salts including organic salts and inorganic salts. Inorganic salts include salts such as aluminum, ammonium, calcium, copper, iron, lithium, magnesium, manganese, potassium, sodium and zinc, of which ammonium, calcium, magnesium, potassium and sodium salts are preferred. Organic salts include salts prepared from primary, secondary or tertiary amines, substituted amines, cyclic amines, basic ion exchange resins, etc., present in nature, and examples thereof include arginine, betaine, caffeine, choline , N, N-dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, N-methylglu Carmine, glucamine, glucosamine, histidine, hydramine, N- (2-hydroxyethyl) piperidine, N- (2-hydroxyethyl) pyrrolidine, isopropylamine, lysine, methylglucamine, mother Polyline, piperazine, piperidine, polyamine resin, procaine, purine, theobromine, triethylamine, trimethylamine, tripropylamine, tromethamine and the like can be mentioned.

If the compounds according to the invention are basic, they may also be prepared in the form of salts of pharmaceutically acceptable non-toxic acids, including organic and inorganic acids. Examples of such acids are acetic acid, adipic acid, aspartic acid, 1,5-naphthalenedisulfonic acid, benzenesulfonic acid, benzoic acid, camphorsulfonic acid, citric acid, 1,2-ethanedisulfonic acid, ethanesulfonic acid, ethylenediaminetetraacetic acid , Fumaric acid, glucoheptonic acid, gluconic acid, glutamic acid, hydroiodic acid, hydrobromic acid, hydrochloric acid, isetionic acid, lactic acid, maleic acid, malic acid, manderic acid, methanesulfonic acid, music acid, 2-naphthalenedisulfonic acid, nitric acid , Oxalic acid, parnoic acid, pentothenic acid, phosphoric acid, pivalic acid, propionic acid, salicylic acid, stearic acid, succinic acid, sulfuric acid, tartaric acid, p-toluenesulfonic acid, undecanoic acid, 10-undecenoic acid, etc. Hydrobromic acid, hydrochloric acid, maleic acid, methanesulfonic acid, phosphoric acid, sulfuric acid and tartaric acid are preferred.

Among the compounds of the formula (1) which exhibit cyclooxygenase-2 inhibitory activity, preferred are those where R ¹ represents hydrogen, methyl or trifluoromethyl, R ² represents methyl or methoxy, and n represents a number from 0 to 3. Compound.

Particularly preferred compounds are those in which R ¹ represents methyl or trifluoromethyl.

Representative compounds among the compounds of the formula (1) include the following compounds.

6- (5-phenyl-3-trifluoromethyl-pyrazol-1-yl) -pyridine-3-sulfonamide;

6- (5-p-tolyl-3-trifluoromethyl-pyrazol-1-yl) -pyridine-3-sulfonamide;

6- [5- (4-methoxy-phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide;

6- [5- (4-Fluoro-phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide;

6- [5- (4-Chloro-phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide;

6- [5- (2,4-Dichloro-phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide;

6- [5- (2,4-Dichloro-5-fluoro-phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide; And

6- (3-Methyl-5-phenyl-pyrazol-1-yl) -pyridine-3-sulfonamide.

On the other hand, the compound of formula 1 according to the present invention can be prepared according to the method as described below. However, methods for preparing the compounds according to the present invention, for example, reaction conditions such as reaction solvents and acids are not limited to those described below, and various synthesis methods described in the present specification or disclosed in the publicly known literature are known. It can be easily prepared by arbitrarily combining, and such combination is a common technique generalized to those skilled in the art to which the present invention belongs.

The compound of formula 1 according to the present invention may be prepared by reacting a hydrazine derivative of formula 2 with a compound of formula 3 in the presence of an acid catalyst in a solvent.

In the above formula

R ¹ , R ² and n are as defined above.

Hydrazine derivatives of formula (II) used as starting materials in the process according to the invention can be prepared by known methods (USP 4,204,870 and J. Org. Chem ., Vol. 56, No. 16, 1991, 4974). . Compounds of formula 3 can also be prepared by known methods ( J. Med . Chem ., Vol. 40, 1997, 1347).

Hereinafter, a method for preparing the compound of Formula 1 according to the present invention will be described in more detail.

