KR20170055626A

KR20170055626A - Pharmaceutical compositions for treatment or prevention of idiopathic pulmonary fibrosis

Info

Publication number: KR20170055626A
Application number: KR1020150158469A
Authority: KR
Inventors: 소홍섭; 오기수; 이수빈
Original assignee: (주)나디안바이오
Priority date: 2015-11-11
Filing date: 2015-11-11
Publication date: 2017-05-22
Also published as: KR101845862B1

Abstract

The present invention relates to a pharmaceutical composition for treating or preventing idiopathic pulmonary fibrosis comprising -lapachone or dunnione, a pharmaceutically acceptable salt thereof, a prodrug thereof, a solvate thereof or an isomer thereof as an active ingredient. According to the present invention, a mouse animal model having the idiopathic pulmonary fibrosis is prepared using bleomycin, and an effect of administering beta-lapachone or dunnione is confirmed. As a result, the infiltration of various inflammatory cells of pulmonary tissues caused by the bleomycin, the density of tissue and the degree of fibrosis due to the accumulation of the extracellular matrix are reduced, an expression of IL-6, TGF-1, and SMA increased by the bleomycin is reduced, a survival rate increases when the dunnione is co-administered after administering the bleomycin, and also, a significant increase in the ratio of NAD^+/NADH is confirmed. Accordingly, the -lapachone or dunnione can be effectively used in treatment or prevention of the idiopathic pulmonary fibrosis.

Description

[0001] The present invention relates to a pharmaceutical composition for the treatment or prevention of idiopathic pulmonary fibrosis,

The present invention relates to a pharmaceutical composition for the treatment of idiopathic pulmonary fibrosis comprising beta-lapachone or dunnione, or a pharmaceutically acceptable salt thereof, prodrug thereof, a solvate thereof or an isomer thereof, To a pharmaceutical composition for the treatment or prevention of idiopathic pulmonary fibrosis.

Interstitial lung disease is an interstitial fibrosis caused by epilepsy, such as alveoli, alveoli, capillaries, bronchioles, and lymphatic vessels. Fibrosis is the most frequent incidence, and it is an important disease when considering the treatment method and prognosis of the disease. The pathogenesis of pulmonary fibrosis has not yet been well known and the treatment method has not yet been established. It is the degree of control of other complications. Although many studies have been done to elucidate the mechanism of pulmonary fibrosis, methods that can effectively control fibrosis are not available.

The indicators of pulmonary fibrosis can be divided into drug therapy and non-drug treatment (lung transplantation, oxygen therapy, respiratory rehabilitation treatment), complication treatment (acute exacerbation, pulmonary hypertension, gastroesophageal reflux disease) As the etiology of this disease is not known exactly, the exact timing of the treatment was not known. More than half of the patients are known to have died within 3 to 5 years due to persistent disease worsening. There is a specificity that does not improve any treatment after the disease progresses and is completely fibrotic.

Many drugs have been tried in the treatment of patients with pulmonary fibrosis, and some drugs have shown promising results, but most of them were found to lack sufficient evidence to demonstrate the efficacy of the treatment or to have a weak effect. According to the International Consolidated Report, some patients may consider the use of N-acetylcysteine alone or combination therapy with steroid / azathioprine, anticoagulant, and pirfenidone, It is not a cure. Clinical studies of pyrennidone have shown that the treatment group significantly slows down the rate of spontaneous abortion and disease progression, and has been approved as a treatment for mild and moderate patients in Japan and Europe, but its efficacy is still questionable. to be.

Beta-lapachone can be obtained from the South American native laphacho tree (Tabebuia avellanedae), while dunnione and alpha-dunnione are also found in the South American native Streptococcus It is obtained from leaves of Streptocarpus dunnii. These natural tricyclic naphthoquinone derivatives have been widely used in Latin America for a long time as an anticancer drug and as a medicine for treating Chagas disease, a typical endemic disease in South America, . Particularly, their pharmacological activity as an anticancer agent began to be noticed in the western world, and as disclosed in US Pat. No. 5,969,163, these tricyclic naphthoquinone derivatives were actually developed as various anti-cancer drugs by various research groups have.

