KR101118768B1 - 5-3-Aryl-1-pyridyl-1H-pyrazol-4-ylmethylene-thiazolidine-2,4-dione derivatives useful as antitumor agent - Google Patents

Abstract

The present invention provides a 5- (3-aryl-1-pyridyl- 1H -pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivative or a pharmaceutically acceptable salt thereof and a method for preparing the same It relates to an anticancer agent composition containing as an active ingredient.

5- (3-aryl-1-pyridyl-1H-pyrazol-4-ylmethylene) -thiazolidine-2,4-dione, cell division inhibitor, CDC25B, dephosphatase, anticancer agent

Description

5- (3-aryl-1-pyridyl-1H-pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivative useful as an anticancer agent {5- (3-Aryl-1-pyridyl-1H- pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivatives useful as antitumor agent}

1 shows in vivo anti-tumor effect of cdc25B inhibitor candidates using a human tumor model.

The present invention relates to a 5- (3-aryl-1-pyridyl-1H-pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivative or a pharmaceutically acceptable salt thereof and a method for preparing the same It relates to an anticancer agent composition containing as an active ingredient.

CDC25B is a very important dephosphatase that determines the progression to the G2 / M cycle in cell division. When CDC25B activity is inhibited, cell division is inhibited and eventually leads to cell death, resulting in anticancer activity. CDC25B can be used for gastric cancer (Jpn. J. Cancer Res. 1997, 88, 9947), colon cancer (Exp. Cell Res. 1998, 240, 236; Lab. Invest. 2001, 81, 465), lung cancer (Cancer Res. 1998, 58, 4082), brain cancer and laryngeal cancer (Cancer Res. 1997, 57, 2366), breast cancer (Cancer statistics in Japan 1997 Foundation for Promotion of Cancer Research, Tokyo, Jpn.) And overexpressed in most tumors, CDC25B Mammary gland hyperplasia was observed in overexpressed genotyping mice (Oncogene 1999, 18, 4561), and breast cancer was very easily induced by treatment with casinogen (9,10-dimethyl-1,2-benzanthracene). Has been reported. Therefore, CDC25B is an important target in the development of anticancer drugs, and there is a high possibility of developing a new anticancer agent that inhibits CDC25B and has low cytotoxicity to normal cells.

Regarding the 5- (substituted-1H-pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivative compounds, EP-A-379979 A1 discloses a pyrazole structure linked to the 5-position of thiazole. It is described that a 5- (substituted-1H-pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivative having the structure of pyrazolopyridine can be used as an adenosine antagonist. In addition, Japanese Patent No. 55029804 discloses 5- (substituted-1H-pyrazol-4-ylmethylene) -thiazolidine-2, in which the pyrazole structure linked to the 5-position of thiazole has a structure of pyrazolone, Compounds of 4-dione derivatives are described as useful dyes that inhibit clouding and luminescence in silver halide sensitizers.

The inventors have found that 5- (3-aryl-1-pyridyl-1H-pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivatives have selective inhibitory activity against CDC25B and other similar dephosphorylations. The present invention has been completed by discovering a novel compound having selectivity and anticancer effect on enzymes.

One object of the present invention is to provide 5- (3-aryl-1-pyridyl-1H-pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivative or a pharmaceutically acceptable salt thereof It is.

Another object of the present invention is a method for preparing 5- (3-aryl-1-pyridyl-1H-pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivative or a pharmaceutically acceptable salt thereof. To provide.

Another object of the present invention includes a 5- (3-aryl-1-pyridyl-1H-pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivative or a pharmaceutically acceptable salt thereof It is to provide an anticancer agent composition.

In one embodiment, the present invention provides a 5- (3-aryl-1-pyridyl-1H-pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivative of formula (1) Pertaining to acceptable salts.

In this formula,

X is hydrogen, C ₁ -C ₄ alkyl group, halogen atom, C ₁ -C ₄ alkoxy group, C ₁ -C ₄ haloalkoxy group, C ₃ -C ₄ alkenyloxy group, C ₂ -C ₄ acyl group, C ₃ -C ₁₀ cycloalkylalkoxy, C ₃ -C ₁₀ cycloalkyloxy, C ₆ -C ₁₀ arylalkoxy, cyan or nitro group substituents;

n is an integer from 1 to 5;

The pyridyl group at the 1-position of pyrazole represents 2-pyridyl, 3-pyridyl or 4-pyridyl;

R represents hydrogen, a C ₁ -C ₄ alkyl group, a C ₂ -C ₄ alkenyl group, a C ₃ -C ₄ alkynyl group, a C ₁ -C ₄ haloalkyl group, a C ₂ -C ₄ alkoxyalkyl group or a benzyl group.

