KR20060086529A - Novel purine compound having lipoxygenase inhibitory activity, the preparation method thereof and a composition containing the same for preventing and treating asthma - Google Patents

Abstract

The present invention provides a novel Purine Derivative (I) compound having inhibitory activity against Lipoxygenase, a preparation method thereof, and a composition for treating and preventing asthma, including the same.

Since the compound according to the present invention shows a strong inhibitory activity against lipoxygenase, the composition containing the same can be used as a medicament for the treatment of asthma.

Lipoxygenase, inhibitory activity, asthma, purine, derivatives

Description

 Novel purine compound having lipoxygenase inhibitory activity, the preparation method about and a composition containing the same for preventing a novel purine derivative compound having an inhibitory activity against lipoxygenase and treating asthma}

The present invention relates to a novel purine derivative compound having inhibitory activity against lipoxygenase, a preparation method thereof, and a composition for treating and preventing asthma comprising the same.

Asthma (ASTHMA) is a respiratory disease characterized by reversible airway obstruction, airway inflammation, and airway hyperresponsiveness, with about 12 million people currently having asthma in the United States. Statistics from 1982 to 1992 show that asthma prevalence has increased from 34.7 per 1,000 to 49.4 per 1000 and mortality has increased by approximately 40%, from 13.4 per million to 18.8 per million. In 1995, domestic research showed that about 10% of children and adolescents have experienced asthma symptoms within the past year and have similar results in adults. In adults, about 10% of the population experiences asthma symptoms (Asthma Research Society, The Korean Society for Internal Medicine, 60 , pp196-205, 2001).

Asthma airway obstruction is caused by a combination of factors such as spasm of airway smooth muscle, swelling of the airway mucosa, increased mucus secretion, airway infiltration of inflammatory cells (especially eosinophils and lymphocytes), and damage and loss of airway epithelial cells. Inflammatory mediators are involved in bronchospasm, mucus secretion, and microvascular leakage. Microvascular leakage leads to submucosal edema, increased airway resistance and is also involved in airway hyperresponsiveness. Inflammatory mediators include histamine and arachidonic acid metabolites (leukotriene and thromboxane). Cysteine leukotrienes LTC ₄ (Leukotriene C ₄ ) and LTD ₄ (Leukotriene D ₄ ) are the most potent bronchial contractors. On the other hand, T cell activation in allergic reactions plays the most important role in airway inflammation, and cytokines produced in CD4T _H 2 cells promote the proliferation, differentiation and activation of inflammatory cells, migration of inflammatory cells into the airways and cell lifespan. (Wills-Kapp, M. et al., Interleukin-13: Central mediator of allergic asthma Science, 282 , pp 2258-2261, 1998).

If you classify asthma according to the cause, it can be classified into exogenous, endogenous, mixed, aspirin-induced, exercise-induced, occupational asthma. Exogenous asthma is symptomatic when exposed to causative antigens. A skin test or bronchial challenge test for causative antigens is positive and the onset age is young. House dust mites are the most common cause antigens, and pollen, animal epithelium, and fungi act as causative antigens. Endogenous asthma is caused or worsened by upper respiratory tract infection, exercise, emotional instability, cold climate and humidity changes, and is common in adult asthma. Mixed asthma is caused by a combination of exogenous and endogenous factors, which are common in pediatric asthma. Aspirin-induced asthma is a specific constitutional response to aspirin and refers to the three major symptoms of bronchial asthma, nasal polyps, and aspirin intolerance. Exercise-induced asthma is associated with the development of hyperventilation due to exercise, and thus loss of heat or water in the airways. Occupational asthma is caused by substances that are inhaled in the workplace, initially without symptoms, then asthmatic symptoms appear months or years later, tend to relieve on weekends or vacations, and worsen when returning to work.

Asthma attacks occur suddenly and wheezing, coughing, shortness of breath, etc., most often cough, chest tightness or pressure, and wheezing can be heard. During acute seizures, various symptoms of respiratory distress occur, prompting rapid breathing and rapid pulse. During chest examination, a loud wheezing can be heard during inspiration and most exhalations and an extension of the exhalation can be observed. More severe asthma attacks can lead to rapid and shallow breathing due to fatigue and severe breathing difficulties, and cyanosis as the symptoms become more severe. If the patient shows conscious turbidity and helplessness, it means progression to respiratory failure due to CO ₂ coma, in which case severe mucus plugging hardly hears wheezing and exacerbates gas exchange by exhaustion. The most obvious signs of a severe seizure include difficulty breathing, unable to talk, and the use of cyanosis, acute veins (> 20-30 mmHg), and apnea even at rest. Severity can be assessed most accurately by measuring arterial blood gases (Higgins, G., Inpharma., 1077 , pp 9-10, 1997).

Asthma medications developed so far include preventive medications that are taken at all times and drugs that quickly relieve sudden symptoms of asthma. The former includes inhaled corticosteroids, metyprednisolone, such as beclomethasone, budnoside, flunisolide, fluticasone, and triamcinolone. Oral corticosteroids such as prednisolone, prednisone, or salmeterol, beta-agonist, nedocromil, sodium chromoglycate, etc. master cell stabilizers such as sodium cromoglycate, methylxanthines such as theophylline, and leukotriene antagonists such as zafirlukast and zileuton Drugs that quickly relieve symptoms include salbutamol, bitolterol, pirbuterol, and terbutaline. Anticholinergic agents such as beta-antagonist, ipratropium bromide and oral corticosteroids such as methylprednisolone, prednisolone, and prednisone Corticosteroids (NAEPP (National Asthma Education and Prevention Program, guideline, 1997)).                         