As prior art, Antje Moeller et al. Have shown that the use of bleomycin animal models for the treatment of idiopathic pulmonary fibrosis is appropriate and that 232 drugs have been studied in the animal model (The International Journal of Biochemistry & Cell Biology 40 Korean Patent Laid-Open No. 10-2007-0136105 discloses the synthesis of a naphtoquinone compound, which prevents degeneration of RGC axons forming retinal ganglion cells (RGC) and optic nerve cells , And RGC and axon recovery, it is possible to effectively use the composition as a pharmaceutical composition for the treatment and prevention of glaucoma. However, the present invention provides a pharmaceutical composition for prevention and treatment of idiopathic pulmonary fibrosis Have not been known to date.

Thus, the inventors of the present invention have developed a mouse animal model having idiopathic pulmonary fibrosis using a method of directly injecting bleomycin into the bronchial tree, and after confirming the changes in the administration of beta-raffin or dunion, The expression of IL-6, TGF-β1 and αSMA increased by bleomycin was decreased, and the expression of IL-6, TGF-β1 and αSMA was decreased by the infiltration of various inflammatory cells of the lung tissue, accumulation of extracellular matrix, The survival rate was increased in combination with dithion after the administration of micin and by confirming a significant increase in the ratio of NAD ⁺ / NADH, beta-lapachone or dunion could be used to treat or prevent idiopathic pulmonary fibrosis The present invention has been completed.

It is an object of the present invention to provide a pharmaceutical composition containing beta-lapachone or dunnione, or a pharmaceutically acceptable salt thereof, a prodrug thereof, a solvate thereof or an isomer thereof, And to provide a pharmaceutical composition for the treatment or prevention of idiopathic pulmonary fibrosis.

In order to accomplish the above object, the present invention provides a pharmaceutical composition comprising a compound represented by the following formula (1), a pharmaceutically acceptable salt thereof, a prodrug thereof, a solvate thereof, or an isomer thereof, A pharmaceutical composition for treating or preventing:

[Chemical Formula 1]

:

In this formula,

R1 and R2 are each independently hydrogen, halogen, alkoxy, hydroxy or lower alkyl of 1 to 6 carbon atoms;

R 3, R 4, R 5, R 6, R 7 and R 8 are each independently hydrogen, hydroxy, alkyl of 1 to 20 carbon atoms, alkene or alkoxy, cycloalkyl, heterocycloalkyl, aryl or heteroaryl, May form an annular structure by mutual bonding;

n is 0 or 1, and when n is 0, adjacent carbon atoms thereof form a cyclic structure by a direct bond.

In addition, the present invention relates to a pharmaceutical composition for preventing or ameliorating interstitial lung disease containing the compound represented by the above-mentioned formula (1), a pharmaceutically acceptable salt thereof, a prodrug thereof, a solvate thereof or an isomer thereof as an active ingredient .

In the present invention, the effect of administration of beta-raffin or dunion was confirmed after the preparation of a mouse animal model with idiopathic pulmonary fibrosis using bleomycin. As a result, it was found that various inflammatory cells of pulmonary tissue Tissue density and degree of fibrosis due to invasion and accumulation of extracellular matrix decreased, and expression of IL-6, TGF-β1 and αSMA increased by bleomycin was decreased. In combination with duomin after bleomycin administration , And by confirming a significant increase in the ratio of NAD ⁺ / NADH, beta-raffin or dunion can be usefully used in the treatment or prevention of idiopathic pulmonary fibrosis.