As a preferred embodiment of the present invention, Formula 1 is

X is C ₁ -C ₄ alkyl group, halogen atom, C ₁ -C ₄ alkoxy group, C ₁ -C ₄ haloalkoxy group, C ₃ -C ₄ alkenyloxy group, C ₃ -C ₁₀ cycloalkylalkoxy, C _3- C ₁₀ cycloalkyloxy, C ₆ -C ₁₀ arylalkoxy or nitro group substituents;

n is an integer from 1 to 3;

The pyridyl group at the 1-position of pyrazole represents 2-pyridyl, 3-pyridyl or 4-pyridyl;

R represents hydrogen, a C ₁ -C ₄ alkyl group, a C ₁ -C ₄ haloalkyl group, a C ₂ -C ₄ alkenyl group or a C ₃ -C ₄ alkynyl group.

As a preferred embodiment of the present invention, Formula 1 is

X is a halogen atom, isopropyl group, tertiary butyl group, bromine, iodine, methoxy group, ethoxy group, propyloxy group, isopropoxy group, allyloxy group, nitro group, cyclopropylmethoxy group or cyclopentyloxy group, Fluoromethoxy group, benzyloxy group, propenyloxy group substituents;

n is an integer from 1 to 3;

The pyridyl group at the 1-position of pyrazole represents 2-pyridyl, 3-pyridyl or 4-pyridyl;

R represents hydrogen, methyl group, ethyl group, propyl group, propenyl group, propynyl group or C ₁ -C ₄ fluoroalkyl group.

As a preferred embodiment of the present invention, Formula 1 is

X is 4-position is methoxy group, ethoxy group, propyloxy group, fluoromethoxy group, propenyloxy group or allyloxy group, 3-position and 5-position are isopropyl group, tertiary butyl group, bromine, Iodine, benzyloxy, fluoromethoxy, cyclopropylmethoxy group or cyclopentyloxy group substituents;

n is an integer from 1 to 3;

The pyridyl group at the 1-position of pyrazole represents 2-pyridyl or 4-pyridyl;

R represents hydrogen, methyl, ethyl or propyl.

In the compound of Formula 1, the position of X may be in the 1 to 5-position of the phenyl group, preferably in the 3, 4 or 5-position.

In a most preferred embodiment of the present invention, the compound of formula 1 is 5- [3- (3,5-dicyclopropylmethoxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene ] -Thiazolidine-2,4-dione, 5- [3- (3,4-difluoromethoxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -thia Zolidine-2,4-dione, 5- [3- (3,5-dibenzyloxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -thiazolidine-2 , 4-dione, 5- [3- (3-bromo-4-ethoxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -thiazolidine-2,4 -Dione, 5- [3- (3,5-dibromo-4-propoxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -thiazolidine-2, 4-dione, 5- [3- (3,5-diisopropyl-4-propoxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -thiazolidine-2 , 4-dione or their hydrochloride salts.

As used herein, the term "alkyl" means a straight or branched, saturated hydrocarbon radical of 1 to 4 carbon atoms, unless otherwise indicated.

As used herein, the term "halogen" or "halo" refers to a halogen atom and includes fluorine, chlorine, bromine, iodine, fluorine and the like.

As used herein, the term "alkoxy" means a 0-alkyl group (alkyl is defined above) unless otherwise indicated.

The term "alkenyl" as used herein, unless otherwise indicated, means an unsaturated hydrocarbon radical having 2 to 4 carbon atoms having a double bond.

As used herein, the term "alkynyl" means an unsaturated hydrocarbon radical having 3 to 4 carbon atoms, unless otherwise indicated.

The term "acyl" as used herein, unless otherwise indicated, refers to "aroyl" groups derived from aliphatic carboxylic acids, such as acetyl, propionyl, and the like.

The term "cycloalkylalkoxy" used in the present invention means a radical having alkoxy bonded to a saturated hydrocarbon ring having 3 to 10 carbon atoms, unless otherwise indicated.

The term "cycloalkyloxy" used in the present invention means a radical having oxygen bonded to a saturated hydrocarbon ring having 3 to 10 carbon atoms, unless otherwise indicated.

As used herein, unless otherwise indicated, the term "alkenyloxy" means a radical wherein an unsaturated hydrocarbon having a double bond, having 3 to 4 carbon atoms, is bonded to oxygen.

As used herein, the term "haloalkyl" means a radical in which the hydrogen of an alkyl group (as defined above) is replaced with a halogen atom, unless otherwise indicated.

As used herein, the term "haloalkoxy" means a radical wherein the hydrogen of alkoxy (as defined above) is replaced with a halogen atom, unless otherwise indicated.

