KR101042697B1 - Inhibitor for hepatic fibrosis - Google Patents

Abstract

It inhibits hepatic fibrosis that occurs continuously in chronic hepatitis, thereby providing a safe hepatic fibrosis inhibitor.

General formula

In the formula, R ¹ is an alkyl group having 4 to 8 carbon atoms, R ² represents a hydrogen atom, an alkylcarbonyl group having 2 to 6 carbon atoms or an alkoxycarbonyl group having 2 to 6 carbon atoms or a cyclodextrin clathrate thereof. It is a hepatic fibrosis inhibitor characterized by including a oxyquinone derivative as an active ingredient.

Liver fibrosis inhibitor

Description

Liver Fibrosis Inhibitor {INHIBITOR FOR HEPATIC FIBROSIS}

The present invention relates to liver fibrosis inhibitors.

Hepatic fibrosis is caused by viral hepatitis, alcoholic liver injury, autoimmune liver disease and metabolic liver disease, which promote the production of fibrous tissue called collagen or extracellular matrix composed of collagen or complex sugars during necrotic hepatocyte repair. As the inflammation progresses, these fibrous tissues gradually accumulate.

In the early stages of inflammation, the proliferating and growing extracellular matrix immediately heals without being absorbed and accumulated, but in chronic hepatitis in which the inflammatory condition persists, the production of the matrix exceeds the degradation / absorption, and the fibrous tissue continues to accumulate. As a result, the vicious cycle that the liver parenchymal cells pressurized by the accumulated fibrous tissue is disturbed and the production of the fibrous tissue is accelerated is repeated, and the cirrhosis of the liver is cured by the fibrous tissue in the near future.

If cirrhosis has been reached, it is extremely difficult to repair it, and in many cases it leads to liver cancer. In particular, hepatitis C, which is a social problem due to infection by administration of contaminated blood products, progresses from chronic hepatitis to cirrhosis to liver cancer at a high rate. Thus, while healing to suppress inflammation and fibrosis is important, there are few side effects and few therapeutic drugs with high efficacy.

Recently, hepatic stellate cells and TGF-β are considered to play an important role in the progression of liver fibrosis. Hepatic stellate cells are activated by cytokines released by inflammatory cells such as macrophages to actively produce extracellular matrices starting with type I collagen. On the other hand, TGF-β is expressed as a cytokine that promotes the transformation and proliferation of normal fibroblasts, and has the effect of promoting the proliferation of activated stellate cells, promoting the production of extracellular matrix and inhibiting the proliferation of hepatic parenchymal cells. Therefore, development of the therapeutic drug which suppresses the action of TGF- (beta), or suppresses excessive production of TGF- (beta) about progression of liver fibrosis is progressing.

From this point of view, for example, integrin inhibitors in patent document 1, ALK5 inhibitors in patent document 2, anti-TGF-β receptor peptides in patent document 3, and the like, have been described as anti-TGF-β action or TGF-β production inhibitory action. It is being examined as an inhibitor. For example, Patent Document 4 discloses that it has a NO production inhibitory effect and that an inhibitor of TGF-β production and a hepatic fibrosis inhibitory effect are recognized by an iNOS activity inhibitor having a specific structure.

On the other hand, the hydroxyquinone derivative represented by the following general formula (1) is a substance having a strong antioxidant action and NO production inhibitory action, antioxidant by using the present invention as an active ingredient (patent document 5), chronic The invention regarding the therapeutic drug (patent document 6), carcinogenic inhibitor (patent document 7), and atherosclerosis therapeutic agent (patent document 8) of intractable inflammatory diseases, such as a rheumatoid arthritis or a nonspecific inflammatory bowel disease, is described.

[Formula 1]

In addition, Patent Literature 9 of the present inventors describes a composition for treating liver disease using the present compound as an active ingredient, and is directed to α-naphthylisothiocyanate (ANIT) -induced acute liver injury in mice. The efficacy test is specifically described by the pharmacological test example, and the safety test example discloses that the safety is also high.