Brief Description of the Drawings Figure 1 is a diagram showing the histological changes of bleomycin-induced lung injury and the effects of beta-lapachone (A) and dunion (B)
Cont: normal group; BLM: Bleomycin; BLM + β-Lapachone: combination treatment of bleomycin with 40 mg / kg of beta-lapachone; β-Lapachone: a group treated with beta-Lapachone 40 mg / kg alone; BLM + Dunnione (20 mg / kg): group treated with bleomycin and dunion 20 mg / kg; BLM + Dunnione (40 mg / kg): group treated with bleomycin and dunion 40 mg / kg; BLM + Dunnione (80 mg / kg): a combination of bleomycin and Dynion 80 mg / kg; Dunnione (80 mg / kg): treated with Dynion 80 mg / kg alone.
Figure 2 shows the histological results of lungs treated with bleomycin, Beta-Lapachone (A) and Dynion (B) in a high power field (100x)
Cont: normal group; BLM: Bleomycin; BLM + β-Lapachone: combination treatment of bleomycin with 40 mg / kg of beta-lapachone; β-Lapachone: a group treated with beta-Lapachone 40 mg / kg alone; BLM + Dunnione (20 mg / kg): group treated with bleomycin and dunion 20 mg / kg; BLM + Dunnione (40 mg / kg): group treated with bleomycin and dunion 40 mg / kg; BLM + Dunnione (80 mg / kg): a combination of bleomycin and Dynion 80 mg / kg; Dunnione (80 mg / kg): treated with Dynion 80 mg / kg alone.
Figure 3 shows the results of an analysis of the effects of bleomycin-induced pulmonary fibrosis and beta-lapachone (A) and duoni (B)
Cont: normal group; BLM: Bleomycin; BLM + β-Lapachone: combination treatment of bleomycin with 40 mg / kg of beta-lapachone; β-Lapachone: a group treated with beta-Lapachone 40 mg / kg alone; BLM + Dunnione (20 mg / kg): group treated with bleomycin and dunion 20 mg / kg; BLM + Dunnione (40 mg / kg): group treated with bleomycin and dunion 40 mg / kg; BLM + Dunnione (80 mg / kg): a combination of bleomycin and Dynion 80 mg / kg; Dunnione (80 mg / kg): treated with Dynion 80 mg / kg alone.
Figure 4 compares quantitative analysis of the effect of bleomycin-induced pulmonary fibrosis and duiomy:
* p <0.05: comparison between normal and bleomycin groups and ** p <0.05: comparison between bleomycin and dunion combination groups.
Figure 5 shows the effect of IL-6 (interleukin-6) expression and dynion on bleomycin:
Cont: normal group; BLM: Bleomycin; BLM + Dunnione (20 mg / kg): group treated with bleomycin and dunion 20 mg / kg; BLM + Dunnione (40 mg / kg): group treated with bleomycin and dunion 40 mg / kg; BLM + Dunnione (80 mg / kg): a combination of bleomycin and Dynion 80 mg / kg; Dunnione (80 mg / kg): treated with Dynion 80 mg / kg alone.
6 is a graph showing the expression of TNF-alpha (tumor necrosis factor-alpha) and IL-1 beta (interleukin-1 beta) and the effect of dyneon on bleomycin:
Cont: normal group; BLM: Bleomycin; BLM + Dunnione (20 mg / kg): group treated with bleomycin and dunion 20 mg / kg; BLM + Dunnione (40 mg / kg): group treated with bleomycin and dunion 40 mg / kg; BLM + Dunnione (80 mg / kg): a combination of bleomycin and Dynion 80 mg / kg; Dunnione (80 mg / kg): treated with Dynion 80 mg / kg alone.
FIG. 7 shows the expression of TGF-beta1 (transforming growth factor-beta 1) and the effects of beta-lapachone (A) and dunion (B) on bleomycin:
Cont: normal group; BLM: Bleomycin; BLM + β-Lapachone: combination treatment of bleomycin with 40 mg / kg of beta-lapachone; β-Lapachone: a group treated with beta-Lapachone 40 mg / kg alone; BLM + Dunnione (20 mg / kg): group treated with bleomycin and dunion 20 mg / kg; BLM + Dunnione (40 mg / kg): group treated with bleomycin and dunion 40 mg / kg; BLM + Dunnione (80 mg / kg): a combination of bleomycin and Dynion 80 mg / kg; Dunnione (80 mg / kg): treated with Dynion 80 mg / kg alone.
8 is a graph showing the effect of α-smooth muscle actin (αSMA) expression and dyneon on bleomycin:
Cont: normal group; BLM: Bleomycin; BLM + Dunnione (20 mg / kg): group treated with bleomycin and dunion 20 mg / kg; BLM + Dunnione (40 mg / kg): group treated with bleomycin and dunion 40 mg / kg; BLM + Dunnione (80 mg / kg): a combination of bleomycin and Dynion 80 mg / kg; Dunnione (80 mg / kg): treated with Dynion 80 mg / kg alone.
9 is a diagram showing mouse survival rate by bleomycin and dunion treatment:
Cont: normal group; BLM: Bleomycin; BLM + Dunnione (10 mg / kg): group treated with bleomycin and 10 mg / kg dunion; BLM + Dunnione (20 mg / kg): group treated with bleomycin and dunion 20 mg / kg; BLM + Dunnione (40 mg / kg): group treated with bleomycin and dunion 40 mg / kg; BLM + Dunnione (80 mg / kg): a combination of bleomycin and Dynion 80 mg / kg; Dunnione (80 mg / kg): treated with Dynion 80 mg / kg alone.
10 shows the effect of bleomycin and beta-lapachone (A) and duithon (B) on the ratio of NAD ⁺ / NADH in lung tissue:
* p <0.05: comparison between normal and bleomycin groups and ** p <0.05 compared with the combination treatment of bleomycin and beta-lafacin or dunion.

