阅读说明：本技术 用于预防或治疗肺纤维化的药剂 (Agent for preventing or treating pulmonary fibrosis ) 是由 A·阿扎拉 吉川公平 于 2020-02-04 设计创作，主要内容包括：用于预防或治疗肺纤维化的药剂,其含有作为活性成分的由下式(I)表示的化合物或其药学上可接受的盐。(An agent for preventing or treating pulmonary fibrosis, which contains a compound represented by the following formula (I) as an active ingredient Or a pharmaceutically acceptable salt thereof.)

1. An agent for preventing or treating pulmonary fibrosis, comprising:

a compound of formula (I)

Or a pharmaceutically acceptable salt thereof.

2. The medicament according to claim 1, wherein the pulmonary fibrosis is an interstitial lung disease with fibrosis.

3. The medicament according to claim 2, wherein the interstitial lung disease with fibrosis is a disease selected from idiopathic pulmonary fibrosis, disease-induced pulmonary fibrosis and other factor-induced pulmonary fibrosis.

4. A method for preventing or treating pulmonary fibrosis, comprising:

administering to a patient in need thereof a prophylactically or therapeutically effective amount of a compound of formula (I)

Or a pharmaceutically acceptable salt thereof.

5. A compound of formula (I), or a pharmaceutically acceptable salt thereof, for use in the prevention or treatment of pulmonary fibrosis,

6. use of a compound of formula (I), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for the prevention or treatment of pulmonary fibrosis,

Technical Field

The present invention relates to an agent for preventing or treating pulmonary fibrosis, and a method for preventing or treating pulmonary fibrosis, and is useful in the medical field.

Background

In international publication No. 2007/089034, a 1, 4-benzoxazine compound is described, and compound (I) is described as an MR antagonist. Further, in international publication No. 2018/062134, compound (I) is described for use in the treatment of non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), and the like. The entire contents of this disclosure are incorporated herein by reference.

Disclosure of Invention

One aspect of the present invention is an agent for preventing or treating pulmonary fibrosis, which contains, as an active ingredient, a compound represented by the following formula (I) (which may be referred to as compound (I) hereinafter)

Or a pharmaceutically acceptable salt thereof.

The agent can be used for preventing or treating pulmonary fibrosis, wherein the pulmonary fibrosis is an interstitial lung disease with fibrosis.

The agent can be used for preventing or treating pulmonary fibrosis, wherein the interstitial lung disease accompanied with fibrosis is a disease selected from idiopathic pulmonary fibrosis, disease-induced pulmonary fibrosis and other factor-induced pulmonary fibrosis.

Another aspect of the present invention is a pharmaceutical composition for preventing or treating pulmonary fibrosis, which contains a compound represented by the following formula (I) as an active ingredient

Or a pharmaceutically acceptable salt thereof, together with a pharmaceutically acceptable carrier.

The pharmaceutical composition may be used for the prevention or treatment of pulmonary fibrosis, wherein the pulmonary fibrosis is an interstitial lung disease with fibrosis.

The pharmaceutical composition may be used for the prevention or treatment of pulmonary fibrosis, wherein the interstitial lung disease with fibrosis is a disease selected from idiopathic pulmonary fibrosis, disease-induced pulmonary fibrosis and other factor-induced pulmonary fibrosis.

Still another aspect of the present invention is a method for preventing or treating pulmonary fibrosis, comprising administering to a patient in need thereof a prophylactically effective amount or a therapeutically effective amount of a compound represented by the following formula (I)

Or a pharmaceutically acceptable salt thereof.

The method may be used for the prevention or treatment of pulmonary fibrosis, wherein the pulmonary fibrosis is an interstitial lung disease with fibrosis.

The method may be used for the prevention or treatment of pulmonary fibrosis, wherein the interstitial lung disease with fibrosis is a disease selected from idiopathic pulmonary fibrosis, disease-induced pulmonary fibrosis and other factor-induced pulmonary fibrosis.

Another aspect of the present invention is a compound represented by the following formula (I)

Or a pharmaceutically acceptable salt thereof, for use in the prevention or treatment of pulmonary fibrosis.