[Patent Document 1] Publication No. 2002-530431

[Patent Document 2] Japanese Patent Application Laid-Open No. 2005-343889

[Patent Document 3] Japanese Patent Application Laid-Open No. 2007-186519

[Patent Document 4] Japanese Patent Application Laid-Open No. 2005-41837

[Patent Document 5] Publication No. 5-301836

[Patent Document 6] Japanese Patent Application Laid-Open No. 2004-352661

[Patent Document 7] Publication No. 6-100441

[Patent Document 8] Japanese Patent Application Laid-Open No. 2002-241366

[Patent Document 9] Japanese Patent Application Laid-Open No. 8-67627

However, all of the compounds described in Patent Documents 1-4 are being examined as liver fibrosis inhibitors, and no effective drug for inhibiting the progression of liver fibrosis and cirrhosis has yet been found. Also about the compound of General formula (1), neither the function of inhibiting liver fibrosis is described in patent document 5-9.

Thus, the present inventors have found that the antibody titer of Naofen (GenBank Accession No .: EF613262), a newly discovered in vivo protein as an anti-Verotoxin II-antibody reactive substance, is a model for inducing cirrhosis and liver fibrosis by carbon tetrachloride (CCl4) in rats. Found to rise. Thus, it was found that Naofen is involved in cirrhosis and liver fibrosis prior to the production of TGF-β because the increase occurs prior to the production of TGF-β and collagen production.

Therefore, in order to find the substance which has the inhibitory effect of hepatic fibrosis, the substance which has the activity which suppresses the rise of the antibody titer of intrahepatic Naofen in this model, earnestly researched. Thus, from the fact that the hydroxyquinone derivative of the general formula (1) has an inhibitory effect on liver damage, it was predicted to have an inhibitory activity of Naofen and further research was conducted.

On the other hand, the causal relationship between hepatic fibrosis, TGF-β production, and NO derived from inducible NO synthase has not been revealed to date, and it is not known whether the NO production inhibitory action acts as an effective hepatic fibrosis inhibitory effect. The present inventors predict that the hydroxyquinone derivative of the general formula (1) has NO production inhibitory activity, inhibits production of TGF-β similarly to the iNOS activity inhibitor, and has hepatic fibrosis inhibitory activity, Further research was conducted.

As a result, the present invention has been found to have a function of inhibiting Naofen expression and also inhibiting TGF-β mRNA expression, inhibiting liver fibrosis, and being useful as a liver fibrosis inhibitor, thereby completing the present invention.

In other words, the present invention is the following general formula (1)

[Formula 2]

In the formula, R ¹ represents an alkyl group having 4 to 8 carbon atoms, R ² represents a hydrogen atom, an alkyl carbonyl group having 2 to 6 carbon atoms or an alkoxycarbonyl group having 2 to 6 carbon atoms. It is to provide a hepatic fibrosis inhibitor comprising as a component.

Thus, these compounds inhibit the expression of Naofen, a substance involved in liver fibrosis prior to TGF-β. The details of the mechanism by which such compounds inhibit Naofen expression have not been revealed to date, but the present inventors have found that the inhibition of the activation of IκB-kinase, similar to the inhibition of NO production through the mechanism of inhibition of phosphorylation of IκB of hydroxyquinone derivatives, is observed. It is predicted that the possibility of suppressing Naofen by the mechanism through. In addition, these compounds have an inhibitory effect on NO production and, similar to iNOS inhibitors, inhibit the production of TGF-β which promotes fibrosis of the liver. By this action, the production of collagen in the fibrotic liver is suppressed, the accumulation of fibrous tissue is suppressed, and the fibrosis of the liver is suppressed.