Hereinafter, the present invention will be described in detail.

[Chemical Formula 1]

:

In this formula,

The compound of formula (1) is preferably selected from compounds of the following formulas (2) and (3)

(2)

;

(3)

:

Wherein R 1, R 2, R 3, R 4, R 5, R 6, R 7 and R 8 are the same as defined in Formula (1).

Preferably, R1 and R2 are each hydrogen, and the compound of Formula 2 is preferably a compound of Formula 2a wherein R1, R2 and R4 are each hydrogen, or a compound of Formula 2a below wherein R1, R2 and R6 are each hydrogen :

(2a)

;

(2b)

.

The compound of Formula 3 is preferably a compound of Formula 3a wherein R1, R2, R5, R6, R7 and R8 are each hydrogen.

[Chemical Formula 3]

.

The compound of formula (1) is most preferably a beta-lapachone or dunnione represented by the following formula (4) or (5)

[Chemical Formula 4]

;

[Chemical Formula 5]

.

In addition, the interstitial lung disease is preferably idiopathic pulmonary fibrosis.

It is preferable that the composition has the activity of infiltration of inflammatory cells in the lung tissue, densification and fibrosis of tissues due to accumulation of extracellular matrix, decrease of expression of IL-6, TGF-beta 1 and alpha SMA.

The cause and pathogenesis of pulmonary fibrosis have not yet been elucidated and various mechanisms have been suggested. One well-known mechanism is repeated inflammatory reaction and accumulation of extracellular matrix increases fibrosis. The inflammatory cytokines (TNF-α, IL-6, interleukin-6, IL-1β, interleukin-1β) TGF-β1 (transforming growth factor-β1) is a well-known factor. In addition, collagens and α-smooth muscle action (αSMA) are the major molecules known to be major markers of fibrosis. Thus, by analyzing the changes in the expression of the above factors, we can confirm the pathway for controlling bleomycin pulmonary fibrosis in Beta-Lapachon and Duunion.

In addition, NAD ⁺ and NADH are known to be important mediators for energy metabolism and cell homeostasis. NAD ⁺ is a cofactor for various enzymes such as sirtuins and the like, thereby regulating the activity of NAD ⁺ Or to treat the disease. Recently, the importance of the NQO1 pathway has been elucidated in pathways that increase the intracellular NAD ⁺ concentration (de novo, salvage and NQO1 (NAD (P) H: quinone oxidoreductase-1) pathway.

In a specific example of the present invention, after the mouse animal model with idiopathic pulmonary fibrosis was produced using bleomycin, the effect of administration of beta-raffin or dunion was confirmed, and as a result, (Fig. 3 and Fig. 4), and decreased expression of IL-6, TGF- [beta] l and [alpha] SMA increased by bleomycin (See FIG. 5 to FIG. 8), survival rates were increased (see FIG. 9), and a significant increase in the ratio of NAD ⁺ / NADH was observed when dyunion was coadministered after bleomycin administration (see FIG. 10) .

Thus, beta-lapachone or dunion may be usefully employed as a pharmaceutical composition for the treatment or prevention of idiopathic pulmonary fibrosis.

The present invention includes not only the compounds represented by the formula (1), but also pharmaceutically acceptable salts thereof, possible solvates, hydrates, racemates or stereoisomers thereof which can be prepared therefrom.