The compound or a pharmaceutically acceptable salt thereof may be used for the prevention or treatment of pulmonary fibrosis, wherein the pulmonary fibrosis is an interstitial lung disease with fibrosis.

The compound or a pharmaceutically acceptable salt thereof can be used for preventing or treating pulmonary fibrosis, wherein the interstitial lung disease accompanied with fibrosis is a disease selected from idiopathic pulmonary fibrosis, disease-induced pulmonary fibrosis and other factor-induced pulmonary fibrosis.

Another aspect of the present invention is a compound represented by the following formula (I)

Or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for the prevention or treatment of pulmonary fibrosis.

The use of the compound or a pharmaceutically acceptable salt thereof, wherein the pulmonary fibrosis is an interstitial lung disease with fibrosis.

The use of the compound or a pharmaceutically acceptable salt thereof, wherein the interstitial lung disease with fibrosis is a disease selected from idiopathic pulmonary fibrosis, disease-induced pulmonary fibrosis and other factor-induced pulmonary fibrosis.

Drawings

A more complete appreciation of the invention and many of the attendant advantages thereof will be readily obtained as the same becomes better understood by reference to the following detailed description when considered in connection with the accompanying drawings, wherein:

FIG. 1 shows the results of evaluation of the amount of deep air intake and elasticity performed in example 1;

figure 2 shows the results of the lung compliance and resistance evaluations performed in example 1;

FIG. 3 shows the hydroxyproline content and positive rate evaluation results in the sirius red staining performed in example 1;

FIG. 4 shows the results of evaluation of collagen gene expression performed in example 1;

FIG. 5 shows the results of the assessment of α SMA, fibronectin and vimentin gene expression performed in example 1;

FIG. 6 shows the results of the evaluation of CTGF, PAI-1 and Ccl2 gene expression performed in example 1;

FIG. 7 shows the results of evaluation of TGF-beta gene expression performed in example 1;

FIG. 8 shows the results of the evaluation of the concentration of Compound (I) in plasma carried out in example 1;

FIG. 9 shows the results of evaluation of the amount of deep suction and the resistance performed in example 2;

FIG. 10 shows the results of the elasticity and compliance evaluation performed in example 2;

FIG. 11 shows the results of evaluation of hydroxyproline content and positive rate in sirius red staining performed in example 2; and

fig. 12 shows the results of the evaluation of the concentration of compound (I) in plasma performed in example 2.

Detailed Description

Embodiments will now be described with reference to the drawings, wherein like reference numerals designate corresponding or equivalent elements in the various drawings.

In the present specification, an example of "pulmonary fibrosis" which is a target disease is "interstitial lung disease accompanied by fibrosis". Further, in the present specification, all "fibers" generated in vivo are denoted as "fibers". Examples of "interstitial lung disease with fibrosis" include "Idiopathic Pulmonary Fibrosis (IPF)", "disease-induced pulmonary fibrosis", and "other-factor-induced pulmonary fibrosis". Here, the term "idiopathic pulmonary fibrosis" refers to pulmonary fibrosis for which the cause cannot be identified. The term "disease-induced pulmonary fibrosis" refers to pulmonary fibrosis that occurs in conjunction with a disease in pulmonary fibrosis. For example, "disease-induced pulmonary fibrosis" is pulmonary fibrosis in which fibrosis has developed from: allergic pneumonia (HP), rheumatoid arthritis-associated interstitial lung disease (RA-ILD), systemic scleroderma-associated interstitial lung disease (SSc-ILD), polymyositis/dermatomyositis-associated interstitial lung disease (PM/DM-ILD), Sjogren's ILD, systemic lupus erythematosus-associated interstitial lung disease (SLE-ILD), mixed connective tissue disease-associated interstitial lung disease (MCTD-ILD), collagen disease-associated interstitial lung disease (CTD-ILD), pulmonary sarcoidosis, and the like.

