WO2015034169A1 - Pharmaceutical composition containing 3-(6-(4-(trifluoromethoxy)phenylamino)pyrimidin-4-yl)benzamide or a pharmaceutically acceptable salt thereof as active ingredient for preventing or treating diabetes - Google Patents

Abstract

The present invention relates to a pharmaceutical composition and functional health food containing 3-(6-(4-(trifluoromethoxy)phenylamino)pyrimidin-4-yl)benzamide or a pharmaceutically acceptable salt thereof as an active ingredient for preventing or treating diabetes. The 3-(6-(4-(trifluoromethoxy)phenylamino)pyrimidin-4-yl)benzamide or the pharmaceutically acceptable salt thereof according to the present invention can be effectively used to treat diabetes including type 1 diabetes and type 2 diabetes, and the compound can be used as a functional health food for preventing or ameliorating diabetes.

Description

Pharmaceutical composition for the prevention or treatment of diabetes containing 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof as an active ingredient

The present invention relates to the prevention of diabetes containing 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof as an active ingredient or The present invention relates to a therapeutic pharmaceutical composition and dietary supplement.

The in vivo tyrosine kinase signaling pathway is important for cell proliferation, differentiation, metabolism, and cell migration regulation mechanisms, and abnormalities in this pathway are known to be closely related to the development of diseases such as cancer.

In particular, TKI development has been researched and developed in connection with cancer conquest, as imatinib (Gleevec or Glivec), a Bcr-Abl tyrosine kinase inhibitors (TKI), is known as a breakthrough in the treatment of chronic myelogeneous leukemia (CML).

3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide, known as GNF-2, is a non-ATP-competitive inhibitor and known as a Bcr-Abl inhibitor (Mian et al., 2012). Representative drugs of TKI include Imatinib, Nilotinib and Dasatinib, and the mechanism of action of these TKIs, known as Imatinib, is known to bind Bcr-Abl kinase to inhibit autophosphorylation and substrate phosphorylation of Bcr-Abl oncoprotein (Mokhtari and Welsh, 2010). TKI targets known to date include Abl-related gene (Arg), stem cell factor (SCF) recptor c-Kit, platelet-derived growth factor receptor (PDGFR), discoidin domain receptor 1/2 (DDR1 / 2) and oxidoreductase NQO2 (NADPH dehydrogenase quinone 2) and others (Hantschel et al., 2008).

Most TKI drugs, including Imatinib, have been shown to be excellent therapeutic agents for CML and to inhibit various cancer cell proliferations, including gastrointestinal stromal tumiors (GISTs). In particular, the study reports that administration of Imatinib to CML patients with chronic myeloid leukemia improves diabetes as well as CML (Breccia et al., 2004; Veneri et al., 2005). Attention was drawn to antidiabetic activity. Studies on the anti-diabetic activity of imatinib have been shown to protect against STZ-induced diabetes, improve diabetes in db / db mice as well as non-obese diabetic (NOD) mice (Han et al. , 2009).

Subsequently, TKIs such as Nilotinib, Sunitinib, Dasatinib, Erlotinib, and Fasudil have been reported to improve the type 1 or type 2 diabetes in association with the inhibition of beta-cell apoptosis and insulin resistance. It was recognized that. The mechanism of action on the anti-diabetic effect of TKI was studied through Imatinib study to inhibit protein kinase C (PKC) delta activation and to protect against pancreatic beta-cell apoptosis by blocking the stress-activated protein kinase pathway. (Hagerkvist et al., 2006).

Imatinib, Nilotinib and Dasatinib, which target Bcr-Abl used for CML treatment, were effective for early CML treatment but have limited treatment in patients with advanced disease stages CML. In particular, if there is a point mutation in the ATP binding site of the Abl kinase domain, these mutations do not act on Bcr-Abl, and thus the therapeutic efficacy of these drugs is reduced, thereby causing resistance to these drugs. That is, these drugs (Imatinib, Nilotinib, Dasatinib) target the ATP binding site of Abl tyrosine kinase, and thus are inhibitors of ATP competitive Abl tyrosine kinase.