The present invention can be used in the form of a compound represented by the formula (1) or a pharmaceutically acceptable salt thereof, and as the salt, an acid addition salt formed by a pharmaceutically acceptable free acid is useful. Acid addition salts include those derived from inorganic acids such as hydrochloric acid, nitric acid, phosphoric acid, sulfuric acid, hydrobromic acid, hydroiodic acid, nitrous acid or phosphorous acid, and aliphatic mono- and dicarboxylates, phenyl-substituted alkanoates, hydroxyalkanoates, Derived from non-toxic organic acids, such as, for example, diesters, aromatic acids, aliphatic and aromatic sulfonic acids. Such pharmaceutically innocuous salts include, but are not limited to, sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, nitrate, phosphate, monohydrogenphosphate, dihydrogenphosphate, metaphosphate, pyrophosphate chloride, bromide, Butyrate, caprate, heptanoate, propiolate, oxalate, malonate, succinate, succinic acid, succinic acid, succinic acid, succinic acid, , Sebacate, fumarate, maleate, butyne-1,4-dioate, hexane-1,6-dioate, benzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate, hydroxybenzoate, Methoxybenzoate, phthalate, terephthalate, benzenesulfonate, toluene sulfonate, chlorobenzene sulfoxide Sulfonate, methanesulfonate, propanesulfonate, naphthalene-1-sulphonate, naphthalene-1-sulphonate, , Naphthalene-2-sulfonate or mandelate.

The acid addition salt according to the present invention can be produced by a conventional method, for example, by dissolving the compound represented by the formula (1) in an excess amount of an acid aqueous solution, and adding the salt to a water-miscible organic solvent such as methanol, ethanol, acetone or acetonitrile To precipitate it. It is also possible to prepare the same by heating the compound represented by the general formula (1) and an acid aqueous solution or alcohol, followed by evaporating the mixture or drying the precipitated salt by suction filtration.

In addition, bases can be used to make pharmaceutically acceptable metal salts. The alkali metal or alkaline earth metal salt is obtained, for example, by dissolving the compound in an excess amount of an alkali metal hydroxide or alkaline earth metal hydroxide solution, filtering the insoluble compound salt, and evaporating and drying the filtrate. At this time, it is preferable for the metal salt to produce sodium, potassium or calcium salt. The corresponding silver salt is also obtained by reacting an alkali metal or alkaline earth metal salt with a suitable salt (such as silver nitrate).

The pharmaceutical composition of the present invention may further contain commonly used excipients, disintegrants, sweeteners, lubricants, flavors and the like, and may be formulated into tablets, capsules, powders, granules, suspensions, Syrups, and other liquid preparations.

Specifically, the pharmaceutical compositions of the present invention may be formulated for oral administration, for example, tablets, troches, lozenges, aqueous or aqueous suspensions, prepared powders or granules, emulsions, hard or soft capsules, It is formulated into elixirs. Binders such as lactose, saccharose, sorbitol, mannitol, starch, amylopectin, cellulose or gelatin, excipients such as dicalcium phosphate, disintegrants such as corn starch or sweet potato starch, magnesium stearate, Lubricating oil such as calcium stearate, sodium stearyl fumarate or polyethylene glycol wax. In the case of a capsule formulation, in addition to the above-mentioned substances, a liquid carrier such as fatty oil is contained.

In addition, the pharmaceutical composition of the present invention can be administered orally or parenterally, and it is preferable to select subcutaneous injection, intravenous injection, intramuscular injection, or intra-thoracic injection injection method for parenteral administration. In order to formulate the composition for parenteral administration, the mixed extract of the present invention is mixed with a stabilizer or a buffer in water to prepare a suspension, which is formulated into a unit dosage form of ampoule or vial, , Or in combination with methods using surgery, radiation therapy, hormone therapy, chemotherapy, and biological response modifiers.

The dosage of the active ingredient according to the present invention is appropriately selected depending on the degree of absorption, inactivation rate and rate of absorption of the active ingredient in the body, age, sex and condition of the patient, and severity of the disease to be treated.

The present invention also relates to a pharmaceutical composition for preventing or ameliorating interstitial lung disease comprising a compound represented by the following formula 1, a pharmaceutically acceptable salt thereof, a prodrug thereof, a solvate thereof or an isomer thereof, Lt; / RTI >

[Chemical Formula 1]

:

In this formula,

[Chemical Formula 4]

;

[Chemical Formula 5]

.

In the present invention, beta-lapachone or dunion has the effect of infiltration of inflammatory cells and accumulation of extracellular matrix in the lung tissue of animal models with idiopathic pulmonary fibrosis, resulting in densification of tissue and fibrosis, , It can be used as a health functional food for prevention or improvement of interstitial lung disease.