The reaction of the hydrazine derivative of the formula (2) with the 1,3-diketone compound of the formula (3) is carried out under the same conditions regardless of the kinds of the substituents R ¹ and R ² .

As the reaction solvent, one kind selected from inert organic solvents commonly used in organic synthesis, that is, dichloromethane, chloroform, tetrahydrofuran, dimethylformamide, benzene, toluene, diethyl ether and alcohols such as methanol, ethanol and isopropanol It is preferable to use the above, and it is most preferable to use double ethanol.

The reaction is essentially in the presence of an acid catalyst. Therefore, the same result is obtained by reacting a hydrazine derivative of Formula 2 with a compound of Formula 3 in the form of a salt with hydroiodic acid, hydrobromic acid or hydrochloric acid, or by separately adding an acid such as hydroiodic acid, hydrobromic acid, hydrochloric acid and the like.

In the reaction, the reaction may be completed by heating up to the boiling point of the solvent used at room temperature. Most preferably, the reaction is completed by heating and refluxing at boiling point using ethanol as a solvent.

After the reaction is completed, the product can be separated and purified by conventional workup methods such as chromatography, recrystallization and the like.

The compound of formula 1 according to the present invention has a selective inhibitory activity against cyclooxygenase-2 and can be usefully used as an inhibitor of this enzyme. Compounds of formula (I) having selective inhibitory activity on cyclooxygenase-2 can be used as a substitute for typical nonsteroidal anti-inflammatory drugs. In particular, as an alternative drug with improved side effects of conventional nonsteroidal anti-inflammatory drugs, peptic ulcer and gastritis It is useful in patients with partial enteritis, ulcerative colitis, diverticulitis, gastrointestinal bleeding, hypoprothrombinemia, etc. and is expected to be useful as a therapeutic agent for inflammatory diseases such as osteoarthritis and rheumatoid arthritis.

Hereinafter, the present invention will be described in more detail with reference to the following Examples and Experimental Examples. However, these Examples and Experimental Examples are only for better understanding of the present invention, and the scope of the present invention is not limited by them in any sense.

Example 1

Preparation of 6- (5-phenyl-3-trifluoromethyl-pyrazol-1-yl) -pyridine-3-sulfonamide

700 mg (3.72 mmol) of 6-hydrazino-pyridine-3-sulfonamide and 804 mg (3.72 mmol) of 1-phenyl-4,4,4-trifluorobutane-1,3-dione were suspended in 70 ml of ethanol and Then, 1.7 ml of an ethanol solution saturated with hydrochloric acid gas was added, followed by heating and reflux for about 12 hours. After cooling to room temperature, the solvent was distilled off under reduced pressure. The residue was diluted with 100 mL of dichloromethane, washed with 100 mL of water and 100 mL of saturated brine, dried over anhydrous magnesium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: ethyl acetate / n-hexane = 1/4, v / v) to give 710 mg (yield 52%) of the title compound as white crystals.

¹ H NMR (400 MHz, CDCl ₃ ) δ 5.05 (s, 2H), 6.75 (s, 1H), 7.25-7.45 (m, 5H), 7.85 (d, J = 8 Hz, 1H), 8.30 (d, J = 8 Hz, 1H), 8.75 (s, 1H)

Melting point: 125-128 ℃

Example 2

Preparation of 6- (5-p-tolyl-3-trifluoromethyl-pyrazol-1-yl) -pyridine-3-sulfonamide

700 mg (3.72 mmol) of 6-hydrazino-pyridine-3-sulfonamide and 856 mg (3.72 mmol) of 1-p-tolyl-4,4,4-trifluorobutane-1,3-dione in 70 ml of ethanol Suspension was added 1.7 mL of an ethanol solution saturated with hydrochloric acid gas, and then heated and refluxed for about 12 hours. After cooling to room temperature, the solvent was distilled off under reduced pressure. The residue was diluted with 100 mL of dichloromethane, washed with 100 mL of water and 100 mL of saturated brine, dried over anhydrous magnesium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: ethyl acetate / n-hexane = 1/4, v / v) to give 782 mg (yield 55%) of the title compound as white crystals.