Meanwhile, 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide (GNF-2) as an ATP noncompetitive, allosteric Bcr-Abl tyrosine kinase inhibitor has been reported (Nat Chem Biol. 2006 Feb. 2 (2): 95-102). GNF-2 has been reported that Abl tyrosine kinase is very effective for transforming cells (Adrian et al., 2006). Combination of GNF-2 with ATP-competitive Bcr-Abl kinase inhibitors (Imatinib / Nilotinib) has been shown to be effective in treating TKI resistant CML such as Bcr-Abl-T3151 (Nature. 2010 Jan 28; 463 (7280)): 501-6.). As mentioned above, the antidiabetic effect of Imatinib, a Bcr-Abl kinase, has been reported (Breccia et al., 2004; Veneri et al., 2005), and GNF-2 is also reported as a Bcr-Abl kinase inhibitor (Nat. Chem). Biol. 2006 Feb; 2 (2): 95-102; J, Biol. Chem. 2009 Oct; 284 (42): 29005-29014).

In addition, the significant involvement of oxidative stress in the development of diabetes has been mentioned in the study of the mechanisms of streptozotocin (STZ) and alloxan drugs used to induce diabetes, and Kubisch et al. Cu / Zn superoxide dismutase Diabetes was suppressed when induction of overexpression was observed, and the correlation between diabetes and vitamin C and vitamin E was also reported in clinical studies. This suggests that oxidative stress induced by hyperglycemia in vivo is associated with the activation of protein kinase C, which leads to increased glycation, increased intracellular osmotic pressure and degeneration following conversion to the polyol pathway, and enhanced expression of pro-inflammatory and inflammatory cytokine genes. It became known.

[Preceding technical literature]

[Patent Documents]

(Patent Document 1) US2012-0329798 A

[Non-Patent Documents]

(Non-Patent Document 1) Allosteric inhibition enhances the efficacy of ABL kinase inhibitors to target unmutated BCR-ABL and BCR-ABL T315I, Afsar Ali Mian et al., BMC Cancer 2012, 12: 411

(Non-Patent Document 2) The Ins and Outs of Bcr-Abl Inhibition, E. Premkumar Reddy et al., Genes & Cancer 3 (5-6) 447-454

The problem to be solved in the present invention is diabetes mellitus containing 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof as an active ingredient ( It is to provide a pharmaceutical composition for the prevention or treatment of diabetes).

Another object to be solved by the present invention is to provide a health functional food for the prevention or improvement of diabetes comprising the compound.

In order to solve the above problems, the present invention is a 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof as an active ingredient It provides a pharmaceutical composition for the prevention or treatment of diabetes containing (diabetes).

The diabetes is preferably type 1 diabetes or type 2 diabetes.

The 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof preferably inhibits apoptosis of pancreatic β-cells.

It is preferable that the 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof inhibits the production of reactive oxygen species (ROS). .

The 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof preferably inhibits JNK phosphorylation.

The 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof preferably inhibits the activation of PKCδ.

The present invention also provides a dietary supplement for preventing or ameliorating diabetes, including 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide.

The diabetes is preferably type 1 diabetes or type 2 diabetes.

3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof according to the present invention includes type 1 diabetes mellitus and type 2 diabetes mellitus It can be effectively used for the treatment of diabetes mellitus, and the compound can also be used as a dietary supplement for the prevention or improvement of diabetes.

1 is a result confirming the protective action of rat pancreatic β-cell of GNF-2.

2 is a result showing the effect on the ROS generation of GNF-2.

Figure 3 is a result showing the effect on stress-activated protein kinase (SAPK) of GNF-2.

Figure 4 is a result showing the effect on apoptosis by Protein kinase C (PKC) of GNF-2.

5 is a result showing the anti-diabetic activity of GNF-2 through in vivo experiments.

EMBODIMENT OF THE INVENTION Hereinafter, this invention is demonstrated in detail.

The inventors of the present invention described the 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide known as GNF-2, an ATP-noncompetitive allosteric Bcr-Abl kinase inhibitor. Antidiabetic activity was confirmed in vitro and in vivo and GNF-2 was found to be used for the prevention or treatment of diabetes.