Compounds of formula (I) of the present invention include pharmaceutically acceptable salts thereof. Such salts can be prepared using conventional techniques in the art. Such pharmaceutically acceptable salts are, for example, salts with organic or inorganic acids such as hydrochloride, bromate, sulfate, phosphate, acetate, citrate, fumarate, lactate, maleate, succinate, tartarate, sodium Salts, salts with alkali metals such as potassium salts, or all pharmaceutically acceptable salts such as ammonium salts.

In another aspect of the present invention, the present invention comprises the steps of preparing a compound of formula 4 by reacting a compound of formula 2 and a compound of formula 3; Preparing a compound of Formula 5 by reacting the compound of Formula 4 with Vilsmeier-Haack; Reacting a compound of formula 5 with thiazolo-2,4-dione to prepare a compound of formula 1 wherein R is H, or reacting a compound of formula 5 with a compound of formula 6 to It relates to a process for preparing a compound of formula (1) or a pharmaceutically acceptable salt thereof, including the step of preparing.

In this formula,

X, n and R are as defined above,

Z represents a halogen atom such as chlorine, bromine or iodine, a toluenesulfonyloxy group, a methanesulfonyloxy group or the like as a leaving group.

Scheme of the production method of the present invention can be represented by the following scheme 1.

a: acetic acid or benzoic acid

b: Alcohols such as methanol and ethanol or water or organic solvent

c: anhydrous dimethylformamide (DMF) / phosphorusoxy chloride (POCl ₃ ), thionyl chloride or oxalyl chloride, etc.

d: sodium hydroxide aqueous solution

e: salts of weak acids such as acetic acid and benzoic acid, weak bases such as pyridine, amines and aniline

f: organic solvents such as benzene and toluene

g: Inorganic bases, such as potassium carbonate, sodium carbonate, sodium hydroxide, calcium hydroxide, cesium carbonate, or organic base, such as triethylamine and pyridine

h: dimethylformamide (DMF), tetrahydrofuran (THF), acetone, water or other organic solvent

First, the hydrazone derivative of Formula 4 may be prepared by reacting the substituted acetophenone of Formula 2 with the substituted hydrazine of Formula 3. In this reaction, the solvent may be alcohols such as methanol or ethanol, water or other organic solvents, preferably alcohols such as methanol or ethanol. The catalyst may use a small amount of weak acid such as acetic acid or benzoic acid.

Subsequently, the hydrazone derivative of Formula 4 was used as an solvent using anhydrous dimethylformamide (DMF) and phosphorusoxy chloride (POCl ₃ ) or thionyl chloride or oxalyl chloride, preferably phosphorusoxy chloride (POCl ₃ ) was used. The 3-aryl-pyridyl-pyrazole-4-carboxaldehyde derivative of the formula (5) can be prepared as a catalyst.

Subsequently, the compound of formula 1, wherein R is H, may be prepared by reacting pyrazolecarboxaldehyde of formula 5 with thiazolidine-2,4-dione. In this reaction, the solvent may be benzene, toluene or other organic solvent, preferably benzene or toluene. The catalyst may be mixed with acetic acid and piperidine, or salts of weak acids such as acetic acid and benzoic acid, or weak bases such as pyridine, amines and aniline, and preferably mixed with acetic acid and piperidine. .

To prepare a compound of formula 1 wherein R has a substituent other than H, it may be reacted with a compound of formula 6 in the presence of a base. In this reaction, the solvent may be dimethylformamide (DMF), tetrahydrofuran (THF), acetone, water or other organic solvent, preferably dimethylformamide (DMF), tetrahydrofuran (THF). . The base may be an inorganic base such as potassium carbonate, sodium carbonate, sodium hydroxide, potassium hydroxide or cesium carbonate, or an organic base such as triethylamine or pyridine, and preferably potassium carbonate or sodium carbonate.

In another embodiment of the invention, the invention provides 5- (3-aryl-1-pyridyl-1H-pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivative of formula (1) or a medicament thereof It relates to an anticancer agent comprising a scientifically acceptable salt as an active ingredient.

The 5- (3-aryl-1-pyridyl-1H-pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivative of Formula 1 determines the progression of the cell division cycle to G2 / M. It has inhibitory activity against CDC25B dephosphorase, which plays a decisive role, and inhibits cell division. It also shows selectivity to other dephosphorase enzymes in the human body and at the same time in human experiments using nude mice. It has been shown to have an effect of destroying cancer cells.

Accordingly, the anticancer agent or pharmaceutical composition comprising the compound of formula 1 according to the present invention as an active ingredient may be prepared by adding a conventional non-toxic pharmaceutically acceptable carrier, adjuvant or excipient to the compound of formula 1, For example, it can be formulated into preparations for oral administration such as tablets, capsules, troches, solutions, and suspensions.