¹ H NMR (400 MHz, CDCl ₃ ) δ 2.41 (s, 3H), 5.00 (s, 2H), 6.73 (s, 1H), 7.15- 7.18 (m, 4H), 7.85 (d, J = 8.6Hz, 1H ), 8.25-8.29 (m, 1 H), 8.80 (s, 1 H)

Melting point: 104-105 ℃

Example 3

Preparation of 6- [5- (4-methoxy-phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide

700 mg (3.72 mmol) of 6-hydrazino-pyridine-3-sulfonamide and 916 mg (3.72 mmol) of 1- (4-methoxy-phenyl) -4,4,4-trifluorobutane-1,3-dione It was suspended in 70 ml of ethanol, 1.7 ml of ethanol solution saturated with hydrochloric acid gas was added, and then heated and refluxed for about 12 hours. After cooling to room temperature, the solvent was distilled off under reduced pressure. The residue was diluted with 100 mL of dichloromethane, washed with 100 mL of water and 100 mL of saturated brine, dried over anhydrous magnesium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: ethyl acetate / n-hexane = 1/4, v / v) to give 711 mg (48% yield) of the title compound as white crystals.

¹ H NMR (400 MHz, CDCl ₃ ) δ 3.85 (s, 3H), 5.10 (s, 2H), 6.70 (s, 1H), 6.90 (d, J = 8 Hz, 2H), 7.20 (d, J = 8 Hz, 2H), 7.80 (d, J = 9 Hz, 1H), 8.25 (d, J = 9 Hz, 1H), 8.80 (s, 1H)

Melting Point: 136-138 ℃

Example 4

Preparation of 6- [5- (4-Fluoro-phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide

700 mg (3.72 mmol) of 6-hydrazino-pyridine-3-sulfonamide and 871 mg (3.72 mmol) of 1- (4-fluoro-phenyl) -4,4,4-trifluorobutane-1,3-dione It was suspended in 70 ml of ethanol, 1.7 ml of ethanol solution saturated with hydrochloric acid gas was added, and then heated and refluxed for about 12 hours. After cooling to room temperature, the solvent was distilled off under reduced pressure. The residue was diluted with 100 mL of dichloromethane, washed with 100 mL of water and 100 mL of saturated brine, dried over anhydrous magnesium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: ethyl acetate / n-hexane = 1/4, v / v) to give 647 mg (yield 45%) of the title compound as white crystals.

¹ H NMR (400 MHz, CDCl ₃ ) δ 5.00 (s, 2H), 6.75 (s, 1H), 7.10 (t, J = 9Hz, 2H), 7.30 (t, J = 9Hz, 2H), 7.95 (d, J = 8Hz, 1H), 8.35 (d, J = 8Hz, 1H), 8.75 (s, 1H)

Melting Point: 141-144 ℃

Example 5

Preparation of 6- [5- (4-Chloro-phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide

700 mg (3.72 mmol) of 6-hydrazino-pyridine-3-sulfonamide and 932 mg (3.72 mmol) of 1- (4-chloro-phenyl) -4,4,4-trifluorobutane-1,3-dione It was suspended in 70 ml of ethanol, 1.7 ml of ethanol solution saturated with hydrochloric acid gas was added, and then heated and refluxed for about 12 hours. After cooling to room temperature, the solvent was distilled off under reduced pressure. The residue was diluted with 100 mL of dichloromethane, washed with 100 mL of water and 100 mL of saturated brine, dried over anhydrous magnesium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: ethyl acetate / n-hexane = 1/4, v / v) to give 914 mg (61% yield) of the title compound as white crystals.

¹ H NMR (400 MHz, CDCl ₃ ) δ 4.80-5.40 (br s, 2H), 6.75 (s, 1H), 7.20 (d, J = 9 Hz, 2H), 7.40 (d, J = 9 Hz, 2H), 7.95 (d, J = 9 Hz, 1H), 8.35 (d, J = 9 Hz, 1H), 8.75 (s, 1H)

Melting Point: 168-170 ℃

Example 6

Preparation of 6- [5- (2,4-Dichloro-phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide

700 mg (3.72 mmol) of 6-hydrazino-pyridine-3-sulfonamide and 1,057 mg (3.72) of 1- (2,4-dichloro-phenyl) -4,4,4-trifluorobutane-1,3-dione mmol) was suspended in 70 mL of ethanol, 1.7 mL of an ethanol solution saturated with hydrochloric acid gas was added, and then heated and refluxed for about 12 hours. After cooling to room temperature, the solvent was distilled off under reduced pressure. The residue was diluted with 100 mL of dichloromethane, washed with 100 mL of water and 100 mL of saturated brine, dried over anhydrous magnesium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: ethyl acetate / n-hexane = 1/3, v / v) to give 651 mg (40% yield) of the title compound as white crystals.