As used herein, the term " health functional food "refers to foods prepared and processed in the form of tablets, capsules, powders, granules, liquids, and circles by using raw materials and components having useful functions in the human body. Here, the term "functionality" means that the structure and function of the human body are controlled to obtain nutritional effects or effects useful for health use such as physiological actions. The health functional food of the present invention can be prepared by a method commonly used in the art and can be prepared by adding raw materials and ingredients that are conventionally added in the art.

The amount of the active ingredient to be mixed can be suitably determined according to the intended use (prevention, health or therapeutic treatment). Generally, the compound of the formula (1) of the present invention is added in an amount of 1 to 10% by weight, preferably 5 to 10% by weight of the raw material composition in the production of food. However, in the case of long-term ingestion intended for health and hygiene purposes or for the purpose of controlling health, the amount can also be used in the above-mentioned range.

The health functional food of the present invention may contain various flavoring agents or natural carbohydrates as an additional ingredient like ordinary foods. The natural carbohydrates are sugar saccharides such as monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, polysaccharides such as dextrin and cyclodextrin, xylitol, sorbitol and erythritol. Examples of sweeteners include natural sweeteners such as tau martin and stevia extract, synthetic sweeteners such as saccharin and aspartame, and the like.

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

However, the following Examples and Experimental Examples are merely illustrative of the present invention, and the content of the present invention is not limited by the following Examples and Experimental Examples.

< Example 1> Preparation of an idiopathic pulmonary fibrosis animal model

8 weeks old C57BL / 6 rats were used in the present invention. All mice were housed in an aseptic animal room at constant temperature (22 ~ 26 ℃) and humidity (55 ~ 60%). Adult animals were fed with normal solid diet (Samtaco, Korea) and water for one week Respectively. All experiments were conducted after approval of the clinical trial management committee according to the laboratory animal management and ethics regulations of Wonkwang University clinical trial management committee. Induction of pulmonary fibrosis in mice is a relatively widely used method in which bleomycin is directly sprayed onto the bronchi and then maintained for 2 to 3 weeks. The experimental group was orally administered daily with PBS (control, 10), bleomycin (2 mg / kg / day) one day prior to bleomycin (BLM, 10 mice) (10 mg / kg, 20 mg / kg, 40 mg / kg, and 80 mg / kg) in the combination treatment with β-lapachone (40 mg / kg, 5 mice) or dunnione (40 mg / kg, 5 doses) or dunion (80 mg / kg, 10 doses) were administered to each group.

< Experimental Example 1> Beta- Lapa Village Dyneon's Bleomycin Confirming the effect of inducing pulmonary fibrosis

<1-1> Duneion Of pulmonary histology

In pulmonary fibrosis, various inflammatory cells infiltrate into pulmonary tissue due to inflammatory reaction, induce repeated inflammatory reaction, and manifest fibrosis with various expression of extracellular matrix.

Specifically, the lung tissue was separated from the animal model prepared by the method of Example 1, fixed in 10% formalin for 24 hours at 4 ° C, washed and dehydrated, Embedded in paraffin and cut to a thickness of 5 탆. The cut tissue was stained with hematoxylin-eosin and confirmed by optical microscope.

As a result, as shown in Fig. 1, when the entire cross sections of the left lung were compared, the pulmonary tissues of the bleomycin group showed various infiltration of inflammatory cells and alveolar tissue accumulation in the alveolar tissues And the concentration of the above-mentioned phenomenon was reduced in a concentration-dependent manner in the group treated with the combination of Beta-Lafacin (A) or Dynion (B) (FIG. 1). Further, as shown in Fig. 2, when observed in detail at a high magnification of a microscope, the tissue changes caused by bleomycin were significantly controlled in the group treated with Beta-Lapachone (A) or Duchenon (B) (Fig. 2).

<1-2> Beta- Lapa Village Duneion Of pulmonary fibrosis

To confirm the regulation of pulmonary fibrosis by beta-lapachone or dunion, the following experiment was performed. In pulmonary fibrosis, accumulation of various extracellular substances such as collagen is increased and the degree of fibrosis is evaluated by analyzing such substances by methods such as sirius red, masson's trichrome and hydroxyproline assay.