"other-factor-induced pulmonary fibrosis" is, for example, pulmonary fibrosis that is a pulmonary disease caused by idiopathic nonspecific interstitial pneumonia (iNSIP), exposure to inorganic substances, exposure to organic substances, drugs or smoking, or the like, and in which fibrosis has been developed.

In the present specification, the term "treatment" includes cure of a disease (all pathological conditions or one or more pathological conditions), amelioration of a disease, and inhibition of progression of disease severity. The term "therapeutically effective amount" refers to a dose of compound (I) sufficient to achieve this purpose.

The therapeutic agents of the present specification may also be used as prophylactic agents. The term "prevention" includes both prevention of the development of a disease (all pathological conditions or one or more pathological conditions) and delay in the development of a disease. The term "prophylactically effective amount" refers to a dose of compound (I) sufficient to achieve this purpose.

Compound (I)

The compound (I) of the present invention is a compound described in International publication No. 2007/089034 (example 9). One skilled in the art can use the methods described in this disclosure or methods consistent therewith to produce compound (I).

In practicing this embodiment, compound (I) may be used in free form or in the form of a pharmaceutically acceptable salt thereof.

In the present specification, examples of such pharmaceutically acceptable salts of compound (I) include salts with acids, such as salts with inorganic acids, for example, hydrochloride, hydrobromide, sulfate and phosphate, and salts with organic acids, such as acetate, fumarate, oxalate, citrate, methanesulfonate, benzenesulfonate, toluenesulfonate and maleate; salts with bases, for example alkali metal salts such as sodium and potassium salts, and alkaline earth metal salts such as calcium salts; salts with amino acids such as glycinate, lysinate, arginate, ornithine, glutamate and aspartate; and so on.

The compound (I) or a pharmaceutically acceptable salt thereof includes intramolecular salts or adducts thereof, and also includes solvates or hydrates thereof.

Further, it is known that a crystal polymorphism exists in the compound (I) (see international publication No. 2014/024950). Therefore, in the present invention, the compound (I) as an active ingredient may be used in any crystalline form based on such a crystalline polymorphism.

Pharmaceutical preparation

In practicing the present invention, compound (I) or a pharmaceutically acceptable salt thereof (hereinafter, these may be collectively referred to as "the compound of the present invention") may be used in a separate form or in the form of a pharmaceutical composition containing the compound of the present invention as an active ingredient together with a pharmaceutically acceptable carrier.

Examples of such pharmaceutical compositions include tablets, pills, powders, granules, capsules and emulsions.

In the present specification, as the "pharmaceutically acceptable carrier", various carriers commonly used in the technical field of pharmaceutical preparations can be used.

As specific examples of the "pharmaceutically acceptable carrier", for example, in solid pharmaceutical preparations, excipients, lubricants, binders, and disintegrants may be used.

In liquid pharmaceutical formulations, vehicles, dissolution aids, suspending agents, isotonic agents and buffers may be used.

If necessary, other necessary additives such as a preservative may be blended.

The pharmaceutical composition of the present invention differs depending on the dosage form, administration method, carrier, etc., but can be produced by adding the compound of the present invention in an amount of usually 0.01 to 99% (w/w), and preferably 0.1 to 85% (w/w) with respect to the total amount of the pharmaceutical preparation. Depending on its form, the pharmaceutical composition may be produced using methods commonly used in the art of pharmaceutical formulation technology. The pharmaceutical composition of the present invention can be molded into sustained-release pharmaceutical preparations containing an active ingredient.

Hereinbefore, the compounds of the present invention and the pharmaceutical compositions of the present invention have been described. The compound of the present invention and the pharmaceutical composition of the present invention are expected to have the following excellent effects.

For example, when steroid-type MR antagonists such as spironolactone or eplerenone are used, there is concern about serious side effects (e.g., gynecomastia, irregular menstruation, erectile dysfunction, etc.). However, with the compound of the present invention and the pharmaceutical composition of the present invention, there is little fear about such serious side effects, and therefore, the compound of the present invention and the pharmaceutical composition of the present invention can have high safety as a medicament.