¹ H NMR (400 MHz, CDCl ₃ ) δ 4.95 (s, 2H), 6.75 (s, 1H), 7.30-7.40 (m, 2H), 7.45 (s, 1H), 8.15 (d, J = 8 Hz, 1H) , 8.35 (d, J = 8 Hz, 1H), 8.65 (s, 1H)

Melting Point: 166-168 ℃

Example 7

Preparation of 6- [5- (2,4-Dichloro-5-fluoro-phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide

700 mg (3.72 mmol) 6-hydrazino-pyridine-3-sulfonamide and 1- (2,4-dichloro-5-fluoro-phenyl) -4,4,4-trifluorobutane-1,3- 1127 mg (3.72 mmol) of dione were suspended in 70 mL of ethanol, 1.7 mL of an ethanol solution saturated with hydrochloric acid gas was added, and then heated and refluxed for about 12 hours. After cooling to room temperature, the solvent was distilled off under reduced pressure. The residue was diluted with 100 mL of dichloromethane, washed with 100 mL of water and 100 mL of saturated brine, dried over anhydrous magnesium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: ethyl acetate / n-hexane = 1/3, v / v) to give 945 mg (56% yield) of the title compound as white crystals.

¹ H NMR (400 MHz, CDCl ₃ ) δ 4.90 (br s, 2H), 6.75 (s, 1H), 7.25 (d, J = 8 Hz, 1H), 7.50 (d, J = 7 Hz, 1H), 8.15 (d , J = 8 Hz, 1H), 8.35 (d, J = 8 Hz, 1H), 8.65 (s, 1H)

Mass (FAB) 455.0 (M + 1)

Melting point: 123-126 ℃

Example 8

Preparation of 6- (3-methyl-5-phenyl-pyrazol-1-yl) -pyridine-3-sulfonamide

700 mg (3.72 mmol) of 6-hydrazino-pyridine-3-sulfonamide and 602 mg (3.72 mmol) of 1-benzoylacetone were suspended in 70 ml of ethanol, and 1.7 ml of ethanol solution saturated with hydrochloric acid gas was added. Heated to reflux. After cooling to room temperature, the solvent was distilled off under reduced pressure. The residue was diluted with 100 mL of dichloromethane, washed with 100 mL of water and 100 mL of saturated brine, dried over anhydrous magnesium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: ethyl acetate / n-hexane = 1/2, v / v) to give 653 mg (yield 56%) of the title compound as white crystals.

¹ H NMR (400 MHz, CDCl ₃ ) δ 2.40 (s, 3H), 4.95 (s, 2H), 6.35 (s, 1H), 7.30-7.40 (m, 5H), 7.70 (d, J = 8Hz, 1H) , 8.15 (d, J = 8 Hz, 1H), 8.75 (s, 1H)

Mass (FAB) 315.0 (M + 1), 629.0 (2M + 1)

Melting Point: 165-167 ℃

Experimental Example

Selective Inhibitory Activity on Cyclooxygenase-2

1. Experiment Method

In order to pharmacologically verify the selective inhibitory activity on the cyclooxygenase-2 enzyme of the compound according to the present invention, the enzyme inhibitory activity on cyclooxygenase-1 and cyclooxygenase-2 is as follows. It was measured by the method.

1) Screening of Inhibitory Effect of Cyclooxygenase-1 and Cyclooxygenase-2 by LPS-Induced Intraperitoneal Macrophage Method

First, in the case of cyclooxygenase-1, it carried out as follows.