Among the above drugs, leukotrienes (LT) antagonists are drugs that inhibit the synthesis or action of LT, and are mainly cysteinyl-LT antagonists and 5-lipoxygenase inhibitors (5- lipoxygenase inhibitors, 5-lipoxygenase activating protein inhibitors, and LTB ₄ antagonists. Lipoxygenase is a type of dioxygenase, classified according to the double bond sites (# 5, 11, 15) where oxygen attacks. Leukotriene is a degradation product of 5-lipoxygenase and plays an important role in inflammation. 5-lipoxygenase converts Arachidonic acid to 5-HPETE and back to Leukotriene A ₄ . From this again LTB ₄ , LTC ₄ is produced and the glutamic acid and glycine residues of LTC ₄ are cut off by dipeptidase and LTD ₄ and LTE ₄ are produced. HETE and LTB ₄ are involved in the action of neutrophils and eosinophils to relay chemoxis. It stimulates adenylyl cyclase, induces PMN and activates lysosomal hydrolytic enzymes. LTC ₄ and LTD ₄ contract the smooth muscle, contract the lungs and contract the muscles of the bronchus and intestine. It also increases the permeability of capillaries, causing edema. LTB ₄ is immunosuppressive. It inhibits CD ₄ ⁺ cells and proliferates CD ₈ ⁺ cells. Also Neutrophil-endothelial promoting the adhesion of cell allergy (Allergy), infection (Inflammation), secretion, cell migration of insulin (Insulin) (Cell movement), cell growth (Cell growth), the flow of Ca ₂ ⁺ (fluxes) To get involved. LTC ₄ and LTD ₄ are more potent allergens than histamine (Fabien, J. et al., Expert Opinion. Ther. Patents , 13 (1) , pp1-13, 2003).

The inventors of the LT antagonist, purine (purine) derivatives modeled after the structure of the representative drug marketed as a 5-lipoxygenase inhibitor, Ziruton and E-6080 (Eisai) under development in Japan By introducing various substituents as a basis, studies have been conducted to synthesize and develop derivatives showing potent activity against 5-lipoxygenase.

The present inventors confirmed that the novel purine derivative compounds newly prepared by the present inventors exhibit potent inhibitory activity against 5-lipoxygenase enzyme, and thus the compounds of the present invention have high development potential as a therapeutic agent for asthma and complete the present invention. Was done.

It is an object of the present invention to provide a purine derivative compound exhibiting potent inhibitory activity against 5-lipoxygenase enzyme, a method for preparing the same, and a composition for the prevention and treatment of asthma comprising the same.

In order to achieve the above object, the present invention provides a purine derivative compound having the structure of formula (I) or a pharmacologically acceptable salt thereof, which is useful for treating and preventing asthma:

(Ⅰ)

(Ⅰ)

Where

R ₁ is one or more substituents selected from a hydrogen atom, a halogen atom, a hydroxyl group, a sulfanyl group or a lower alkyl group,

R ₂ is a C ₁ -C ₆ lower acyl or benzoyl group unsubstituted or substituted with one or more R ′,

R 'is a substituent which is a halogen atom or a C ₁ -C ₆ lower alkyl group.

Among the compound groups belonging to the general formula (I), preferably, R ₁ is a hydrogen atom, a hydroxyl group or a sulfanyl group; R ₂ is a compound group which is an acetyl group, isobutylyl group, propionyl group or benzoyl group unsubstituted or substituted with one or more halogen atoms or C ₁ -C ₃ lower alkyl groups.

General formula Particularly preferred groups of compounds in (I) include the following compounds and pharmaceutically acceptable salts thereof:

N- (9H-purin-8-yl) -benzamide,

N- (6-hydroxy-9H-purin-8-yl) -benzamide,

N- (2,6-dihydroxy-9H-purin-8-yl) -benzamide,

N- (6-hydroxy-2-mercapto-7H-purin-8-yl) -benzamide,

N- (7H-purin-8-yl) -isobutylamide,

N- (6-hydroxy-9H-purin-8-yl) -isobutylamide,

N- (9H-purin-8-yl) -3,5-bis-trifluoromethyl-benzamide,

N- (6-hydroxy-9H-purin-8-yl) -3,5-bis-trifluoromethyl-benzamide,

N- (9H-purin-8-yl) -acetamide,

N- (9H-purin-8-yl) -propionamide,

N- (6-hydroxy-9H-purin-8-yl) -propionamide,

N- (2,6-dihydroxy-9H-purin-8-yl) -propionamide,

2,2-dimethyl-N- (9H-purin-8-yl) -propionamide,

N- (6-hydroxy-9H-purin-8-yl) -2,2-dimethyl-propionamide,                     

N- (2,6-dihydroxy-9H-purin-8-yl) -2,2-dimethyl-propionamide,

N- (6-hydroxy-2-mercapto-7H-purin-8-yl) -2,2-dimethyl-propionamide.

The compounds of the present invention represented by the general formula (I) may be prepared with pharmaceutically acceptable salts and solvates according to conventional methods in the art.