In addition, eplerenone is metabolized primarily by CYP3a4, and thus, is prohibited from being used in combination with a strong CYP3a4 inhibitor. However, in the case of the compound of the present invention, the metabolic pathways are different, and therefore, the compound of the present invention is not so limited and can be used in combination with a wide range of other drugs. Therefore, the compounds of the present invention and the pharmaceutical compositions of the present invention are highly useful in clinical practice.

Further, the compound of the present invention and the pharmaceutical composition of the present invention have a characteristic in terms of pharmacokinetics that a constant drug level in plasma can be maintained for a long period of time, and therefore, a sustained effect can be exerted even at a low dose. Therefore, also from this viewpoint, the compound of the present invention and the pharmaceutical composition of the present invention can be used as a medicament having low toxicity and high safety.

Subject to be administered

As described above, the compound of the present invention has low toxicity and is expected to have few side effects, and also has excellent properties as a pharmaceutical agent. Thus, the compounds of the present invention can be safely administered to mammals (particularly humans).

Route of administration

In practicing the present invention, the compounds of the present invention may be administered orally or parenterally (e.g., intravenously, intramuscularly, subcutaneously, intraorganoleptically, intranasally, intradermally, intraocularly, intracerebrally, intrarectally, intravaginally, and intraperitoneally, as well as administration to a lesion), either independently, or as a pharmaceutical composition.

Dosage form

The dose of the compound of the present invention varies depending on the subject to be administered, the administration route, and the age and symptoms of the subject to be administered, but is not particularly limited. For example, when the compound of the present invention is orally administered to an adult patient with pulmonary fibrosis (having a body weight of about 40-80 kg, e.g., 60 kg), the daily dose thereof is, for example, 1-30mg, preferably 1-25mg, even more preferably 2.5-25mg, and particularly preferably 7.5-25 mg. This amount may be administered on a dosing schedule of 1-3 times per day.

Used in combination with other drugs

As described above, the compound of the present invention has extremely low toxicity and can be used for the prophylaxis or treatment of pulmonary fibrosis in combination with other drugs, and an excellent prophylactic effect and/or therapeutic effect by combination with other drugs can be expected. Further, such combination therapy can be expected to reduce the dosage of other drugs and reduce the side effects of other drugs.

Examples of such drugs (hereinafter, abbreviated as concomitant drugs) that can be used in combination with the compound of the present invention include steroid drugs (e.g., prednisolone, methylprednisolone, etc.), immunosuppressive agents (e.g., cyclophosphamide, cyclosporine, etc.), and anti-fibrotic agents (e.g., nintedanib, pirfenidone, etc.).

In actual combination therapy, concomitant drugs may be appropriately selected in view of the type of disease of a patient, the severity of symptoms thereof, and the like.

The dosage form of the concomitant drug of the present invention is not particularly limited, and the compound of the present invention and the concomitant drug may be combined at the time of administration. For example, concomitant medication may be used in the form of: (1) administering a pharmaceutical formulation comprising a combination of a compound of the invention and a concomitant drug; (2) administering two pharmaceutical preparations obtained by separately formulating the compound of the present invention and the concomitant drug via the same administration route, simultaneously or separately; and (3) administering two pharmaceutical preparations obtained by separately formulating the compound of the present invention and the concomitant drug via different administration routes, simultaneously or separately. The preferred form can be appropriately selected according to the actual situation in clinical practice.

From the above-described pharmaceutical compositions containing the compounds of the present invention, pharmaceutical preparations containing a combination of the compounds of the present invention and concomitant drugs can be appropriately produced by those skilled in the art.

The dosage of the concomitant drug can be appropriately selected based on the clinically used dosage. Further, the compounding ratio of the compound of the present invention and the concomitant drug may be appropriately selected depending on the disease and symptom of the subject to be administered, the administration route, the type of the concomitant drug to be used, and the like. In general, the compounding ratio can be determined as appropriate in accordance with actual circumstances in clinical practice based on a general clinical dose of a concomitant drug to be used.

Examples

Hereinafter, the present invention is described in more detail based on examples. However, these examples do not limit the scope of the present invention. Further, unless otherwise specified, reagents, devices and materials used in the present invention are commercially available or can be appropriately prepared by those skilled in the art.