In addition, the compound represented by the formula (2) is 2,3,5-trimethylhydroxyquinone-1-hexyl ether or 2,3,5-trimethylhydroxyquinone-1-hexyl ether 4-acetate to provide a liver fibrosis inhibitor will be.

The hydroxyquinone derivative of the present invention composed of the compound represented by the formula (2) significantly inhibits hepatic fibrosis by inhibiting Naofen expression related to hepatic fibrosis and also inhibiting the production of TGF-β. It is also safe for the liver. For this reason, the composition of this invention which uses such a hydroxyquinone derivative as an active ingredient can be effectively used as a liver fibrosis inhibitor.

In addition, 2,3,5-trimethylhydroxyquinone-1-hexyl ether or 2,3,5-trimethylhydroxyquinone-1-hexyl ether 4-acetate is particularly effective in terms of pharmacological activity and biocompatibility. Can be used.

Hereinafter, the present invention will be described in detail.

The compound represented by the general formula (2) contained in the hepatic fibrosis inhibitor of the present invention may be any linear, branched or cyclic alkyl group having 4 to 8 carbon atoms represented by R ¹ , and examples thereof include various butyl groups and various pens. And a hexyl group, various hexyl groups, various heptyl groups, various octyl groups, cyclobutyl groups, cyclopentyl groups, cyclohexyl groups, cycloheptyl groups, and cyclooctyl groups. From the viewpoint of pharmacological activity and biocompatibility, a straight chain having 4 to 7 carbon atoms is preferable, and n-hexyl group is particularly suitable.

In addition, the C2-C6 alkyl carbonyl group in R <^2> may be linear, branched, or cyclic, and an acetyl group, a propionyl group, butyryl group, an isobutyryl group etc. are mentioned as an example. In addition, the C2-C6 alkoxycarbonyl group in R <^2> may be linear, branched, or cyclic | annular, and the methoxycarbonyl group, an ethoxycarbonyl group, propoxycarbonyl group, isopropoxycarbonyl group etc. are mentioned as an example.

Among the compounds of the formula (2), particularly preferred compounds in terms of pharmacological activity are 2,3,5-trimethylhydroxyquinone-1-butyl ether, 2,3,5-trimethylhydroxyquinone-1-hexyl ether or 2,3 And 5-trimethylhydroxyquinone-1-hexyl ether 4-acetate.

The compound represented by the said Formula (2) can be manufactured by the method of patent document 5, for example.

The hepatic fibrosis inhibitor of the present invention contains the hydroxyquinone derivative represented by the above-mentioned formula (2) as an active ingredient, and is formulated by adding an additive that is acceptable as an additive for pharmaceuticals such as a carrier for carrier or an excipient. Oral dosage forms of formulations suitable for absorption from the digestive tract, such as tablets, granules, capsules, oral solution, parenteral administrations such as injectables, suppositories, and transdermal absorbents such as tape and pape, and also solid, liquid, circulating, Due to its preservation properties, all forms conventionally practiced, such as dissolving a solid solvent with a suitable solvent at the time of use, can be appropriately used. In addition, in order to improve the bioavailability or stability of the present compound, it is also possible to use a delivery system including preparation techniques such as microcapsules, micropowders, and inclusions.

The dosage can not be collectively defined because it varies depending on the target therapeutic effect, the method of administration, the age, the weight, and the like, but the daily parenteral dosage is usually about 0.01 to 100 mg per body weight, preferably Is about 0.05-10 mg. Orally about 0.1 to 300 mg, preferably about 0.5 to 100 mg, it may be administered by dividing it 1 to 5 times per day.

[Example]

Next, although a test example demonstrates this invention further in detail, this invention is not limited at all by a test example.