In addition, the dosage of the compound of Formula 1 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. To 400 mg / day, and may be divided once to several times a day at regular time intervals according to the judgment of a doctor or a pharmacist.

Hereinafter, the present invention will be described in more detail with reference to the following examples. However, the following examples are merely to aid the understanding of the present invention, and the scope of the present invention is not limited to the following examples.

Example 1: 5- [3- (3,5-Dicyclopentyloxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -thiazolidine-2,4-dione Preparation of (Compound No. 9)

(1) (3 ', 5'-dicyclopentyloxy) acetophenone (Formula 2)

(3 ', 5'-dihydroxy) acetophenone (2.50 g, 16.43 mmol) was dissolved in 50 mL of anhydrous acetone and cyclopentane bromide (4.40 mL, 41.08 mmol), potassium carbonate (5.00 g, 36.13 mmol), catalyst Potassium iodide (27.28 mg, 0.16 mmol) was added as a reflux for 5 hours, and then slowly cooled to room temperature. Ethyl acetate (50 mL) was added to the mixture, followed by washing with water (20 mL × 2 times) and brine (20 mL × 2 times). The organic layer was separated, dried over anhydrous magnesium sulfate, filtered and concentrated under reduced pressure. The residue was purified by silica gel chromatography (n-hexane / ethyl acetate = 10: 1) to give 4.22 g (89%) of the desired product.

¹ H NMR (300 MHz, CDCl ₃ ) δ 1.50-1.68 (m, 4H), 1.72-1.95 (m, 12H), 2.55 (s, 3H), 4.74-4.83 (m, 2H), 6.59 (s, 1H ), 7.03 (s, 2 H).

(2) Pyridin-2-ylhydrazone of (3 ', 5'-dicyclopentyloxy) acetophenone (Formula 4)

(3 ', 5'-dicyclopentyloxy) acetophenone (3.0 g, 10.40 mmol) was dissolved in 30 mL of anhydrous ethanol and 2-hydrazinopyridine (1.13 g, 10.40 mmol), glacial acetic acid (28.82 μl, 0.52 as catalyst) mmol) and stirred at room temperature for 10 hours. After adding 20 mL of water, the resulting solid was filtered and washed with water (10 mL × 3 times). The filtered solid was dried to prepare 3.54 g (90%) of the desired product.

(3) 3- (3,5-dicyclopentyloxy) -1-pyridin-2-yl-1H-pyrazole-4-carboxyaldehyde (Formula 5)

Phosphorus oxychloride (0.45 mL, 7.02 mol) was added to 2 mL of anhydrous dimethylformamide and stirred at 0 ° C. for 1 hour. Pyridin-2-ylhydrazone (3.54 g, 9.35 mmol) of (3 ', 5'-dicyclopentyloxy) acetophenone dissolved in 5 mL of anhydrous dimethylformamide was slowly added dropwise, and then the temperature was 70-80 ° C. Raised to and stirred for 6 hours. The temperature was cooled to 0 ° C. with ice water, slowly added dropwise with 30% aqueous sodium hydroxide solution to pH 7-8, and the resulting solid was filtered and washed with water (10 mL × 3 times). The filtered solid was dried to give 2.73 g (70%) of the desired product.

¹ H NMR (200 MHz, CDCl 3) δ 1.21-1.95 (m, 16H), 4.72-4.88 (m, 2H), 6.50 (s, 1H), 6.97 (s, 2H), 7.20-7.35 (m, 1H) , 7.88 (t, 1H, J = 5.7 Hz), 8.12 (d, 1H, J = 8.1 Hz), 8.48 (d, 1H, J = 1.4 Hz), 9.17 (s, 1H), 10.10 (s, 1H) .

(4) 5- [3- (3,5-dicyclopentyloxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -thiazolidine-2,4-dione ( Compound number 9)

3- (3,5-Dicyclopentyloxy-phenyl) -1-pyridin-2-yl-1H-pyrazole-4-carboxyaldehyde (1.00 g, 2.40 mmol), thiazolidine-2,4-dione (282.00 mg, 2.40 mmol) was added to 50 mL of anhydrous toluene and glacial acetic acid (6.78 μL, 0.12 as a catalyst). mmol), piperidine (14.29 μl, 0.14 mmol), and refluxed for 12 hours while removing water using Dean Starktrap. The temperature was lowered to room temperature and stirred for 6 hours. The resulting solid was filtered and washed with diethyl ethyr (10 mL × 3 times). The filtered solid was dried to prepare 0.97 g (78%) of the desired product.