As salts are acid addition salts formed with pharmaceutically acceptable free acids. Acid addition salts are prepared by conventional methods, for example by dissolving a compound in an excess of aqueous acid solution and precipitating the salt using a water miscible organic solvent, such as methanol, ethanol, acetone or acetonitrile. Equimolar amounts of the compound and acid or alcohol (eg, glycol monomethylether) in water can be heated and the mixture can then be evaporated to dryness or the precipitated salts can be suction filtered.

In this case, organic acids and inorganic acids may be used as the free acid, hydrochloric acid, phosphoric acid, sulfuric acid, nitric acid, tartaric acid, etc. may be used as the inorganic acid, and methanesulfonic acid, p -toluenesulfonic acid, acetic acid, trifluoroacetic acid, Citric acid, maleic acid, succinic acid, oxalic acid, benzoic acid, tartaric acid, fumaric acid, manderic acid, propionic acid, citric acid, lactic acid, glycolic acid, gluconic acid ( gluconic acid), galacturonic acid, glutamic acid, glutaric acid, glucuronic acid, glucuronic acid, aspartic acid, ascorbic acid, carbonic acid, vanic acid, hydroiodic acid, and the like.

Bases can also be used to make pharmaceutically acceptable metal salts. Alkali metal or alkaline earth metal salts are obtained by, for example, dissolving a compound in an excess alkali metal hydroxide or alkaline earth metal hydroxide solution, filtering the insoluble compound salt, and then evaporating and drying the filtrate. At this time, as the metal salt, it is particularly suitable to prepare sodium, potassium or calcium salts, and the corresponding silver salt is obtained by reacting an alkali metal or alkaline earth metal salt with a suitable silver salt (for example, silver nitrate).

Pharmaceutically acceptable salts of the above general formula (I) include salts of acidic or basic groups which may be present in compounds of general formula (I), unless otherwise indicated. For example, pharmaceutically acceptable salts include sodium, calcium and potassium salts of the hydroxy group, and other pharmaceutically acceptable salts of the amino group include hydrobromide, sulfate, hydrogen sulphate, phosphate, hydrogen phosphate, dihydrogen Phosphate, acetate, succinate, citrate, tartrate, lactate, mandelate, methanesulfonate (mesylate) and p -toluenesulfonate (tosylate) salts, and methods or processes for preparing salts known in the art It can be prepared through.

Another object of the present invention is to provide a method for preparing the compound of Formula (I), which may be chemically synthesized by the method shown in the following schemes, but is not limited thereto.

The following schemes represent the preparation steps of the representative compounds of the present invention. The various compounds of the present invention may be prepared by small changes such as changing the reagents, solvents, and reaction sequences used in the synthesis of Schemes 1 to 3. have. Some compounds of the present invention were synthesized according to procedures not included in the scope of Schemes 1 to 3, and detailed synthesis procedures for these compounds are described in their respective examples.

First, a first step of preparing pyrimidinediamine (b) by reacting the O -nitropyrimidineamine (a) substituted with R ₁ in the presence of a reducing agent such as hydrogen (H ₂ ) under a Pd / C catalyst;

The Oh minche (c) carbon disulfide (CS ₂₎ in the optionally substituted with R _2, methane iodide (MeI), and the reaction was gradually sodium hydride (NaH) mixture in the solid state under an argon gas at O ℃ to room temperature, N- (bis A second step of preparing methylsulfanylmethylene) amide derivative compound (d);

The pyrimidine diamine obtained in Scheme 1 and the N- (bis-methylsulfanylmethylene) amide derivative obtained in Scheme 2 (d) are refluxed under an inert organic solvent to provide various substituents of R ₁ to R ₂ . Purine derivative compounds of the present invention of general formula (I) having

Accordingly, the present invention provides a method for preparing the purine derivative compound of the general formula (I).

      Compounds of general formula (I) obtained by the above production method are useful as a therapeutic agent capable of treating or preventing asthma diseases by confirming that they exhibit potent asthma inhibitory activity through experiments on inhibitory activity against lipoxygenase enzyme.

Accordingly, the present invention provides a pharmaceutical composition for the treatment and prevention of asthma comprising the purine derivative compound of formula (I) as an active ingredient.

     Compositions comprising a compound of the present invention may further comprise a suitable carrier, excipient or diluent according to conventional methods.                     

Carriers, excipients and diluents that may be included in the compositions of the present invention include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, Cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil.

The compositions comprising the compounds of the present invention are each formulated in the form of oral dosage forms, external preparations, suppositories, or sterile injectable solutions, such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, etc., in accordance with conventional methods. Can be used.

Specifically, when formulated, it may be prepared using diluents or excipients such as commonly used fillers, extenders, binders, wetting agents, disintegrating agents, surfactants, and the like. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and such solid preparations may contain at least one excipient such as starch, calcium carbonate, sucrose, or the like. ), Lactose, gelatin and the like can be mixed. In addition to simple excipients, lubricants such as magnesium stearate, talc can also be used. Liquid preparations for oral use include suspensions, solvents, emulsions, and syrups, and include various excipients such as wetting agents, sweeteners, fragrances, and preservatives, in addition to commonly used simple diluents such as water and liquid paraffin. Can be. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations and suppositories. As the non-aqueous solvent and suspending agent, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, injectable esters such as ethanolate and the like can be used. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin butter, glycerogelatin and the like can be used.