Example 1: anti-fibrotic effects in bleomycin intraairway administration-induced pulmonary fibrosis model in mice (1)

(1) Test method

C57/BL6NTAC mice (male; body weight at The start of The test: 25g) (manufactured by The Jackson Laboratory) were divided into The following 4 groups (12 mice each):

group 1: normal group

Group 2: group without drug administration (control group)

Group 3: drug (Compound (I)) administration group

Group 4: drug (eplerenone; positive control) administration group

On the day of test initiation (day 0), bleomycin (3.25U/kg) was administered dropwise via the airway to mice of groups 2 to 4 as the test group. In place of bleomycin, saline was dropwise administered to mice of group 1 as a normal group through the airway. On day seven after the start of the test (day 7), oral gavage administration of vehicle (0.1% HCO60+ 0.5% CMC) was initiated for mice of group 1 and group 2 (twice in the morning and afternoon) using a dosing schedule twice daily in the morning and afternoon; for group 3 mice, a 2mg/ml solution of compound (I) was prepared and oral gavage administration (morning) of it and vehicle (0.1% HCO60+ 0.5% CMC) at a dose of 10mg/kg was initiated (afternoon); and oral gavage administration of eplerenone at a dose of 50mg/kg was initiated for group 4 mice (twice in the morning and afternoon). Thereafter, until the twenty-first day after initiation of the test (day 21), administration was continued with the same dosing schedule. After completion of administration (day 21), the following evaluation items were checked.

a) Assessment of pulmonary function

The following evaluation items were measured using commercially available respiratory and pulmonary function evaluation systems (measurement site: around airway). The 4 evaluations included: 1) deep gas uptake (mL); 2) compliance (mL/cmH)₂O); 3) elasticity (cmH)₂O/mL); and 4) resistance (cmH)₂O.s/mL)。

The "deep inspiratory volume" corresponds to the amount of air expelled from the lungs at the stage from slow expiration to completion of breathing, and is an indicator of the lung volume. As pulmonary fibrosis progresses and the lung becomes stiffer, lung volume shrinks and the amount of deep inspiration decreases.

"compliance" indicates a change in lung volume due to some change in pressure. Large compliance means that changes in lung volume vary greatly with respect to unit pressure and indicate that the lung is prone to stretching. As pulmonary fibrosis progresses and the lung hardens, lung volume shrinks and compliance decreases.

"elasticity" is a value represented by the inverse of compliance, and is an index representing difficulty in stretching the lung. As pulmonary fibrosis progresses and the lung hardens, elasticity increases.

By "resistance" is meant airway resistance. Resistance is the resistance experienced by airflow in breathing, and greater resistance means that air is more difficult to pass through the airway.

b) Assessment of lung pathology

(i) Evaluation by sirius red staining (left lung lobe)

Pulmonary fibrosis is caused by the accumulation of activated fibroblasts at the site of fibrosis and the production of large amounts of type I collagen. Thus, based on the extent of collagen accumulation in lung tissue, the extent of pulmonary fibrosis can be assessed. Type I collagen and type III collagen in tissues were stained by sirius red staining. Therefore, image diagnosis of the situation of collagen accumulation can be performed, and quantitative evaluation can also be performed by calculating the staining positive rate.

Specifically, the above staining is performed on a tissue section of the lung, and the area of the stained site (fibrosis site) is measured and evaluated.

(ii) Evaluation of hydroxyproline content (μ g) (lower and middle lung lobes)

Hydroxyproline is the major component of collagen and is substantially absent in other proteins. Thus, by measuring the hydroxyproline content in the tissue, the amount of collagen can be assessed.

(iii) Assessment of Gene expression fluctuations associated with pulmonary fibrosis (quantitation of mRNA; fold increase) (upper lung lobes)

mRNA was extracted from homogenized upper lung leaves according to conventional methods, and very small amounts of mRNA were quantified using a real-time PCR system.

c) Evaluation of blood samples

At 2 hours (before the above evaluation) and 18 hours from the last administration (day 21) of compound (I), a small amount of blood was collected from the tail vein under anesthesia, and the plasma was divided with a refrigerated centrifuge and the concentration (nM) of compound (I) in the plasma was measured using a conventional method.