¹ H NMR (200 MHz, CDCl ₃ ) δ 1.64-2.01 (m, 16H), 4.82-4.93 (m, 2H), 6.55 (s, 1H), 6.70 (s, 2H), 6.99-7.54 (m, 1H ), 7.63 (s, 1H), 8.07 (t, 2H, J = 7.1 Hz), 8.57 (d, 2H, J = 3.9 Hz), 8.77 (s, 1H), 12.56 (br, 1H, NH).

Example 2: 5- [3- (3,5-Dicyclopentyloxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -3-methyl-thiazolidine-2 Preparation of, 4-dione (Compound No. 10)

5- [3- (3,5-Dicyclopentyloxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -thiazolidine-2,4-dione (Compound No. 9 , 0.10 g, 0.21 mmol) was dissolved in 1.0 mL of anhydrous diformamide, sodium carbonate (24.38 mg, 0.23 mmol) was added under nitrogen gas, and after 10 minutes, methane iodide (21.65 µl, 0.29 mmol) was added thereto and stirred at room temperature for 2 hours. I was. After the reaction was completed, water was added to the resulting solid was filtered and washed with water (10 mL × 3 times). The filtered solid was dried to give 80.7 mg (87%) of the desired product.

¹ H NMR (200 MHz, CDCl ₃ ) δ 1.60-2.00 (m, 16H), 4.75-4.82 (m, 2H), 6.52 (s, 1H), 6.72 (s, 2H), 7.22-7.32 (m, 1H ), 7.85 (d, 1H, J = 7.8 Hz), 7.97 (s, 1H), 8.10 (d, 1H, J = 8.3 Hz), 8.48 (d, 1H, J = 5.5 Hz), 8.88 (s, 1H ).

Example 3: 5- [3- (3,5-Dicyclopentyloxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -3-ethyl-thiazolidine-2 Preparation of 4,4-dione (Compound No. 11)

5- [3- (3,5-Difluoro-phenyl) -1-phenyl-1H-pyrazol-4-ylmethylene] -thiazolidine-2,4-dione (Compound No. 9, 0.10 g, 0.21 mmol) was dissolved in 1.0 mL of anhydrous diformamide, sodium carbonate (26.5 mg, 0.25 mmol) was added under nitrogen gas, and after 10 minutes, ethyl bromide (23.90 μl, 0.32 mmol) was added thereto, followed by stirring at room temperature for 4 hours. After the reaction was completed, water was added, and the resulting precipitate was filtered, washed with water (10 mL × 3 times), and dried (85.3 mg, 80%) to prepare a target substance.

¹ H NMR (200 MHz, CDCl ₃ ) δ 0.95 (t, 3H, J = 7.5 Hz), 1.20-1.96 (m, 18H), 3.72 (t, 3H, J = 7.3 Hz), 4.74-4.84 (m, 2H), 6.51 (s, 1H), 6.73 (s, 2H), 7.21-7.31 (m, 1H), 7.85 (d, 1H, J = 7.7 Hz), 7.95 (s, 1H), 8.10 (d, 1H , J = 7.3 Hz), 8.48 (d, 1H, J = 4.8 Hz), 8.88 (s, 1H).

Example 4: 5- [3- (3-Bromo-4-ethoxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -thiazolidine-2,4- Preparation of Dione (Compound No. 24)

(1) Pyridin-2-ylhydrazone of (3'-bromo-4'-ethoxy) acetophenone (Formula 4)

(3'-bromo-4'-ethoxy) acetophenone (4.00 g, 16.45 mmol) was dissolved in 100 mL of absolute ethanol and 2-hydrazinopyridine (1.80 g, 16.45 mmol), glacial acetic acid (9.16 μl, 0.16 mmol) was added thereto, followed by stirring at room temperature for 5 hours. After adding 20 mL of water, the resulting solid was filtered and washed with water (10 mL × 3 times). The filtered solid was dried to give 3.68 g (92%) of the desired product.

(2) 3- (3-bromo-4-ethoxy-phenyl) -1-pyridin-2-yl-1H-pyrazole-4-carboxyaldehyde (Formula 5)

Phosphorus oxychloride (2.68 mL, 23.4 mmol) was added to 2 mL of anhydrous dimethylformamide, and the mixture was stirred at 0 ° C. under nitrogen gas for 1 hour. Pyridin-2-ylhydrazone (3.68 g, 15.14 mmol) of (3'-bromo-4'-ethoxy) acetophenone dissolved in 5 mL of anhydrous dimethylformamide was slowly added dropwise, and then the temperature was 70-80. It was raised to ℃ and stirred for 6 hours. The temperature was cooled to 0 ° C. with ice water, slowly added dropwise with 30% aqueous sodium hydroxide solution to pH 7-8, and the resulting solid was filtered and washed with water (10 mL × 3 times). The filtered solid was dried to give 2.89 g (51.3%) of the desired product.