Preferred dosages of the compounds of the present invention depend on the condition and weight of the patient, the extent of the disease, the form of the drug, the route of administration and the duration, but may be appropriately selected by those skilled in the art. However, for the desired effect, the compound of the present invention may be administered in an amount of 0.0001 to 100 mg / kg, preferably in an amount of 0.001 to 100 mg / kg once to several times daily. The compound of the present invention in the composition should be present in an amount of 0.0001 to 10% by weight, preferably 0.001 to 1% by weight, based on the total weight of the total composition.

In addition, the pharmaceutical dosage forms of the compounds of the present invention may be used in the form of their pharmaceutically acceptable salts, and may be used alone or in combination with other pharmaceutically active compounds as well as in a suitable collection.

The pharmaceutical composition of the present invention can be administered to mammals such as mice, mice, livestock, humans, etc. 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 will be described in more detail based on the following examples and experimental examples, but the present invention is not limited thereto.

Reference Example 1. Experiment Preparation and Devices

1-1. Analyzer

The instrument used to confirm the structure of the product obtained in this experiment is as follows. Nuclear magnetic resonance spectra (H NMR, C NMR) were used at 300 MHz or 400 MHz, and the solvent used was CDC, DMSO-d ₆ . Coupling constants ( J ) are expressed in Hz. Mass spectra were ESI and expressed in m / z.

1-2. TLC and tube chromatography

Thin layer chromatography (TLC) was used as silica gel (Merck F254) manufactured by E. Merck, and silica (Merck EM9385, 230-400 mesh) was used for column chromatography. In addition, using a UV lamp (= 254 nm) or soaked in anisealdehyde (Anisaldehyde), potassium permanganate (KMnO ₄ ) coloring reagent to identify the material separated on the TLC, the plate was confirmed by heating.

1-3. Used reagents

The reagents used in this experiment were purchased from Sigma-Aldrich, Lancaster, and Fluka, and the solvent used in the reaction was Sigma-Aldrich, Merck. A first grade reagent from Merck, Junsei Chemical Co. was used without purification. The THF used in the solvent was used when Na was added to Na metal and benzophenone in the argon stream and heated to reflux to become blue. In addition, dichloromethane (CH ₂ Cl ₂ ) was used by heating under reflux with CaH ₂ in the argon stream. Ethyl acetate and hexane were purified by heating under reflux in an argon stream.

Reference Example 2 Preparation of Intermediates 1a to 1c

2-1. Preparation of 4,5-diaminopyridine (1a)

Dissolve 5-nitro-pyrimidin-4-yl amine (500 mg, 3.57 mmol) in MeOH (26 ml) in a 250 ml round flask and add Pd / C ( 0.03 mg, 0.26 mmol) was added. A hydrogen balloon was attached and air was completely drained 3-4 times using a guest pump, replaced with hydrogen, and stirred at room temperature for 5 hours. After the reaction, the mixture was filtered to completely remove Pd / C, the filtrate was concentrated under reduced pressure, and the residue was separated by column chromatography (EtOAc: n-Hexane = 1: 1) to obtain 4,5-diaminopyridine compound (1a) 95.2. Obtained in% yield (400 mg).

2-2. Preparation of 4-hydroxy-pyrimidine-5,6-diamine (1b)

Experiment with 4-hydroxy-6-amino-5-nitro-pyrimidine (4-hydroxy-6-amino-5-nitro-pyrimidine 500 mg, 3.20 mmol) for 12 hours in the same manner as in Reference Example 2-1. To prepare 4-hydroxy-pyrimidine-5,6-diamine having the following physical properties (1b) The compound was obtained in a yield of 79.1% (319 mg).

2-3. Preparation of 2,4-dihydroxy-pyrimidine-5,6-diamine (1c)

Reference Example 2- above with 2,4-dihydroxy-6-amino-5-nitro-pyrimidine (2,4-dihydroxy-6-Amino-5-nitro-pyrimidine; 500 mg, 2.64 mmol) for 12 hours. The experiment was carried out in the same manner as in 1 to give a 2,4-dihydroxy-pyrimidine-5,6-diamine (1c) compound having a yield of 79.1% (319 mg).

Reference Example 3. Preparation of Intermediates (2a to 2f)

3-1. N- (bis-methylsulfanyl-methylene) -acetamide (2a)

Acetamide (500 mg, 8.46 mmol) in a round flask substituted with argon gas was dissolved in 33.9 ml of DMF, followed by carbon disulfide (2.0 ml, 33.8 mmol) and iodine at 0 ° C. Domethane (Iodomethane; 1.7 ml, 27.1 mmol) and sodium hydroxide (Sodium hydride; 445 mg, 18.6 mmol) were added sequentially and stirred at room temperature for 5 hours. The reaction was terminated with distilled water and extracted with EtOAc. The organic layer was dried over anhydrous MgSO, filtered, concentrated under reduced pressure, separated by column chromatography EtOAc: n-Hexane = 1: 7), and separated with N- (bis-methylsulfanyl-methylene) -acetamide (Physical Properties). 2a) The compound was obtained in a yield of 25.2% (163 mg).

¹ H NMR (300 MHz, CDCl ₃ ): 2.47 (s, 6H), 2.23 (s, 3H)

3-2. N- (bis-methylsulfanyl-methylene) -benzamide (2b)

Benzamide (2.00 g, 16.51 mmol) in a round flask substituted with argon gas was dissolved in DMF (66 ml), followed by carbon disulfide (4 ml, 66.04 mmol) and iododomethane (3.3 ml, 52.83 mmol) at 0 ° C. ), Sodium hydroxide 60% (1.32 g, 33.02 mmol) was added sequentially and stirred at room temperature for 5 hours. The reaction was terminated with distilled water and extracted with EtOAc. The organic layer was dried over anhydrous MgSO ₄ , filtered and concentrated under reduced pressure, and separated by column chromatography (EtOAc: n-Hexane = 1: 7) to obtain N- (bis-methylsulfanyl-methylene) -benz having the following physical properties. An amide (2b) compound was obtained in a yield of 40.8% (1.52 g).