Accordingly, the present invention is directed to diabees containing 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof as an active ingredient. Provided is a prophylactic or therapeutic pharmaceutical composition.

The diabetes is preferably type 1 diabetes or type 2 diabetes.

The 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof preferably inhibits apoptosis of pancreatic β-cells.

It is preferable that the 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof inhibits the production of reactive oxygen species (ROS). .

The 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof preferably inhibits JNK phosphorylation.

The 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof preferably inhibits the activation of PKCδ.

As the pharmaceutically acceptable salt of the compound of the present invention, acid addition salts formed by free acid are useful. 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. Equal molar amounts of the compound and acid or alcohol (eg, glycol monomethyl ether) 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, and hydrochloric acid, phosphoric acid, sulfuric acid, nitric acid, tartaric acid, and the like may be used as the inorganic acid, and methanesulfonic acid, p-toluenesulfonic acid, acetic acid, refluoroacetic 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, galacturonic acid, glutamic acid, glutaric acid, glucuronic acid, aspartic acid, ascorbic acid , Carbonic acid, vanic acid, hydroiodic acid and the like can be used.

Bases can also be used to make pharmaceutically acceptable metal salts. An alkali metal or alkaline earth metal salt is 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 salt, 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 compounds of the invention include acidic or basic salts which may be present in the compounds of the invention 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. It can be prepared through.

The pharmaceutical compositions of the present invention may be formulated in various forms, such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, oral formulations, injections of sterile injectable solutions, etc. Can be used.

Such pharmaceutical compositions may further include carriers, excipients or diluents, and examples of suitable carriers, excipients or diluents that may be included include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, Starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, amorphous cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil Etc. can be mentioned. In addition, the pharmaceutical composition of the present invention may further include a filler, an anticoagulant, a lubricant, a humectant, a perfume, an emulsifier, a preservative, and the like.

In a preferred embodiment, solid preparations for oral administration include tablets, pills, powders, granules, capsules and the like, which solid preparations comprise at least one excipient such as starch, calcium carbonate, Sucrose, lactose, gelatin and the like are mixed and formulated. In addition, lubricants such as magnesium stearate, talc and the like may also be used in addition to simple excipients.

As a preferred embodiment, oral liquid preparations may be exemplified by suspensions, solvents, emulsions, syrups, and the like, and various excipients, for example, wetting agents, sweeteners, Fragrances, preservatives and the like.

As a preferred embodiment, the preparation for parenteral administration may include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilizers, suppositories and the like. Non-aqueous solvents and suspending agents may include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, injectable esters such as ethyl oleate, and the like. Injectables may include conventional additives such as solubilizers, isotonic agents, suspending agents, emulsifiers, stabilizers, preservatives, and the like.

The pharmaceutical composition of the present invention is administered in a pharmaceutically effective amount. In the present invention, “pharmaceutically effective amount” means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment, and an effective dose level means the type, severity, activity of the drug, Sensitivity to drug, time of administration, route of administration and rate of release, duration of treatment, factors including concurrent use of drugs, and other factors well known in the medical arts. The pharmaceutical compositions of the present invention may be administered as individual therapeutic agents or in combination with other therapeutic agents, may be administered sequentially or simultaneously with conventional therapeutic agents, and may be administered as single or multiple doses. Taking all of the above factors into consideration, it is important to administer an amount that can obtain the maximum effect in a minimum amount without side effects, which can be easily determined by those skilled in the art.

In a preferred embodiment, the effective amount of the compound in the pharmaceutical composition of the present invention may vary depending on the age, sex and weight of the patient, and generally 1 to 5,000 mg, preferably 100 to 3,000 mg per kg body weight daily or It can be administered every other day or divided into 1 to 3 times a day. However, the dosage may be increased or decreased depending on the route of administration, the severity of the disease, sex, weight, age, etc., and the above dosage does not limit the scope of the present invention in any way.

The pharmaceutical composition of the present invention can be administered to a subject through various routes. All modes of administration can be expected, for example by oral, rectal or intravenous, intramuscular, subcutaneous, intrauterine dural or intracerebroventricular injection.