¹ H NMR (300 MHz, CDCl ₃ ) δ 1.53 (t, 3H, J = 6.9 Hz), 4.20 (q, 2H, J = 6.9 Hz), 6.93-8.40 (m, 7H), 9.17 (s, 1H) , 10.07 (s, 1 H).

(3) 5- [3- (3-bromo-4-ethoxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -thiazolidine-2,4-dione (Compound No. 24)

3- (3-bromo-4-ethoxy-phenyl) -1-pyridin-2-yl-1H-pyrazole-4-carboxyaldehyde (1.00 g, 2.69 mmol), thiazolidine-2,4- Dione (316.00 mg, 2.69 mmol) was added to 50 mL of anhydrous toluene and glacial acetic acid (7.60 μL, 0.13 as a catalyst). mmol), piperidine (16.08 μl, 16.44 mmol), and refluxed for 3 hours while removing water using Dean Starktrap. The temperature was lowered to room temperature and stirred for 6 hours. The resulting solid was filtered off and washed with diethyl ether (10 mL × 3 times). The filtered solid was dried to prepare 1.04 g (82%) of the desired product.

¹ H NMR (200 MHz, CDCl ₃ ) δ 1.41 (t, 3H, J = 6.9 Hz), 4.21 (q, 2H, J = 6.9 Hz), 7.20-8.60 (m, 8H), 8.79 (s, 1H) , 12.50 (br, 1 H, NH).

Example 5: 5- [3- (3-Bromo-4-ethoxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -3-methyl-thiazolidine- Preparation of 2,4-dione (Compound No. 26)

5- [3- (3-Bromo-4-ethoxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -thiazolidine-2,4-dione (Compound No. 24, 0.10 g, 0.21 mmol) was dissolved in 1.00 mL of anhydrous dimethylformamide, sodium carbonate (33.92 mg, 0.32 mmol) was added under nitrogen gas, and after 5 minutes, methane iodide (19.78 µL, 0.32 mmol) was added thereto at room temperature for 2 hours. Stirred. After the reaction was completed, 10 mL of water was added to the resulting solid, which was filtered, washed with water (10 mL × 3 times), and dried to prepare 68 mg (67%) of the title compound.

¹ H NMR (300 MHz, CDCl ₃ ) δ 1.52 (t, 3H, J = 7.0 Hz), 3.25 (s, 3H), 4.20 (q, 2H, J = 7.0 Hz), 6.99-8.88 (m, 9H) .

Example 6: 5- [3- (3-Bromo-4-ethoxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -3-ethyl-thiazolidine- Preparation of 2,4-dione (Compound No. 27)

5- [3- (3-Bromo-4-ethoxy-phenyl) -1-pyridin-2-yl-1H-pyrazol-4-ylmethylene] -thiazolidine-2,4-dione (Compound No. 24, 0.10 mg, 0.21 mmol) was dissolved in 1.00 mL of anhydrous dimethylformamide, sodium carbonate (33.92 mg, 0.32 mmol) was added under nitrogen gas, and after 5 minutes, ethane bromide (23.72 μL, 0.32 mmol) was added thereto at room temperature for 2 hours. Stirred. After the reaction was completed, 10 mL of water was added to the resulting solid, which was filtered, washed with water (10 mL × 3 times), and dried to prepare 57 mg (54%) of the title compound.

¹ H NMR (300 MHz, CDCl ₃ ) δ 1.26-1.57 (m, 6H), 3.82 (q, 2H, J = 7.2 Hz), 4.19 (q, 2H, J = 6.9 Hz), 6.99-8.87 (m, 9H).

Compounds listed in Table 1 below were synthesized in a similar manner to Examples 1-6.

Example 7: Formulation

The compound of formula 1 according to the present invention can be formulated in various forms according to the purpose. The following examples show several formulation methods containing the compound of formula 1 according to the present invention as an active ingredient, but the present invention is not limited thereto.

Formulation 1: tablet (direct pressure)

After sifting 5.0 mg of active ingredient, 14.1 mg of lactose, 0.8 mg of Crospovidone USNF, and 0.1 mg of magnesium stearate were mixed and pressed to form a tablet.

Formulation 2: Tablet (Wet Granulation)

After sifting 5.0 mg of the active ingredient, 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.

Formulation 3: Powders and Capsules

5.0 mg of the active ingredient was sieved, followed by mixing 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.