¹ H NMR (300 MHz, CDCl ₃ ): 8.13 (d, J = 7.5 Hz, 2H), 7.65 (t, J = 6.6 Hz, 1H), 7.55 (t, J = 6.6 Hz, 2H), 2.47 (s , 6H), 1.19 (s, 3H), 1.17 (s, 3H)

3-3. N- (bis-methylsulfanyl-methylene) -isobutylamide (2c)

Isobutylamide (1.00 g, 11.47 mmol) in a round flask substituted with argon gas was dissolved in DMF (46 ml), and carbon disulfide (2.8 ml, 45.91 mmol), iodomethane (2.2 ml, 36.73) at 0 ° C. mmol) and sodium hydroxide 60% (0.92 g, 22.96 mmol) were added sequentially and stirred at room temperature for 5 hours. The reaction was terminated with distilled water and extracted with EtOAc. The organic layer was dried over anhydrous MgSO 4, filtered and concentrated under reduced pressure, separated by column chromatography (EtOAc: n-Hexane = 1: 7), and separated by N- (bis-methylsulfanyl-methylene) -isobutyl having the following physical properties. An amide (2c) compound was obtained in 64.2% yield (1.41 g).

¹ H NMR (300 MHz, CDCl ₃ ): 2.71-2.62 (m, 1H), 2.47 (s, 6H), 1.19 (s, 3H), 1.17 (s, 3H)

3-4, N- (bis-methylsulfanyl-methylene) -3,5-bis-trifluoromethyl-benzamide (2d)

3,5-bis (trifluoromethyl) benzamide (3,5-Bis (trifluoromethyl)-benzamide; 500 mg, 1.94 mmol), carbon disulfide (0.47 ml, 7.78 mmol), iodomethane (0.39 ml, 6.22 mmol), sodium hydroxide (156 mg, 3.89 mmol), and DMF (7.8 ml) in the same manner as in Reference Example 3-1, where N- (bis-methylsulfanyl-methylene) -3 having the following physical properties: 5-Bis-trifluoromethyl-benzamide (2d) compound was obtained in 32.0% yield (225 mg).

¹ H NMR (400 MHz, CDCl ₃ ): 8.64 (s, 2H), 8.03 (s, 1H), 1.26 (s, 6H)

3-5. N- (bis-methylsulfanyl-methylene) -propionamide (2e)

Propionamide (1.0 g, 13.68 mmol), carbon disulfide (3.3 ml, 54.72 mmol), iodomethane (2.7 ml, 43.78 mmol), sodium hydroxide (1.09 g, 27.36 mmol), 54.7 ml DMF In the same manner as in 3-1, an N- (bis-methylsulfanyl-methylene) -propionamide (2e) compound having the following physical properties was obtained in a yield of 35.1% (850 mg).

¹ H NMR (400 MHz, CDCl ₃ ): 2.73 (q, J = 7.2 Hz, 2H), 2.33 (t, J = 3.3 Hz, 3H), 1.16 (s, 6H)

3-6. N- (bis-methylsulfanyl-methylene) -2,2-dimethyl-propionamide (2f)

Trimethylacetamide (1.0 g, 9.89 mmol), carbon disulfide (2.4 ml, 39.54 mmol), iodomethane (2.0 ml, 31.64 mmol), sodium hydroxide (0.79 g, 19.77 mmol), see above in 40 ml DMF In the same manner as in Example 3-1, an N- (bis-methylsulfanyl-methylene) -2,2-dimethyl-propionamide (2f) compound having the following physical properties was obtained in a yield of 71.9% (1.46 g). .

¹ H NMR (400 MHz, CDCl ₃ ): 2.48 (s, 6H), 1.21 (s, 9H)

Example 1 N- (9H-Purin-8-yl) -benzamide

N- (bis-methylsulfanyl-methylene) -benzamide (159 mg, 0.71 mmol) in Reference Example 3-2 was dissolved in DMF (3 ml), and then 4,5-diamino-pyrimidine (4,5 -Di-amino pyrimidine (78 mg, 0.71 mmol) was added and refluxed for 8 hours. After the reaction, the mixture was cooled to room temperature and the solvent was removed by distillation under reduced pressure. Thereafter, the mixture was filtered through MeOH to obtain an N- (9H-purin-8-yl) -benzamide compound having the following physical properties in 71.1% yield (120 mg).

¹ H NMR (300 MHz, DMSO-d ₆ ): (br, NH, NH), 8.81 (s, 1H), 8.81 (s, 1H), 8.13 (d, J = 7.5 Hz, 2H), 7.68 (t , J = 7.4 Hz, 1H), 7.57 (t, J = 7.4 Hz, 2H)

Example 2. N- (6-hydroxy-9H-purin-8-yl) -benzamide

N- (bis-methylsulfanyl-methylene) -benzamide (200 mg, 0.89 mmol), 4,5-diamino-6-hydroxypyrimidine-hemisulphate (4,5- of Reference Example 3-2 Diamino-6-hydroxypyrimidine hemisulfate; 155 mg, 0.89 mmol) N- (6-hydroxy-9H-purin-8-yl having the following physical properties by the same experiment as in Example 1 with DMF (4 ml) ) -Benzamide compound was obtained in a yield of 45.6% (103 mg).