(Action in Rat Carbon Tetrachloride-Induced Cirrhosis Model)

(Test Example 1)

Six-week-old Wister male rats were subcutaneously administered with 40% carbon tetrachloride-containing olive oil or olive oil twice a week for 3 weeks at 3 ml / kg. In addition, 2,3,5-trimethylhydroxyquinone-1-hexyl ether (Compound 1) was dissolved in drinking water at a concentration of 0.3 mg / ml in the hydroxyquinone derivative represented by General Formula (1). In parallel with the administration of carbon tetrachloride, the group administered daily and the group administered only water were set. That is, four groups of the olive oil administration group, the olive oil + compound administration group, the carbon tetrachloride administration group, and the carbon tetrachloride + compound administration group were composed, and 10 animals were assigned to each group. Animals were slaughtered 8 weeks after the start of carbon tetrachloride administration and examined. As a test item, the antibody titer in serum against Naofen was measured by ELISA, and the mRNA expression level of Naofen in liver was measured by RT-PCR method.

As shown in FIG. 6, the inventors found that Naofen expression was significantly increased in the early stage of cirrhosis progression in the carbon tetrachloride-administered group. Moreover, since the expression occurs earlier than the expression of TGF-β shown in FIG. 7 and the expression of intrahepatic collagen shown in FIG. 8, it was also found that the expression occurs earlier than the expression of TGF-β in the organization of cirrhosis.

The mRNA expression amount in this test is shown in FIG. It was found that the increase in the amount of Naofen expression was suppressed when Compound 1 was administered (6 weeks (+), 8 weeks (+)).

The Fischer Direct Probability Table of the measurement result of the antibody titer against Naofen in serum is shown in Table 1. In the compound 1 administration group (+), the antibody value increased more than 16 times compared with the olive oil administration group (-).

TABLE 1

HX V> = 16 4 <V <16 V <= 4 Sum - 7 0 One 8 + 2 One 4 7 Sum 9 One 5 15

From these results, Compound 1 inhibits the elevated expression of Naofen in liver cirrhosis and hepatic fibrosis model, that is, inhibits the production of liver cirrhosis and liver fibrosis in a path different from the inhibition of TGF-β by inhibiting NO production. It was shown.

(Test Example 2)

A rat carbon tetrachloride-induced cirrhosis model was used under the same conditions as in Test Example 1 except that the concentration of Compound 1 was 0.1 mg / ml. As a test item, measurement of blood AST, ALT, albumin (ALB), total bilirubin (T-Bil) and hepatic fibrosis markers, hyaluronic acid (HA) concentration, autopsy, body weight and organ weight, The TGF-β1 mRNA and type I collagen α1 mRNA expression levels in hepatocytes were measured by RT-PCR and histopathological examination was performed.

In the test results, the results of the measurement of blood AST, ALT, and HA concentrations are shown in Table 2, the liver and spleen weights, and the number of ascites in Table 3, and the results of measurement of the expression levels of TGF-β1 mRNA and type I collagen α1 mRNA in FIG. And 3, liver tissue photographs by Masson staining and immunohistochemical staining are shown in FIGS. 4 and 5 (A: olive oil administration, B: carbon tetrachloride administration, C: olive oil + compound administration 1, D: carbon tetrachloride + compound administration Yes, the lower right horizontal line is shown at 100 μm).

TABLE 2

Effect on Hemochemical Test Values in Rat Carbon Tetrachloride-Induced Cirrhosis Model

Inspection items olive oil Olive Oil + Compound 1 CCl ₄ CCl ₄ + Compound 1 AST (IU / L) 195.7 ± 90.4 192.7 ± 93.4 2215.5 ± 110.2 ^*** 603.6 ± 126.3 ^{** ++} ALT (IU / L) 94.7 ± 35.1 93.7 ± 34.1 1418.3 ± 181.2 ^*** 422.4 ± 60.5 ^{** ++} ALB (g / dL) 4.3 ± 0.1 4.3 ± 0.1 3.1 ± 0.1 ^** 3.8 ± 0.2 ^{* +} T-Bil (mg / dL) 0 0 0.34 ± 0.05 ^* 0.03 ± 0.02 ⁺ HA (ng / mL) 131.0 ± 19.5 136.7 ± 16.6 435.0 ± 19.7 ^** 237.7 ± 36.6 ^{* +}