Intraperitoneal solution in which macrophage was suspended in mouse abdominal cavity was extracted and centrifuged at 4 ° C. and 1000 rpm for 2 minutes. The supernatant was removed, suspended in 20 ml of incomplete RPMI (including penicillin streptomycin) and centrifuged again under the same conditions. After two more washes, the cell pellet was suspended in 10 ml of incomplete (no serum) RPMI 1640 to obtain a cell suspension, after which the cell count was measured by a hemocytometer to 1 × 10 ⁶ cells / ml. The final suspension was made. 100 μl of this suspension was added to each well of a 96-well plate and left for 5 hours in a 5% CO ₂ , 37 ° C. incubator to attach macrophages. The attached macrophages were washed twice with PBS and then treated with the appropriate concentration of the test sample, and 3% FBS-RPMI 1640 medium was added so that the total volume was 200 µl. 5% CO ₂ , incubated for about 12-16 hours in a 37 ℃ incubator. Arachidonic acid was added to a final concentration of 10 μM and further incubated at 37 ° C. for 10 minutes, and the reaction supernatant (˜180 μl) was recovered to terminate the reaction. The ELISA method provided by Cayman Chemicals was used to quantify PGE2 for the sample, and from this result, the inhibition rate (% inhibition) of cyclooxygenase-1 of each compound was calculated.

In the case of cyclooxygenase-2,

Intraperitoneal solution in which macrophage was suspended in mouse abdominal cavity was extracted and centrifuged at 4 ° C. and 1000 rpm for 2 minutes. The supernatant was removed, suspended in 20 ml of incomplete RPMI (including PC / SM) and centrifuged again under the same conditions. After two more washes, the cell pellet was suspended in 10 ml of incomplete (no serum) RPMI 1640 to obtain a cell suspension, after which the cell count was measured by a hemocytometer to 1 × 10 ⁶ cells / ml. The final suspension was made. This suspension was treated with aspirin to a final concentration of 500 μM, and then 100 μl was added to each well of a 96-well plate and left for 5 hours in a 5% CO ₂ , 37 ° C. incubator to attach macrophages. Attached macrophages were washed twice with PBS and then treated with appropriate concentration of test samples, and 3% FBS-RPMI 1640 medium containing 10 µg / ml LPS was added to each well. 5% CO ₂ , incubated for about 12-16 hours in a 37 ℃ incubator. Arachidonic acid was added to a final concentration of 10 μM and further incubated at 37 ° C. for 10 minutes, and the reaction supernatant (˜180 μl) was recovered to terminate the reaction. The ELISA method provided by Cayman Chemicals was used to quantify PGE2 for the sample, and from this result, the inhibition rate (% inhibition) of cyclooxygenase-2 of each compound was calculated.

2) Screening of Inhibitory Effect of Cyclooxygenase-1 Using U-937

U-937 cells (Human lymphoma cell; Korea Cell Line Bank; Accession No. 21593) were collected by centrifugation and cultured in 90% RPMI1640 medium and 10% fetal calf serum (FBS) inactivated by heat, followed by 1xHBSS (Hank's balanced salt solutin). 1 ml per well was dispensed into 12-well plates by diluting to 1 × 10 ⁶ cells / ml. 5 μl of a sample solution and DMSO vehicle, which were dissolved in DMSO and diluted to an appropriate concentration, were added and mixed. Incubated for 15 min in a CO ₂ incubator at 37 ℃. A stock solution prepared by dissolving arachidonic acid used as a substrate in ethanol at a concentration of 10 mM was diluted 10-fold with 1 × HBSS to prepare a 1 mM solution. 10 μl of 1 mM arachidonic acid solution was added to each well treated with the test sample, followed by incubation for 30 minutes in a 37 ° C. CO ₂ incubator. Cell solutions of each well were collected in a centrifuge tube and centrifuged at 4 ° C. and 10,000 rpm for 5 minutes. The concentration of PGE2 in the supernatant separated from the cells collected by centrifugation was quantified using a monoclonal kit (Cayman Chemicals), and the concentration of the experimental substance treated group was compared with that of the vehicle group to generate PGE2 of each substance. By determining the inhibition rate, the inhibitory effect of the substance on the cyclooxygenase-1 enzyme was determined.

3) Screening of Inhibitory Effect of Cyclooxygenase-2 Using Raw 264.7 Cell Line

Raw 264.7 cells (Korea Cell Line Bank; Accession No. 40071) were seeded 2 × 10 ⁶ per well of a 12 well plate, treated with aspirin at 250 μM, and incubated for 2 hours at 37 ° C. [Medium: Dulbecco's modified Eagle's medium 90% and heat inactivated fetal bovine serum (FBS) 10%]. After changing to fresh medium, each sample was treated and incubated for 30 minutes. Here, the cells were treated with interferon γ (100 units / ml) and lipopolysaccharide (LPS, 100ng / ml) per well, and then cultured for 18 hours. Thereafter, the medium was transferred to another tube and PGE2 was quantified using an EIA kit (Cayman Chemicals).