As used herein, "administration" means providing a patient with any substance by any suitable method, wherein the route of administration of the pharmaceutical composition of the present invention is oral or parenteral via all common routes as long as the target tissue can be reached. Oral administration. In addition, the composition of the present invention may be administered using any device capable of delivering an active ingredient to a target cell.

"Subject" in the present invention is not particularly limited, but includes, for example, humans, monkeys, cattle, horses, sheep, pigs, chickens, turkeys, quails, cats, dogs, mice, rats, rabbits or guinea pigs. And preferably mammals, and more preferably humans.

The present invention also provides a dietary supplement for preventing or ameliorating diabetes, including 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide.

The diabetes is preferably type 1 diabetes or type 2 diabetes.

The dietary supplement containing the compound of the present invention includes, for example, various foods, beverages, gums, teas, vitamin complexes, and the like. It may also be added to foods or beverages for the purpose of preventing diabetic diseases. At this time, the amount of the compound in food or beverage may be added in 0.01 to 15% by weight of the total weight of the food, beverages may be added in a ratio of 0.02 to 5g, preferably 0.3 to 1g based on 100ml.

Health functional food of the present invention includes the form of tablets, capsules, pills, liquids and the like.

The dietary supplement of the present invention is not particularly limited to other ingredients containing the above-mentioned preferred ratios of the compound, and may contain various flavors or natural carbohydrates as additional ingredients, such as conventional foods or drinks. Examples of the above-mentioned natural carbohydrates include monosaccharides such as glucose, fructose and the like; Disaccharides such as maltose, sucrose and the like; And conventional sugars such as polysaccharides such as dextrin, cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents other than those mentioned above, natural and synthetic flavoring agents can also be used advantageously. The proportion of said natural carbohydrates is generally about 1 to 20 g, preferably about 5 to 12 g per 100 ml of the composition of the present invention.

In addition to the above, the compounds of the present invention include various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic flavors and natural flavors, coloring and neutralizing agents (such as cheese and chocolate), pectic acid and salts thereof, alginic acid and its Salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, carbonation agents used in carbonated drinks and the like. In addition, the compounds of the present invention may contain fruit flesh for the production of natural fruit juices and fruit juice beverages and vegetable beverages. These components can be used independently or in combination. The proportion of such additives is not so critical but is usually selected in the range of 0 to about 20 parts by weight per 100 parts by weight of the compound of the present invention.

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

EXAMPLE

1. Materials and Methods

Cell experiments and animal experiments were performed in this study. First, cell experiments were performed by treating streptozotocin (STZ) on rat pancreatic β-cell line (INS-1) and rat pancreatic β of GNF-2 for apoptosis caused by oxidative stress. The protective activity against cell death of -cell line (INS-1) was investigated.

Inhibition of JNK phosphorylation in the SAPK pathway where the cytoprotective activity of allosteric Bcr-Abl tyrosine kinase inhibitor, 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide, is activated by stress We compared the expression and phosphorylation trends of each protein by using Wester blot for the relationship between the activation of protein kinase delta and caspase-3.

In the animal experiments, mice were fed with GNF-2 (30mg / kg) and non-feeding groups with STG 150mg / kg in single-dose groups, and then weight and blood glucose levels were measured and compared.

2. Results and Discussion

2.1. GNF-2 protects rat pancreatic β-cell

GNF-2 is an ATP-competitive, unlike Imatinib, Nilotinib and Dasatinib as allosteric Bcr-Abl kinase inhibitors. The antidiabetic effect of Icrinib, a Bcr-Abl kinase, has been reported, GNF-2 has been reported as a Bcr-Abl kinase inhibitor, and according to clinical reports that several TKIs have antidiabetic activity, the antidiabetic activity of GNF-2 has been reported in rat pancreatic. The assay was carried out using β-cell line (Ins-1) and C57BL / 6J mice.

first Cytotoxicity of GNF-2 against INS-1 cells was measured. As shown in FIG. 1A, it was found that INS-1 cells were killed by STZ, whereas cells pretreated with GNF-2 at 7.5 μM for 1 hour were protected from cell damage by STZ. This could be confirmed by PI and DAPI staining (FIG. 1B).

