Pharmaceuticals to prevent or treat fibrosis.

Abstract

The invention relates to a pharmaceutical that is useful for preventing or treating fibrosis. According to the invention, there is indication that the effect of preventing or treating fibrosis can be further enhanced by the use of the first and second components in combination.

Description

Pharmaceutical composition for the prevention or treatment of fibrosis

[0001] The present invention relates to a pharmaceutical composition that can be usefully used for the prevention or treatment of fibrosis.

[0002] Fibrosis refers to the hardening of a portion of an organ for any reason. Pulmonary fibrosis and liver fibrosis are representative examples. Pulmonary fibrosis is most often caused by radiation exposure or pulmonary edema, but some people can develop pulmonary fibrosis. There is currently no cure for fibrosis, and treatment options are still being developed and researched.Types of fibrosis include interstitial lung disease (ILD), scleroderma, keloid, hypertrophic scar, non-alcoholic fatty liver disease, primary sclerosing cholangitis (PSC), primary biliary cholangitis (PBC), diabetic retinopathy, age-related macular degeneration (AMD), hypertrophic cardiomyopathy, myocardial infarction, muscular dystrophy, diabetic kidney disease, focal segmental glomerulosclerosis (FSGS), inflammatory bowel disease, IBD) and interstitial lung diseases include idiopathic pulmonary fibrosis (IPF), systemic sclerosis associated interstitial lung disease (SSc-ILD), and chronic fibrosing interstitial lung diseases with a progressive phenotype (PF-ILD).

[0003]

[0004] Idiopathic pulmonary fibrosis (IPF) is a rare, chronic, progressive interstitial lung disease known for its poor prognosis and lack of proven treatment. To date, no clear cause has been identified, and the five-year survival rate after diagnosis is only 43%, and the ten-year survival rate is approximately 15%. Despite extensive research, no treatment has yet been shown to improve survival. Considering that other interstitial lung diseases, such as nonspecific interstitial pneumonia (NSIP) and idiopathic organizing pneumonia (COP), have relatively good prognoses with appropriate treatment, IPF can be considered one of the most unfavorable interstitial lung diseases. The most common causes of death are respiratory failure (39%) and cardiac disease (27%), with other causes including lung cancer, pulmonary embolism, and pneumonia. The prognosis is worse in older age, male gender, those with poor lung function at diagnosis, or those with a high number of fibroblastic foci in biopsies.

[0005]

[0006] Treatment for idiopathic pulmonary fibrosis (IPF) is similar to that for nonspecific interstitial pneumonia (NSIP), with steroids and cytotoxic drugs sometimes used. Recently, antifibrotic agents have become the mainstay of treatment, with various trials underway. Currently, pirfenidone and nintedanib are approved for IPF. These drugs do not provide a cure, but rather delay pulmonary fibrosis and alleviate symptoms. Therefore, there is a need for more effective medications that can improve patients' quality of life.

[0007]

[0008] Systemic sclerosis-associated interstitial lung disease (SSc-ILD) is a condition in which patients with systemic sclerosis (SSc) develop interstitial lung disease (ILD) as a complication, and lung function decline is the leading cause of death in SSc. Nintedanib and tocilizumab, which have been shown to reduce lung function decline, are currently approved treatments for this condition in the United States. However, as with idiopathic pulmonary fibrosis, there is a need for more effective medications that can improve patients' quality of life.

[0009]

[0010] Chronic fibrosing interstitial lung diseases with a progressive phenotype (PF-ILD) refers to various progressive fibrosing interstitial lung diseases excluding idiopathic pulmonary fibrosis, including autoimmune interstitial lung disease and idiopathic interstitial pneumonia. Nintedanib was approved for treatment in the United States after being confirmed to be effective in reducing lung function decline in patients with various fibrosing lung diseases. Since fibrosing interstitial lung diseases can present with a progressive phenotype such as pulmonary fibrosis, lung function decline, and worsening quality of life, regardless of classification and underlying disease, if a drug is proven to be effective in reducing lung function decline in a specific interstitial lung disease, it can be expected to be effective in reducing lung function decline in other interstitial lung diseases as well.

[0011]

[0012] Meanwhile, prolyl-tRNA synthetase (PRS) is a member of the aminoacyl-tRNA synthetase (ARS) family of enzymes that activates amino acids for protein synthesis. Specifically, ARS forms aminoacyl adenylate (AA-AMP) and then transfers the activated amino acid to the 3-terminal of the corresponding tRNA (translational function). Because ARS plays a key role in protein synthesis, ARS inhibition inhibits the growth and development of all cells. Accordingly, ARS is recognized as a promising target for antibiotics and therapeutic agents that require inhibition of cell overexpression (Nature, 2013, 494: 121-125).