Specifically, the lung tissue was separated from the animal model prepared by the method of Example 1, fixed in 10% formalin for 24 hours at 4 ° C, washed, washed and dehydrated, and then embedded in paraffin And cut into a thickness of 5 탆. The cut tissues were stained with Sirius red and confirmed by optical microscope.

As a result, as shown in Fig. 3, when the entire cross sections of the left lung lobe were compared, in the lung tissue of the bleomycin group, strong red staining was observed around the alveolar tissue in comparison with the normal group, Was reduced in a concentration-dependent manner in the group treated with Beta-Lapachone (A) or Duchenon (B) (FIG. 3).

In addition, for quantitative comparison of pulmonary fibrosis, hydroxyproline analysis was performed from lung tissue obtained from each group. Hydroxyproline is a nonproteinogenic amino acid and is known to exist only in substrates such as collagen and elastin, and is a useful target for quantitative analysis of fibrosis. Specifically, lung tissue (10 mg) was homogenized in 100 μL of distilled water, and 100 μL of concentrated hydrochloric acid solution was added thereto, followed by reaction at 120 ° C. for 3 hours. Thereafter, the obtained reaction solution was mixed well and decanted for 5 minutes at a speed of 10,000 × g, and the supernatant was used for analysis.

As a result, as shown in FIG. 4, the amount of hydroxyproline was significantly increased in the bleomycin group as compared with that in the normal group according to the fibrosis, and a significant decrease was observed when the combination of Dynion 80 mg / kg I could confirm. On the other hand, the dunion alone treatment group showed no significant difference with the normal group (Fig. 4).

< Experimental Example 2> Beta- Lapa Village Dyneon's Bleomycin Identification of the pathway for induction

In order to confirm the regulatory pathway of the bleomycin-induced pulmonary fibrosis of Beta-Lapachon and Dunion, an analysis of expression changes of markers related to inflammatory response and fibrosis changes was performed.

Specifically, the lung tissue was isolated from the animal model prepared in Example 1, fixed in 10% formalin for 24 hours at 4 ° C, washed and rinsed, and embedded in paraffin to obtain 5 Mu] m. Subsequently, the excised tissue was subjected to immunohistochemical analysis using the corresponding antibody to confirm the expression of the target proteins.

As a result, as shown in Fig. 5, the expression of IL-6 (Santa Cruz Biotechnology, CA, USA), one of the inflammatory cytokines, was significantly increased in the bleomycin group as compared with the normal group, And decreased in a concentration-dependent manner in one group (FIG. 5). On the other hand, as shown in FIG. 6, significant expression changes of TNF-? And IL-1? (Santa Cruz Biotechnology, CA, USA) could not be confirmed (FIG. 6). The cause of this phenomenon is thought to be the early change of expression immediately after the reaction due to the kinetic difference of the expression depending on the target factors of the inflammatory reaction.

The TGF-β1 signal transduction pathway is known to be a representative mechanism of fibrosis change. To confirm the effect of bleomycin-induced fibrosis and beta-lymphocyte and dunion, TGF- Immunohistochemical analysis was performed using anti-β1 antibody (Abcam, MA, USA).

As a result, as shown in FIG. 7, significant expression of TGF-β1 was confirmed in the bleomycin group, and the change was dose-dependently decreased by the treatment with Beta-Lafacin (A) or Dynion (B) (Fig. 7).

On the other hand, αSMA is known to be one of the main products of fibrosis. As shown in FIG. 8, an increase in expression of αSMA (Abcam, MA, USA) was observed in the bleomycin group, (Fig. 8).

< Experimental Example 3> Bleomycin Survival rate of mice with induced pulmonary fibrosis Dyneon's effect

Pulmonary fibrosis is an irreversible change. It is known that there is no clear remedy until now and it is a poor prognosis. Therefore, the effect of dunion on the survival rate of mice with bleomycin-induced pulmonary fibrosis in the animal model of the present invention was confirmed.

Specifically, mouse survival rate over time was measured using an animal model prepared by the method of Example 1, and it is shown in FIG. As a result, the survival rate of mice after bleomycin administration was drastically decreased after administration of bleomycin, and the survival rate was less than 40% for 2 weeks. In the case of combination treatment with dunion, the survival rate was more than twice as high as that of bleomycin (Fig. 9).