¹ H NMR (400 MHz, DMSO-d ₆ ): 11.83 (br, NH), 11.47 (br, NH), 7.148 (d, J = 8.1 Hz, 1H)

Example 3. N- (2,6-dihydroxy-9H-purin-8-yl) -benzamide

N- (bis-methylsulfanyl-methylene) -benzamide (126 mg, 0.56 mmol), 5,6-diamino-2,4-dihydroxypyrimidine sulfate (5,6) in Reference Example 3-2 -Diamino-2,4-dihydroxypyrimidi nesulfate; 79 mg, 0.56 mmol), DMF (2.2 ml) was tested in the same manner as in Example 1 N- (2,6-dihydroxy- having the following physical properties 9H-Purin-8-yl) -benzamide compound was obtained in a yield of 30.9% (467 mg).

¹ H NMR (400 MHz, DMSO-d ₆ ): 10.72 (br, NH), 10.47 (br, NH), 8.04 (d, J = 7.2 Hz, 1H), 7.64 (t, J = 7.6 Hz, 1H) , 7.55 (t, J = 7.6 Hz, 1H), 7.46-7.39 (m, 2H); m / e 271 (M)

Example 4.N- (6-hydroxy-2-mercapto-7H-purin-8-yl) -benzamide

N- (bis-methylsulfanyl-methylene) -benzamide (123 mg, 0.55 mmol) and 4,5-diamino-6-hydroxy-2-mercaptopyrimidine hemisulfate hydrate of Reference Example 3-2 (4,5-Diamino-6-hydroxy-2-mercaptopyrimidinehemisulfatehydrate; 124 mg, 0.55 mmol) in the same manner as in Example 1 with DMF (2.2 ml) An N- (6-hydroxy-2-mercapto-7H-purin-8-yl) -benzamide compound having the following physical properties was obtained in a yield of 36.0% (566 mg).

¹ H NMR (400 MHz, DMSO-d ₆ ): 11.89 (br, NH, NH), 8.05 (d, J = 8.0 Hz, 2H), 7.65 (t, J = 7.4 Hz, 1H), 7.54 (t, J = 7.4 Hz, 2H); m / e 287 (M)

Example 5. N- (7H-purin-8-yl) -isobutylamide

N- (bis-methylsulfanyl-methylene) -isobutylamide (100 mg, 0.52 mmol), 4,5-diaminopyrimidine (4,5-Diaminopyrimidine; 58 mg, 0.52 mmol) of Reference Example 3-3 Experiment with DMF (2.1 ml) in the same manner as in Example 1 An N- (7H-purin-8-yl) -isobutylamide compound having the following physical properties was obtained in a yield of 9.1% (9.8 mg).

¹ H NMR (400 MHz, DMSO-d ₆ ): 12.8-12.4 (br, NH), 12.4-12 (br, NH), 8.76 (s, 1H), 8.72 (s, 1H), 3-2.6 (m , 1H), 1.15 (d, J = 6.8 Hz, 6H); ^{ESI m / e 206.1 (M +} ), 204.0 (M -)

Example 6. N- (6-hydroxy-9H-purin-8-yl) -isobutylamide

N- (bis-methylsulfanyl-methylene) -isobutylamide (150 mg, 0.78 mmol), 4,5-diamino-6-hydroxypyrimidine-hemisulphate (137 mg, in Reference Example 3-3 0.78 mmol) and DMF (3.1 ml) in the same manner as in Example 1 An N- (6-hydroxy-9H-purin-8-yl) -isobutylamide compound having the following physical properties was obtained in a yield of 47.8% (82.9 mg).

¹ H NMR (400 MHz, DMSO-d ₆ ): 12.8 (br, NH), 12.4 (br, NH), 8.76 (s, 1H), 8.72 (s, 1H), 3-2.6 (m, 1H), 1.15 (d, J = 6.8 Hz, 6H); 13 C NMR (DMSO-d ₆ ): 177.33, 162.30, 158.75, 152.02, 151.23, 138.55, 124.25; + C ESI (m / e) 206.1, -C ESI (m / z) 204.0.

Example 7.N- (9H-Purin-8-yl) -3,5-bis-trifluoromethyl-benzamide

N- (bis-methylsulfanyl-methylene) -3,5-bis-trifluoromethyl-benzamide (70 mg, 0.19 mmol) and 4,5-diaminopyrimidine (21 mg) of Reference Examples 3-4 , 0.19 mmol), in the same manner as in Example 1 with DMF (0.8 ml) An N- (9H-purin-8-yl) -3,5-bis-trifluoromethyl-benzamide compound having the following physical properties was obtained in a yield of 70.3% (51.1 mg).

¹ H NMR (400 MHz, DMSO-d ₆ ): 13.1-12.8 (br, NH), 8.83 (s, 1H), 8.81 (s, 1H), 8.74 (s, 1H), 8.43 (s, 1H); m / e 375 (M)

Example 8. N- (6-hydroxy-9H-purin-8-yl) -3,5-bis-trifluoromethyl-benzamide

N- (bis-methylsulfanyl-methylene) -3,5-bis-trifluoromethyl-benzamide (75 mg, 0.21 mmol), 4,5-diamino-6-hydroxy of Reference Example 3-4 Experiment with pyrimidine-hemisulphate (36 mg, 0.21 mmol) and DMF (1 ml) in the same manner as in Example 1 N- (6-hydroxy-9H-purin-8-yl having the following physical properties ) -3,5-bis-trifluoromethyl-benzamide compound was obtained in a yield of 28.9% (23.5 mg).