(2) Test results

a) Assessment of pulmonary function

The test results are shown in fig. 1 and 2.

As shown in fig. 1, bleomycin reduced deep inspiratory capacity and increased elasticity (group 2). Eplerenone significantly ameliorated this decrease in lung function due to bleomycin (group 4).

In contrast, compound (I) also showed a significant improvement effect in all lung functions, and was found to significantly improve in particular the deep inspiratory capacity to a degree exceeding the effect of eplerenone (group 3).

As shown in fig. 2, bleomycin decreased lung compliance and increased resistance (group 2). Eplerenone significantly ameliorated this decrease in lung function due to bleomycin (group 4).

In contrast, compound (I) also showed a significant effect of improving all lung functions, and it was found that in particular the resistance was very significantly improved to the level of the normal group, exceeding the effect of eplerenone (group 3).

From the above, in the present test, from the viewpoint of lung function, it was confirmed that compound (I) has an excellent improving effect on pulmonary fibrosis even compared with eplerenone.

b) Assessment of lung pathology

(i) Evaluation by sirius red staining (left lung lobes) and (ii) hydroxyproline content (μ g) (lower and middle lung lobes)

The test results are shown in fig. 3.

Bleomycin increased collagen accumulation in the lung as shown by an increase in hydroxyproline content (hydroxyproline; μ g) in the lower and middle lung lobes, and an increase in the positive rate in sirius red staining (PSR positive%) in the left lung lobe (group 2). This fibrosis by bleomycin was significantly improved by eplerenone (group 4).

In contrast, compound (I) also showed a significant improvement in the effect of pulmonary fibrosis in all the evaluation items, and it was found that particularly the hydroxyproline content of the lung tissue was significantly improved to a degree exceeding the effect of eplerenone (group 3).

From the above, in the present test, even from the histopathological viewpoint, it was confirmed that compound (I) has an excellent ameliorating effect on pulmonary fibrosis even compared to eplerenone.

(iii) Assessment of Gene expression fluctuations associated with pulmonary fibrosis

The test results are shown in fig. 4-7. By assessing fluctuations in gene expression associated with pulmonary fibrosis, the therapeutic and prophylactic effects of compound (I) can be predicted.

Bleomycin increases gene expression of type I collagen, type III collagen and type IV collagen. Eplerenone does not reduce expression of these genes.

In contrast, compound (I) significantly reduced the expression of these genes (fig. 4).

From the above, in the present test, even from the viewpoint of controlling collagen production directly involved in pulmonary fibrosis, it was confirmed that compound (I) has an excellent improving effect on pulmonary fibrosis even compared to eplerenone.

Bleomycin slightly increased alpha SMA gene expression. Eplerenone does not reduce this gene expression. In contrast, compound (I) significantly reduced this gene expression (fig. 5-1).

Bleomycin significantly increased fibronectin and vimentin gene expression. Eplerenone significantly reduced expression of these genes compared to bleomycin.

In contrast, compound (I) significantly reduced the expression of these genes (fig. 5-2 and 5-3).

From the above, in the present test, it was confirmed that compound (I) has excellent control ability with respect to gene expression of α SMA, fibronectin and vimentin involved in pulmonary fibrosis.

Bleomycin significantly increased gene expression of CTGF, PAI-1 and Ccl 2. Eplerenone significantly reduced expression of these genes compared to bleomycin.

In contrast, compound (I) significantly reduced the expression of these genes (fig. 6-1-6-3).

From the above, in the present test, it was confirmed that the compound (I) has excellent control ability with respect to gene expression of CTGF, PAI-1 and Ccl2 involved in pulmonary fibrosis.

Bleomycin had no effect on the gene expression of TGF β. Eplerenone showed a trend towards increased gene expression, while compound (I) showed a trend towards decreased gene expression (fig. 7).

c) Evaluation of blood samples

The test results are shown in fig. 8.