Example 8: CDC25B Enzyme Assay

The portion corresponding to the catalytic domain (aa 378-566) of CDC25B was expressed as a GST fusion protein in E. coli and used as an enzyme source. Enzyme assays were performed with a final reaction volume of 200 μl per well on a 96-well plate. To the reaction, 170 μl of buffer (30 mM Tris buffer, pH 8.5, 75 mM NaCl, 0.67 mM EDTA, 1 mM DTT), 20 μl of CDC25B enzyme (0.2 μg) and 10 μl of test compound dissolved in DMSO were added. Was adjusted to 1 μg / mL CDC25B enzyme, 20 μM FDP, and cultured for 1 hour at room temperature. At this time, FDP and DTT were added to the buffer just before the assay, so that the assay was always performed in the fresh state. After 1 hour of incubation, the fluorescence generated by the enzyme reaction was measured at 485 nm (excitation) and 538 nm (emission).

The test compound was first treated with a final concentration of 10 μM, followed by primary screening, and then, an IC ₅₀ value was determined by selecting a compound showing excellent inhibitory ability. Compounds screened by primary screening were generally treated with final concentrations of 1, 2, 5, and 10 μM to obtain% inhibition values. In the case of), the concentration of the compound was increased to 20 μM, and the percentage inhibition rate was determined to obtain an IC ₅₀ value.

IC ₅₀ values for CDC25B are listed in Table 2 below for compounds having excellent activity among the compounds synthesized in Table 1.

Compound number IC ₅₀ ([mu] M) 5 0.55 9 4.33 17 2.39 21 6.05 22 0.27 23 0.43 24 2.42 25 1.37 29 7.40 43 3.66 50 0.52 51 0.43 52 0.43

Example 9: Cell cytotoxicity assay

(1) Cancer Cell Culture

The cells used for measuring anticancer activity are A549 and MCF-7. All of these cancer cells were human-derived tumor cell lines that were obtained from the National Cancer Institute (NCI) in the US and passaged at the Korea Institute of Chemical Research. These cells were all cultured in a 37 ° C. constant temperature and 5% CO ₂ , incubator using RPMI 1640 culture supplemented with 5% fetal bovine serum as culture. Cells were passaged once every 3 to 4 days, and 0.25% trypsin and 3 mM trans-1,2-diaminocyclohexane-N, N, in a solution of phosphate buffered saline (PBS) to separate the cells from the adherent surface. A solution in which N, N-tetraacetic acid (CDTA) was dissolved was used.

(2) Activity measurement

In 1989, the US National Cancer Institute used the sulforhodamine B assay method developed to measure the in vitro anticancer activity of drugs. Cells were passaged from the implant using trypsin-CDTA solution for use in the passaged cells for experiments and seeded at 96-well microplates (96 well microplate, manufactured by Falcon) at a cell count of about 2 × 10 ^{3 per} well. . The aliquoted cells were incubated for 24 hours in a CO ₂ incubator and attached to the bottom, followed by removal of the culture solution with an aspirator, and a log dose of 6 concentrations of Doxorubicin as a reference compound with the compounds of compounds Nos. 17 and 25. Compound solutions diluted in the culture medium (log dose) were put into 3 wells of 100 mL each into the wells containing cells, and further cultured for 72 hours. Dimethyl sulfoxide (DMSO) was used as needed to dissolve the compound. This diluted compound solution was filtered through a 0.22 mL filter prior to addition to the cells to maintain the sterility of the experiment. After incubating the cells with the drug for 72 hours, the culture solution of each well was removed, and 100 mL of 10% trichloroacetic acid (TCA) was added thereto and left at 4 ° C. for 1 hour to fix the cells on the bottom of the plate. After the fixation of the cells, the plate was washed 5 to 6 times with water to completely remove the remaining TCA solution, and dried at room temperature to ensure no remaining water. Completely dried plates were stained for 30 minutes by adding dye solution dissolved in 0.4% SRB to 100 mL of 1% acetic acid solution per well, and washed again 5 to 6 times with 1% acetic acid solution to remove SRB that did not bind to cells. Removed. The stained cell plates are dried again at room temperature and then eluted with dye by shaking for 10 minutes with a titer plate shaker by adding 100 mL of 10 mM unbuffered trisma base solution per well. After absorbance was measured at 520 nm using a microplate reader (microplate reader). To calculate the effect of the drug on cancer cells, the cell number (Tz) at the time of drug addition, the cell number (C) at 72 hours incubation with the drug-free medium, and each concentration of drug 72 The cell number (T) at the time of incubation was measured and calculated by the following Equations 1 and 2 below.

Using the data regression of the LOTUS program from the calculated values, the degree to which the drug inhibits the growth of cancer cells is shown in Table 3 below as an IC ₅₀ value.