2.2. Effect of GNF-2 on Reactive Oxygen Species Production

Free radicals are produced through metabolism of the living body, which is unstable and acts on DNA and protein of surrounding cells, causing mutations or degeneration, causing cell damage and inducing cell death.

STZ is a powerful DNA methylating agent known to act as a free radical donor in the pancreas (Larsen and Grude, 1978; Newsholme et al. , 2007). That is, in the diabetes model induced by STZ treatment, ROS are generated by STZ, and the generated ROS attack pancreatic β-cells to induce β-cell apoptosis, which leads to diabetes. Therefore, we investigated whether the cytoprotective activity of GNF-2, which protects INS-1 cells from STZ, is associated with inhibition of ROS production induced by STZ as inhibitory activity against DCF production, a fluorescent form of DHCF-DA.

As shown in Figure 2, when the ROS generated by streptozotocin treatment to 100% when GNF-2 for 1 hour pretreatment before streptozotocin treatment was confirmed that the inhibition of ROS production by 73.4%.

2.3. Effect of GNF-2 on Stress-activated Protein Kinase (SAPK)

Apoptosis is a mechanism in which cells are killed when cells are exposed to external stress, followed by cell membrane contraction, mitochondria depolarization, proteolysis, chromosome condensation, and DNA fragmentation. Such apoptosis signaling systems are known to involve mitogen-activated protein kinase family (MAPK) and protein kinase C (PKC).

First, the MAPK pathway is largely divided into extracellular signal-regulated kinases (ERK1 / 2), c-Jun NH ₂ -terminal kinases (JNK-1 / 2/3), and p38 MAPK proteins (p38α / β / γ / δ). . Double _{c-Jun NH 2 -terminal kinases (} JNK-1/2/3) and the p38 MAPK proteins (p38α / β / γ / δ) is the cells are UV light, γ-irradiation, DNA damage, heat and osmotic shock or oxidative Apoptosis pathway induced when under stress (English et al., 1999; Hagemann and Blank, 2001; Huang et al., 1999).

Phosphorylation of Bcr-Abl is known to induce activation of stress-activated protein kinase (JNK and p38 MAP-kinases) and to induce apoptosis by activating caspase 9. Imatinib is known to inhibit phosphorylation of nonreceptor protein kinase c-Abl. JNK in stress-activated protein kinase It has been reported to have antidiabetic activity by inhibiting phosphorylation and protecting apoptosis of pancreatic β-cell (Hagerkvist et al., 2006). In addition, the cytoprotective activity of Imatinib was found to inhibit apoptosis through inhibition of cNK and phosphorylation of JNK under oxidative stress by H ₂ O ₂ of myotubes as well as panceatic β-cell. It has been reported to block apoptosis by blocking downstream signaling system JNK pathway following Abl activation (Hagerkvist et al., 2006). Therefore, the effect of GNF-2, a Bcr-Abl kinase inhibitor, on the JNK pathway was investigated.

As shown in FIG. 3, treatment of INS-1 cells with 1 mM STZ for 8 hours resulted in almost no change in total JNK, but drastically induced phosphorylation of JNK-1 and -2. Such activation of JNK resulted in GNF-2 7.5. Pretreatment with μM for 1 hour or treatment with J600 inhibitor SP600125 confirmed that it was rapidly inhibited. From these results, GNF-2, like Imatinib, inhibited JNK phosphorylation and inhibited cell death.

2.4. Effect of GNF-2 on Apoptosis by Protein Kinase C Delta

When cells are subjected to genotoxic stress, they determine cell survival and death depending on the extent of their damage and are either recovered or removed through cell death. To date, the complete mechanism for this is unknown, but protein kinase Cδ isoform is activated by c-Abl phosphorylation in apoprotic cell death, and then PKCδ cleaved by activated caspase-3 moves into the nucleus and induces apoptosis. Reported (Ghayur et al., 1996).

Therefore, as a result of investigating the changes of PKCδ cleavage and caspase-3 according to streptozotocin treatment in INS-1 cells, cleaved PKCδ and caspase 3 were activated when treated with STZ 1 mM for 8 hours as shown in FIG. 4, and their activation was GNF- It was found to be inhibited by 2.