Sufficient concentrations of compound (I) in plasma were detected at 2 and 18 hours after the last administration.

This finding of hemodynamics indicates that sufficient exposure in the blood was obtained after administration of compound (I), and indicates the usefulness of compound (I) as a pulmonary fibrosis drug.

These results indicate that compound (I) has a prophylactic or therapeutic effect on pulmonary fibrosis equal to or higher than eplerenone.

Example 2: anti-fibrotic effects in bleomycin subcutaneous continuous infusion induced pulmonary fibrosis model in mice (2)

(1) Test method

On the day of test initiation (day 0), an ALZET 1007D pump (in which bleomycin: 100U/kg, or saline: 100 μ l) (manufactured by ALZET) was stored) was embedded in the abdominal cavity of each C57/BL6NTAC mouse (male, body weight 25-30 g at test initiation) (manufactured by Taconic Biosciences ltd.) under anesthesia, and administration of bleomycin to the test group (48 mice) was initiated, and on the other hand, administration of saline to the normal group (24 mice) (group 1) was initiated, and thereafter, administration was continued for 7 days (flow rate: 0.5 μ l/hr) (pump was removed on the tenth day after test initiation (day 10). The body weights of the mice of the test group were measured every day, and at the seventh day after the start of the test (day 7), the amount of decrease in body weight was used as an index, and the mice of the test group were randomly grouped as follows.

Group 2: group without drug administration (control group) (12 mice)

Group 3: drug (Compound (I), 3mg/kg) administration group (12 mice)

Group 4: drug (Compound (I), 10mg/kg) administration group (12 mice)

Group 5: drug (Compound (I), 30mg/kg) administration group (12 mice)

Starting from the eighth day (day 8) after the start of the test, oral gavage administration of the starting vehicle (0.1ml/10g body weight) to the group 1 mice as the normal group and the group 2 mice as the drug-free administration group was initiated, and on the other hand, oral gavage administration of the starting predetermined dose of compound (I) to the drug-administration group mice of groups 3 to 5 (once a day in the morning for all groups) was initiated. Thereafter, administration continued until the twenty-first day (day 21) after initiation of the test. After completion of the administration, the following items were evaluated.

a) Assessment of pulmonary function

The 4 evaluations included: 1) deep gas uptake (mL); 2) compliance (mL/cmH)₂O); 3) elasticity (cmH)₂O/mL); and 4) resistance (cmH)₂O.s/mL)。

b) Assessment of lung pathology

i) Evaluation by sirius red staining (left lung lobe)

ii) evaluation of hydroxyproline content (μ g) (right lung lobe)

c) Evaluation of blood samples

Evaluation of compound (I) concentration (nM) in plasma 2 hours (before the above evaluation) and 24 hours after the last administration (day 21) (evaluation of blood samples obtained by tail vein blood sampling, similar to example 1.)

(2) Test results

a) Assessment of pulmonary function

The test results are shown in fig. 9 and 10.

As shown in fig. 9, bleomycin reduced deep inspiratory volume and increased resistance (group 2). In contrast, compound (I) at doses of 10mg/kg and 30mg/kg significantly improved the deep inspiratory capacity (groups 4 and 5), and compound (I) at a dose of 30mg/kg significantly improved the resistance (group 5).

As shown in fig. 10, bleomycin increased elasticity and decreased compliance (group 2). Compound (I) significantly ameliorated this reduction in bleomycin in both lung functions in a dose-dependent manner (groups 3-5).

From the above, in the present test, from the viewpoint of lung function, it was confirmed that compound (I) had an excellent ameliorating effect on pulmonary fibrosis in a dose-dependent manner.

b) Assessment of lung pathology

The test results are shown in fig. 11.

Bleomycin increased collagen accumulation in the lung as shown by increased hydroxyproline content in the right lung, and increased positive rate in sirius red staining in the left lung (group 2). In contrast, compound (I) at the doses of 3mg/kg and 30mg/kg significantly reduced hydroxyproline content (groups 3 and 5), and compound (I) at all doses significantly reduced the positive rate in sirius red staining (groups 3-5).