IC ₅₀ ([mu] M) Compound Number Cancer Cell Line MCF-7 A549 25 13.4 14.0 17 2.7 6.3

According to Table 3, 5- (3-aryl-1-pyridyl-1H-pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivative of formula 1 according to the present invention is A-549 Cytotoxicity against (non-small cell lung cancer), MCF-7 (breast cancer).

Example 10: In Vivo Antitumor Effect of CDC25B Inhibitor Candidates Using a Human Tumor Model

In order to verify the in vivo antitumor effect of Compound 1 of the present invention, C33A human uterine cancer cell line was injected subcutaneously into the abdominal wall of male nude mice 6 to 8 weeks old. When tumors grew about 50-80 mm ³ , compound 17 was intraperitoneally administered with 0.2% Tween 80 used as a solvent as a negative control, and tumor growth was observed. As a positive control, cisplatin (4 mg / kg), which is widely used in clinical practice as an anticancer agent, was used. The amount of the solvent was 0.25 ml per horse per dose and repeated administration for 7 days once a day. Tumor volume was calculated by the following equation 3 after measuring the long axis and short axis of the tumor using a caliper.

Volume of tumor = (short mm) ² x long axis mm x 0.523.

The result is shown in FIG. According to this, compound 17 exhibited anti-cancer effect of cisplatin level, and there was no reaction of anorexia and anorexia due to cisplatin toxicity. Therefore, the 5- (3-aryl-1-pyridyl-1H-pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivative of the formula (1) can be used as a low toxicity anticancer agent with less side effects.

Contains 5- (3-aryl-1-pyridyl- 1H -pyrazol-4-ylmethylene) -thiazolidine-2,4-dione derivative of the formula (1) according to the present invention and a pharmaceutically acceptable salt thereof Compound shows excellent inhibitory activity against CDC25B enzyme, an important dephosphorylation enzyme that determines the progression of cell division to G2 / M cycle, and can be used as a low-toxic anticancer agent with less side effects such as decreased motility due to toxicity and anorexia reaction. Can be.

Claims (4)

A compound of formula 1 or a pharmaceutically acceptable salt thereof. Formula 1

In this formula, X is hydrogen, C ₁ -C ₄ alkyl group, halogen atom, C ₁ -C ₄ alkoxy group, C ₁ -C ₄ haloalkoxy group, C ₃ -C ₄ alkenyloxy group, C ₂ -C ₄ acyl group, C ₃ -C ₁₀ cycloalkylalkoxy, C ₃ -C ₁₀ cycloalkyloxy, C ₆ -C ₁₀ arylalkoxy, cyan or nitro group substituents; n is an integer from 1 to 5; The pyridyl group at the 1-position of pyrazole represents 2-pyridyl, 3-pyridyl or 4-pyridyl; R represents hydrogen, a C ₁ -C ₄ alkyl group, a C ₂ -C ₄ alkenyl group, a C ₃ -C ₄ alkynyl group, a C ₁ -C ₄ haloalkyl group, a C ₂ -C ₄ alkoxyalkyl group or a benzyl group. The method of claim 1, X is C ₁ -C ₄ alkyl group, halogen atom, C ₁ -C ₄ alkoxy group, C ₁ -C ₄ haloalkoxy group, C ₃ -C ₄ alkenyloxy group, C ₃ -C ₁₀ cycloalkylalkoxy, C _3- C ₁₀ cycloalkyloxy, C ₆ -C ₁₀ arylalkoxy or nitro group substituents; n is an integer from 1 to 3; The pyridyl group at the 1-position of pyrazole represents 2-pyridyl, 3-pyridyl or 4-pyridyl; A compound of formula (1) or a pharmaceutically acceptable salt thereof, wherein R represents hydrogen, a C ₁ -C ₄ alkyl group, a C ₁ -C ₄ haloalkyl group, a C ₂ -C ₄ alkenyl group or a C ₃ -C ₄ alkynyl group. Reacting a compound of Formula 2 with a compound of Formula 3 to prepare a compound of Formula 4; Preparing a compound of Formula 5 by reacting the compound of Formula 4 with Vilsmeier-Haack; Reacting a compound of formula 5 with thiazolo-2,4-dione to prepare a compound of formula 1 wherein R is H, or reacting a compound of formula 5 with a compound of formula 6 to A process for preparing a compound of Formula 1 or a pharmaceutically acceptable salt thereof, comprising the step of preparing. Formula 1

Formula 2

Formula 3

Formula 4

Formula 5

6

In this formula, X, n and R are as defined in claim 1, Z represents a leaving group. A pharmaceutical composition for anticancer comprising a pharmaceutically effective amount of a compound of formula 1 or a pharmaceutically acceptable salt thereof. Formula 1

In this formula, X, n and R are as defined in claim 1.

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