< Experimental Example 4> Beta- Lapa Village Dyneon's NAD ⁺ / NADH Confirm adjustment

In order to confirm the NAD ⁺ / NADH modulating effect of beta-lapachone and dunion, the amounts of NAD ⁺ and NADH were measured and analyzed for the combination of bleomycin, beta-lapachone and dunion.

Specifically, 10 mg of lung tissue obtained from the animal model prepared by the method of Example 1 was homogenized by adding 100 μL of NAD ⁺ or NADH extraction solution, and then heat-treated at 60 ° C for 5 minutes. Then, the reaction solution and the extraction solution were added, decanted for 5 minutes at 14,000 rpm, and the supernatant was used for NAD ⁺ / NADH analysis.

As a result, as shown in Fig. 10, the NAD + / NADH ratio was significantly decreased in the bleomycin group compared with the normal group, and in the beta-lapachin (A) or duinon (B) (Fig. 10). Therefore, beta-lapachone and dunion increased the NQO1 enzyme activity and increased the intracellular NAD ⁺ / NADH ratio and increased enzymatic activities such as sirt1 and sirt3, using various enzymes using NAD ⁺ as a cofactor, , Which is one of the posttranslational modification (acetylation of acetylation of NF-κB p65, p53) of several target molecules, is effective in inhibiting and controlling inflammation and fibrosis.

Claims

A pharmaceutical composition for the treatment or prevention of interstitial lung disease comprising a compound represented by the following formula (1), a pharmaceutically acceptable salt thereof, a prodrug thereof, a solvate thereof or an isomer thereof as an active ingredient:
[Chemical Formula 1]

:
In this formula,
R1 and R2 are each independently hydrogen, halogen, alkoxy, hydroxy or lower alkyl of 1 to 6 carbon atoms;
R 3, R 4, R 5, R 6, R 7 and R 8 are each independently hydrogen, hydroxy, alkyl of 1 to 20 carbon atoms, alkene or alkoxy, cycloalkyl, heterocycloalkyl, aryl or heteroaryl, May form an annular structure by mutual bonding;
n is 0 or 1, and when n is 0, adjacent carbon atoms thereof form a cyclic structure by a direct bond.

The pharmaceutical composition for treating or preventing interstitial lung disease according to claim 1, wherein the compound of formula (1) is selected from the following compounds (2) and (3)
(2)

;
(3)

:
Wherein R 1, R 2, R 3, R 4, R 5, R 6, R 7 and R 8 are the same as defined in Formula (1).

2. The pharmaceutical composition for treating or preventing interstitial lung disease according to claim 1, wherein R1 and R2 are each hydrogen.

The compound of claim 2, wherein the compound of formula 2 is a compound of formula 2a wherein R1, R2 and R4 are each hydrogen, or a compound of formula 2a wherein R1, R2 and R6 are each hydrogen. A pharmaceutical composition for treating or preventing:
(2a)

;
(2b)

.

A pharmaceutical composition for the treatment or prevention of interstitial lung disease according to claim 2, wherein said compound of formula (3) is a compound of formula (3a) wherein R1, R2, R5, R6, R7 and R8 are each hydrogen. :
[Chemical Formula 3]

.

The pharmaceutical composition for treating or preventing interstitial lung disease according to claim 1, wherein the compound of formula (1) is a compound of formula (4) or (5)
[Chemical Formula 4]

;
[Chemical Formula 5]

.

The pharmaceutical composition for treating or preventing interstitial lung disease according to claim 1, wherein the interstitial lung disease is idiopathic pulmonary fibrosis.

The composition according to claim 1, wherein the composition has activity of infiltrating inflammatory cells in the lung tissue, decreasing tissue density and fibrosis due to accumulation of extracellular matrix, and decreasing expression of IL-6, TGF-beta 1 and alpha SMA A pharmaceutical composition for the treatment or prevention of sexual pulmonary disease.

A health functional food for preventing or ameliorating an interstitial lung disease comprising a compound represented by the following formula (1), a pharmaceutically acceptable salt thereof, a prodrug thereof, a solvate thereof or an isomer thereof:
[Chemical Formula 1]

10. The health functional food for preventing or improving interstitial lung disease according to claim 9, wherein the interstitial lung disease is idiopathic pulmonary fibrosis.