2. Experimental Results

The inhibitory effect of cyclooxygenase by LPS-induced mouse intraperitoneal macrophage method is shown in Table 1 below, and the inhibitory effect of cyclooxygenase using U-937 and Raw 264.7 cell lines is shown in Table 2 below.

Cyclooxygenase (COX) Inhibitory Effect (% inhibition) Example COX-1 (%) COX-2 (%) 10 μM 3 μM 1 μM 300 nM 100nM 30 nM SC-58635 (control material) 87.37 84.23 84.72 70.12 57.70 37.40 One 80.35 72.00 71.98 74.63 66.08 45.56 2 74.84 73.65 63.20 67.51 52.07 49.08 3 83.64 84.84 83.61 81.94 77.64 77.29 4 81.67 79.98 79.15 58.84 60.85 54.05 5 85.13 78.23 80.36 49.09 61.91 53.76

Cyclooxygenase (COX) Inhibitory Effect (% inhibition) Example COX-1 (%) COX-2 (%) 10 μM 3 μM 1 μM 300 nM 100nM 30 nM SC-58635 (control material) 85.3 76.0 46.0 97.8 75.1 7.0 6 67.6 50.9 30.8 87.4 36.8 0 7 0 7.8 2.7 7.7 7.7 0 8 47.1 32.0 14.7 4.7 0 0

Looking at the results of in vitro experiments on the inhibition of cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) are as follows.

The compounds of Examples 1 to 5 according to the invention showed an equivalent or superior cyclooxygenase inhibitory effect overall compared to the control. In particular, 6- (5-phenyl-3-trifluoromethyl-pyrazol-1-yl) -pyridine-3-sulfonamide of Example 1 and 6- [5- (4-methoxy- of Example 3 In the case of phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide, the inhibitory effect of cyclooxygenase-2 was superior to that of the control, and the inhibitory effect of cyclooxygenase-1 was Was reduced and proved to be excellent selectivity.

The novel compound according to the present invention is an alternative drug with improved side effects of the existing nonsteroidal anti-inflammatory drugs, and is used in patients with peptic ulcer, gastritis, partial enteritis, ulcerative colitis, diverticulitis, gastrointestinal bleeding, hypoprothrombinemia, etc. It is useful and is expected to be useful as a therapeutic agent for inflammatory diseases such as osteoarthritis and rheumatoid arthritis.

Claims (4)

A compound of Formula 1 and pharmaceutically acceptable salts thereof [Formula 1] In the above formula, R ¹ represents hydrogen, methyl or trifluoromethyl, R ² represents halogen, C ₁ -C ₃ -alkyl, halogenomethyl, C ₁ -C ₃ -alkoxy, hydroxy, hydroxycarbonyl or nitro, n represents the number of 0-5. The compound of claim 1, wherein R ¹ is hydrogen, methyl or trifluoromethyl, R ² is methyl or methoxy, and n is a number from 0 to 3. 3. The method of claim 1, 6- (5-phenyl-3-trifluoromethyl-pyrazol-1-yl) -pyridine-3-sulfonamide; 6- (5-p-tolyl-3-trifluoromethyl-pyrazol-1-yl) -pyridine-3-sulfonamide; 6- [5- (4-methoxy-phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide; 6- [5- (4-Fluoro-phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide; 6- [5- (4-Chloro-phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide; 6- [5- (2,4-Dichloro-phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide; 6- [5- (2,4-Dichloro-5-fluoro-phenyl) -3-trifluoromethyl-pyrazol-1-yl] -pyridine-3-sulfonamide; 6- (3-methyl-5-phenyl-pyrazol-1-yl) -pyridine-3-sulfonamide. A process for preparing a compound of formula 1 as defined in claim 1 characterized in that the hydrazine derivative of formula 2 is reacted with a compound of formula 3 in the presence of an acid catalyst in a solvent: [Formula 2] [Formula 3] In the above formula R ¹ , R ² and n are as defined in claim 1.