¹ H NMR (400 MHz, DMSO-d ₆ ): 12.30 (br, NH, NH), 8.71 (s, 2H), 8.43 (s, 1H), 7.98 (s, 1H)

Example 9 N- (9H-Purin-8-yl) -acetamide

N- (bis-methylsulfanyl-methylene) -acetamide (110 mg, 0.67 mmol), 4,5-diaminopyrimidine (74 mg, 0.67 mmol) and DMF (2 ml) in Reference Example 3-1. Experiment as in Example 1 N- (9H-purin-8-yl) -acetamide compound having the following physical properties was obtained in a yield of 72.7 mg (74.8 mg).

¹ H NMR (400 MHz, DMSO-d ₆ ): 12.48 (br, NH), 12.06 (br, NH), 8.77 (s, 1H), 8.71 (s, 1H), 2.21 (s, 3H)

Example 10. N- (9H-Purin-8-yl) -propionamide

Compound 61 (85 mg, 0.48 mmol), 4,5-diaminopyrimidine (53 mg, 0.48 mmol) and DMF (2 ml) were tested in the same manner as in Example 1, and the recrystallization was performed with EtOAc. An N- (9H-purin-8-yl) -propionamide compound having the following physical properties was obtained in a yield of 47.8% (43.8 mg).

¹ H NMR (400 MHz, DMSO-d ₆ ): 12.51 (br, NH), 12.02 (br, NH), 8.76 (s, 1H), 8.71 (s, 1H), 2.5 (q, 2H), 1.12 ( t, 3H); m / e 191 (M)

Example 11 N- (6-hydroxy-9H-purin-8-yl) -propionamide

N- (bis-methylsulfanyl-methylene) -propionamide (105 mg, 0.59 mmol) and 4,5-diamino-6-hydroxypyrimidine-hemisulphate (104 mg, 0.59) in Reference Examples 3-5 mmol) and DMF (2.4 ml) in the same manner as in Example 1, and recrystallization was performed with EtOAc. An N- (6-hydroxy-9H-purin-8-yl) -propionamide compound having the following physical properties was obtained in a yield of 68.7% (84.6 mg).

¹ H NMR (400 MHz, DMSO-d ₆ ): 8.04 (s, 1H), 2.32 (m, 2H), 1.09 (t, J = 7.6 Hz, 3H)

Example 12.N- (2,6-Dihydroxy-9H-purin-8-yl) -propionamide

N- (bis-methylsulfanyl-methylene) -propionamide (93 mg, 0.52 mmol), 5,6-diamino-2,4-dihydroxypyrimidine sulfate (75 mg, in Reference Example 3-5) 0.52 mmol) and DMF (2.1 ml) in the same manner as in Example 1, and the recrystallization was performed with EtOAc. N- (2,6-dihydroxy-9H-purin-8-yl) -propionamide compound having the following physical properties was obtained in a yield of 52.2% (61.1 mg).

¹ H NMR (400 MHz, DMSO-d ₆ ): 2.32 (m, 2H), 1.09 (t, J = 7.6 Hz, 3H); m / e 223 (M)

Example 13. 2,2-Dimethyl-N- (9H-purin-8-yl) -propionamide

N- (bis-methylsulfanyl-methylene) -2,2-dimethyl-propionamide (125 mg, 0.61 mmol), 4,5-diaminopyrimidine (67 mg, 0.61 mmol) of Reference Example 3-6, Experiment in the same manner as in Example 1 with DMF (2.4 ml) and recrystallized with EtOAc A 2,2-dimethyl-N- (9H-purin-8-yl) -propionamide compound having the following physical properties was obtained in a yield of 66.5% (88.7 mg).

¹ H NMR (400 MHz, DMSO-d ₆ ): 12.59 (br, NH), 11.71 (br, NH), 8.77 (s, 1H), 8.71 (s, 1H), 1.29 (s, 9H); m / e 219 (M)

Example 14. N- (6-hydroxy-9H-purin-8-yl) -2,2-dimethyl-propionamide

N- (bis-methylsulfanyl-methylene) -2,2-dimethyl-propionamide (115 mg, 0.56 mmol), 4,5-diamino-6-hydroxypyrimidine-hemisul in Reference Example 3-6 Experiment with Pate (98 mg, 0.56 mmol) and DMF (2.2 ml) in the same manner as in Example 1 and recrystallization with EtOAc N- (6-hydroxy-9H-purin-8-yl) -2,2-dimethyl-propionamide compound having the following physical properties was obtained in a yield of 73.7% (96.8 mg).

¹ H NMR (400 MHz, DMSO-d ₆ ): 7.97 (s, 1H), 3.32 (br, OH), 1.24 (s, 9H)

Example 15. N- (2,6-Dihydroxy-9H-purin-8-yl) -2,2-dimethyl-propionamide

N- (bis-methylsulfanyl-methylene) -2,2-dimethyl-propionamide (129 mg, 0.76 mmol), 5,6-diamino-2,4-dihydroxypyrimi of Reference Example 3-6 Din sulfate (82 mg, 0.76 mmol) and DMF (3.0 ml) were tested in the same manner as in Example 1, and the recrystallization was performed with EtOAc. N- (2,6-dihydroxy-9H-purine having the following physical properties: -8-yl) -2,2-dimethyl-propionamide compound was obtained in a yield of 35.2% (67 mg).