Compound 1: 2,3,5-trimethylhydroxyquinone-1-hexyl ether

Values are mean ± standard deviation (n = 10)

^** : p <0.01 ^*** : p <0.001 (compared to olive oil group or olive oil + compound 1 administration group)

⁺ : p <0.05 ⁺⁺ : <0.01 (compared to CCl ₄ group)

TABLE 3

Effect on Body Weight, Hepatic and Spleen Weight, and Ascites in Rat Carbon Tetrachloride-Induced Cirrhosis Model

Inspection items olive oil Olive Oil + Compound 1 CCl ₄ CCl ₄ + Compound 1 Weight (g) 358.3 ± 6.1 361.5 ± 1.5 226.5 ± 7.9 ^** 240.8 ± 8.7 ^* Soy sauce relative weight 14.4 ± 1.6 14.8 ± 1.5 10.9 ± 2.1 ^* 16.1 ± 1.8 ⁺ Spleen Relative Weight 0.20 ± 0.01 0.20 ± 0.01 0.38 ± 0.06 ^* 0.28 ± 0.01 ⁺ Multiple numbers of storage animals 0 0 8 One

Compound 1: 2,3,5-trimethylhydroxyquinone-1-hexyl ether

Values are mean ± standard deviation (n = 10)

^* : p <0.05 ^*** : p <0.001 (compared to olive oil group or olive oil + compound 1 group)

⁺ : p <0.05 (compared to CCl ₄ group)

From the results shown in Tables 2 and 3 and Figs. 2 and 3, Compound 1 was shown to inhibit the increase in AST, ALT and hyaluronic acid concentrations, and the increase in the expression levels of TGF-β1 mRNA and type I collagen α1 mRNA by administration of carbon tetrachloride. appear. In histopathological examinations shown in FIGS. 4 and 5, it was confirmed that the accumulation of fibrous tissue and the expression of TGF-β1 were inhibited by the administration of Compound 1.

In addition, in both Test Examples 1 and 2, no particular abnormality was found in the olive oil + Compound 1 administration group, and it was found that Compound 1 had high safety and was extremely useful as a liver fibrosis inhibitor.

BRIEF DESCRIPTION OF THE DRAWINGS It is a figure which shows the effect | action of the compound 1 on the expression amount of Naofen mRNA in the liver.

Fig. 2 is a diagram showing the action of compound 1 on the expression level of TGF-β1 mRNA in the liver.

Fig. 3 shows the action of compound 1 on the expression level of type I collagen α1 mRNA in the liver.

4 is a diagram showing that accumulation of fibrous tissue is reduced by administration of Compound 1. FIG.

FIG. 5 shows that TGF-β1 expression is inhibited by administration of Compound 1. FIG.

It is a figure which shows the expression amount of Naofen mRNA in the liver of the rat carbon tetrachloride-induced cirrhosis model.

Fig. 7 shows the expression levels of TGF-β1 mRNA in the liver of rat carbon tetrachloride-induced cirrhosis model.

It is a figure which shows the expression amount of collagen mRNA in the liver of the rat carbon tetrachloride-induced cirrhosis model.

Claims (2)

(A) general formula

2,3,5-trimethyl in the compound represented by (wherein, R ¹ represents an alkyl group having 4 to 8 carbon atoms, R ² represents a hydrogen atom, an alkylcarbonyl group having 2 to 6 carbon atoms or an alkoxycarbonyl group having 2 to 6 carbon atoms) A hydroxyquinone derivative composed of hydroxyquinone-1-hexyl ether or 2,3,5-trimethylhydroxyquinone-1-hexyl ether 4-acetate as an active ingredient and having an action of inhibiting Naofen expression Hepatic fibrosis inhibitor. delete

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