From the above, in the present test, even from the histopathological viewpoint, it was confirmed that compound (I) has an excellent ameliorating effect on pulmonary fibrosis in a dose-dependent manner.

c) Evaluation of blood samples

The test results are shown in fig. 12.

Compound (I) showed a dose-dependent concentration in plasma 2 hours after the last administration. Sufficient concentrations in plasma were detected 24 hours after the last administration for compound (I) at a dose of 30 mg/kg.

This finding of hemodynamics indicates that compound (I) exposure in the blood is obtained depending on the dose, and indicates the usefulness of compound (I) as a pulmonary fibrosis drug.

These results indicate that compound (I) has a dose-dependent effect and indicate the usefulness of compound (I) as a pulmonary fibrosis drug.

Pulmonary fibrosis is primarily advanced fibrosis of the lung and is a pulmonary disease that causes restricted ventilation disorders. Pulmonary fibrosis is thought to be caused by: due to the repetition of inflammation in the lung interstitium, abnormal injury repair responses with continued alveolar epithelial cell injury are repeated.

There may be cases where the cause for inflammation in the pulmonary interstitium, such as infection, collagen disease, radiation, drugs, dust, etc., can be identified, and there may be cases where the cause cannot be identified. However, when the inflamed tissue has become fibrotic, it progresses to pulmonary fibrosis. Here, pulmonary fibrosis for which the cause cannot be identified is referred to as idiopathic pulmonary fibrosis. Idiopathic pulmonary fibrosis is also frequent and has poor prognosis, and most often develops in adults 50 years or older, and causes irreversible fibrosis in the lung, and is fatal by reducing respiratory function.

For the treatment of pulmonary fibrosis, in general, steroid drugs and immunosuppressive agents have been used. However, the following findings have been accumulated: particularly for idiopathic pulmonary fibrosis, fibrosis worsens when steroid drugs or immunosuppressants are used over a long period of time. For pulmonary fibrosis of various or unknown causes, in general, there is no drug treatment that can be widely recommended for the treatment of pulmonary fibrosis.

In recent years, as a new type of idiopathic pulmonary fibrosis therapeutic agent, an anti-fibrosis agent (e.g., pirfenidone, nintedanib) is marketed. However, there is a problem that these two drugs have strong side effects. For example, with respect to pirfenidone, the possibility of skin carcinogenesis due to exposure to light must be considered for use of the drug, and treatment options remain practically limited for this severe disease. On the other hand, spironolactone, which is a steroid-type mineralocorticoid receptor antagonist (MR antagonist), has been reported to improve hydroxyproline accumulation, which is an index of tissue fibrosis in bleomycin-induced pulmonary fibrosis mice (see European Journal of Pharmacology, 718(2013), 290-298; PLOS ONE, 8(11) (2013), e 81090; and Nanomedicine (Lond.), 11(11) (2016), 1393-1406).

As described above, effective treatment for pulmonary fibrosis is not yet fully established at present, and early treatment establishment through development of a new drug with high safety and high therapeutic effectiveness is strongly required.

In international publication No. 2007/089034, a 1, 4-benzoxazine compound containing compound (I) of the present invention is described, and compound (I) is described as an MR antagonist. Further, in international publication No. 2018/062134, compound (I) is described for use in the treatment of non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), and the like. However, none of the documents specifically mentions the use of the compounds (I) of the present invention for pulmonary fibrosis.

As described above, drug therapy for pulmonary fibrosis has not been fully established, and development of novel prophylactic or therapeutic agents has become an urgent task in the medical technical field.

According to the present invention, an agent which contains compound (I) as an active ingredient and can be effectively and safely used for preventing or treating pulmonary fibrosis can be provided.

Unless otherwise defined, technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

The present invention provides a novel agent which contains compound (I) as an active ingredient and can be effectively and safely used for preventing or treating pulmonary fibrosis.

Obviously, many modifications and variations of the present invention are possible in light of the above teachings. It is therefore to be understood that within the scope of the appended claims, the invention may be practiced otherwise than as specifically described herein.