¹ H NMR (400 MHz, DMSO-d ₆ ): 1.22 (s, 9H); m / e 251 (M)

Example 16. N- (6-hydroxy-2-mercapto-7H-purin-8-yl) -2,2-dimethyl-propionamide

N- (bis-methylsulfanyl-methylene) -2,2-dimethyl-propionamide (122 mg, 0.55 mmol) and 4,5-diamino-6-hydroxy-2-mercaptopy of Reference Example 3-6 The same experiment as in Example 1 was carried out with limidinehemisulfate hydrate (113 mg, 0.55 mmol) and DMF (2.2 ml), and the recrystallization was performed with N- (6-hydroxy-2-) having the following physical properties as EtOAc. Mercapto-7H-purin-8-yl) -2,2-dimethyl-propionamide compound was obtained in a yield of 52.2% (611 mg).

¹ H NMR (400 MHz, DMSO-d ₆ ): 1.22 (s, 9H); m / e 267 (M)

Experimental Example 1. Evaluation of inhibition of lipoxygenase at the cellular level

In order to confirm the inhibitory activity against the lipoxygenase of the compound of the Example was carried out the following experiment by the method described in the reference.

1-1. Preparation of BMMC (Bone-marrow mast cell)

Two female BALB / c mice, 5-6 weeks old, were prepared, sterilized with 71% EtOH, and leg bones were separated. Insert the 25 G needle into the center of the leg bone, push out the bone marrow with 25 ml RPMI, centrifuge (1000 rpm, 5 mins), discard the supernatant and incubate with RPMI 1640/10% FBS. After a day or so, observed and passaged and confirmed more than 90% BMMC, and the following experiment was performed.

1-2. Rukotrien (LTC) ₄ ) Assay

Transfer the BMMC more than 3 weeks into a 50 ml tube (falcon), centrifuge (1000 rpm, 5 min, 4 ° C.) to discard the supernatant and add only RPMI 1640. 50 ul supernatant samples, 50 ul LTC ₄ AChE tracer and 50 ul LTC ₄ immunoserum were dispensed into the wells, and dilution was performed with EIA buffer. The NSB wells were dispensed with 100ul EIA buffer and 50ul LTC ₄ AChE tracer, and the B ₀ wells were dispensed with 50ul EIA buffer, 50ul LTC ₄ AChE tracer and 50ul LTC ₄ immunoserum. After incubation at room temperature for 16 to 20 hours, wash 5 times with 200 ul of wash buffer. 200 ul of Elman's reagent was added to each well, and after incubation for 60 minutes, the plate was read at 405 nm.

As a result of the experiment, it was confirmed that the purine derivative compounds of Examples 1, 2, 5 and 6 had excellent inhibitory activity against lipoxygenase (see Table 1).

Compounds (%) Inhibition at 1uM (%) Inhibition at 10uM Example 1 33 41 Example 2 50 48 Example 5 56 54 Example 6 57 60

The present invention relates to a purine derivative compound having a novel chemical structure having inhibitory activity against lipoxygenase, and a pharmaceutical composition comprising the compound containing the same as an active ingredient can be used as a medicament for treating and preventing asthma.

Claims (3)

Purine derivative compounds of the general formula (I) or pharmacologically acceptable salts thereof

(Ⅰ) Where R ₁ is one or more substituents selected from a hydrogen atom, a halogen atom, a hydroxyl group, a sulfanyl group or a lower alkyl group, R ₂ is a C ₁ -C ₆ lower acyl or benzoyl group unsubstituted or substituted with one or more R ′, R 'is a substituent which is a halogen atom or a C ₁ -C ₆ lower alkyl group. The method of claim 1, N- (9H-Purin-8-yl) -benzamide, N- (6-hydroxy-9H-purin-8-yl) -benzamide, N- (2,6-dihydroxy-9H-purine- 8-yl) -benzamide, N- (6-hydroxy-2-mercapto-7H-purin-8-yl) -benzamide, N- (7H-purin-8-yl) -isobutylamide, N -(6-hydroxy-9H-purin-8-yl) -isobutylamide, N- (9H-purin-8-yl) -3,5-bis-trifluoromethyl-benzamide, N- (6 -Hydroxy-9H-purin-8-yl) -3,5-bis-trifluoromethyl-benzamide, N- (9H-purin-8-yl) -acetamide, N- (9H-purin-8 -Yl) -propionamide, N- (6-hydroxy-9H-purin-8-yl) -propionamide, N- (2,6-dihydroxy-9H-purin-8-yl) -propionamide, 2,2-dimethyl-N- (9H-purin-8-yl) -propionamide, N- (6-hydroxy-9H-purin-8-yl) -2,2-dimethyl-propionamide, N- ( 2,6-dihydroxy-9H-purin-8-yl) -2,2-dimethyl-propionamide, N- (6-hydroxy-2-mercapto-7H-purin-8-yl) -2, 2-dimethyl-propionamide. A pharmaceutical composition for the treatment and prevention of asthma diseases, comprising the compound of formula (I) according to claim 1 or a pharmacologically acceptable salt thereof as an active ingredient.