Taken together, GNF-2 inhibits JNK phosphorylation and PKCδ activation against oxidative stress produced by STZ in INS-1 cells, thereby inhibiting pancreatic β-cell apoptosis. It was confirmed to suppress.

2.5. In vivo  Antidiabetic activity assay by GNF-2 through experiment

In vitro experiments showed that GNF-2 inhibits apoptosis of INS-1 cells by inhibiting JNK activation, PKCδ and caspase 3 activation against genotoxic stress induced by STZ.

Therefore, in vivo experiments were conducted using C57BL / 6J mice to determine whether there was antidiabetic activity associated with the oxidative stress protective activity of these pancreatic β-cells. First, GNF-2 (30mg / kg) dissolved in PBS was gavaged in C57BL / 6J mice, and streptozotocin was administered at 150mg / kg after 2 hours of gavage to induce diabetes. In the same way, anti-diabetic activity was compared with the negative and positive control groups that gavaged the same amount of PBS and Imatinib instead of GNF-2. After the first gavage, the same amount of PBS, GNF-2 and Iamtinib were gaved once daily for 14 days, and after 2 hours after gavage, the blood glucose level was checked with a One Touch glucometer (Lifescan, Milpitas, CA) and the weight was measured.

As shown in A of FIG. 5, the weight change did not change significantly in the control group for 14 days, whereas the GNF-2 treated group showed temporary weight loss at 6 days after gavage and then recovered. At this time, Imatinib-treated group also showed weight loss around 6 days as GNF-2 treated group, but unlike GNF-2, body weight did not recover.

Anti-diabetic activity of GNF-2 is maintained in the blood glucose level up to 400 or more after 4 days with streptozotocin treatment as shown in FIG. 5B, whereas the GNF-2 administration group has a blood glucose level from day 5 of GNF-2 administration. This was adjusted to around 200. At this time, the Imatinib-treated group improved blood sugar level to 400 when the control blood sugar level was 600 on the 7th day of administration, but did not reach the antidiabetic activity of GNF-2.

In summary, the allosteric Bcr-Abl kinase GNF-2 also inhibits the phosphorylation and activation of PKCδ by JNK, which induces apoptosis against oxidative stress, such as Imatinib, Nilotinib and Dasatinib, which target Bcr-Abl. It has been confirmed that it inhibits apoptosis of pancreatic β-cell, and this protective effect from apoptosis of pancreatic β-cell has an excellent effect on diabetes induced by STZ treatment, especially type 1 diabetes. .

[National R & D project supporting this invention]

[Project unique number] A111345

[Ministry of Health] Ministry of Health and Welfare, Republic of Korea

[Professional Research Institute] Korea Health Industry Development Institute

[Name of research project] Health and medical R & D project-Leading specialization project (non-capital area)

[Contribution rate] 100%

[Organization] Kyungpook National University Hospital

[Research Period] 2012. 10. 01 ~ 2016. 03.31

Claims (8)

1. Pharmaceutical for the prophylaxis or treatment of diabetes mellitus containing 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof as an active ingredient Composition.
2. The pharmaceutical composition of claim 1, wherein the diabetes is type 1 diabetes or type 2 diabetes.
3. The method of claim 1, wherein the 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof is characterized by apoptosis of pancreatic β-cells. Pharmaceutical composition, characterized in that inhibiting.
4. The method of claim 1, wherein the 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof is used to generate a reactive oxygen species (ROS). Pharmaceutical composition, characterized in that to inhibit.
5. The method of claim 1, wherein the 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof is characterized in that it inhibits JNK phosphorylation. Pharmaceutical compositions.
6. The method of claim 1, wherein the 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide or a pharmaceutically acceptable salt thereof is characterized in that it inhibits the activation of PKCδ. Pharmaceutical compositions.
7. A dietary supplement for the prevention or improvement of diabetes, comprising 3- (6- (4- (trifluoromethoxy) phenylamino) pyrimidin-4-yl) benzamide.
8. The health functional food according to claim 7, wherein the diabetes is type 1 diabetes or type 2 diabetes.

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