[0013]

[0014] PRS exists and functions as a multisynthetase complex (MSC) in the form of EPRS (Glutamyl-Prolyl-tRNA Synthetase). In particular, among various MSCs, EPRS functions as a translational silencer that suppresses the production of vascular endothelial growth factor A (VEGF A), a key factor in angiogenesis, and has also been reported to be closely related to various solid cancers (Nat. Rev. Cancer, 2011, 11, 708-718).

[0015]

[0016] Accordingly, the inventors of the present invention have conducted extensive research on a method for preventing or treating fibrosis and have confirmed that more effective prevention or treatment of fibrosis is possible when a specific PRS inhibitor is used in combination with an existing fibrosis treatment agent, thereby completing the present invention.

[0017] The present invention provides a pharmaceutical composition that can be usefully used for the prevention or treatment of fibrosis.

[0018] To solve the above problem, the present invention provides a pharmaceutical composition for preventing or treating fibrosis as follows:

[0019] 1) A first component of a compound represented by the following chemical formula 1 or a pharmaceutically acceptable salt thereof, and

[0020] 2) A second component comprising any one selected from the group consisting of a compound represented by the following chemical formula 2, a pharmaceutically acceptable salt thereof, a compound represented by the following chemical formula 3, and a pharmaceutically acceptable salt thereof.

[0021] A pharmaceutical composition for preventing or treating fibrosis,

[0022] The first component and the second component are co-administered in the same formulation or different formulations,

[0023] Pharmaceutical composition:

[0024] [Chemical Formula 1]

[0025]

[0026] [Chemical Formula 2]

[0027]

[0028] [Chemical Formula 3]

[0029] .

[0030]

[0031] The pharmaceutical composition according to the present invention comprises the first component and the second component as described above, so that the preventive or therapeutic effects of each component are combined with each other to enable more effective prevention or treatment of fibrosis.

[0032]

[0033] The first component is a compound described in Korean Patent Registration No. 10-2084772, specifically, a compound described as Example 40 of the specification. The first component, as a PRS inhibitor, can be used together with the second component, which has been used for the prevention or treatment of fibrosis as described below, to exhibit a more effective effect in the prevention or treatment of fibrosis.

[0034]

[0035] The second component is a component used for the prevention or treatment of fibrosis, and the chemical formulae 2 and 3 are components known as pirfenidone and nintedanib, respectively. Compared to the previously known effect of preventing or treating fibrosis of the second component, when used in combination with the first component as in the present invention, the effect is further enhanced.

[0036]

[0037] The weight ratio of the first component to the second component is preferably 1:0.5 to 1:30. Within this range, the respective effects of the first component and the second component interact with each other to enhance the prevention or treatment of fibrosis. More preferably, the weight ratio of the first component to the second component is 1:0.6 to 1:25.

[0038]

[0039] Preferably, the second component is a compound represented by the chemical formula 2, or a pharmaceutically acceptable salt thereof, and the weight ratio of the first component and the second component is 1:2 to 1:25, more preferably 1:2 to 1:20, 1:2 to 1:12, 1:2 to 1:8, 1:6 to 1:12, or 1:6 to 1:8.

[0040]

[0041] Preferably, the second component is a compound represented by the chemical formula 3, or a pharmaceutically acceptable salt thereof, and the weight ratio of the first component and the second component is 1:0.6 to 1:10, more preferably 1:0.6 to 1:6, 1:0.6 to 1:1.5, or 1:1 to 1:1.5.

[0042]

[0043] Additionally, in the pharmaceutical composition according to the present invention, the first component is included in an amount of 100 to 150 mg. In addition, the content of the second component can be adjusted according to the content of the first component.

[0044]

[0045] Preferably, in the pharmaceutical composition according to the present invention, the second component is a compound represented by the chemical formula 2, or a pharmaceutically acceptable salt thereof, and contains 100 to 150 mg of the first component and 200 to 800 mg of the second component.

[0046]

[0047] Preferably, in the pharmaceutical composition according to the present invention, the second component is a compound represented by the chemical formula 3, or a pharmaceutically acceptable salt thereof, and comprises 100 to 150 mg of the first component and 100 to 150 mg of the second component.

[0048]

[0049] Preferably, the first component and the second component are each administered twice a day or three times a day. Preferably, the first component is administered twice a day and the second component is administered three times a day, or the first component is administered three times a day and the second component is administered twice a day.

[0050]

[0051] Meanwhile, the compounds represented by the above chemical formulas 1 to 3 can each be used in the form of a pharmaceutically acceptable salt, and as a salt, an acid addition salt formed by a pharmaceutically acceptable free acid is useful. Inorganic acids and organic acids can be used. Inorganic acids that can be used include hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, etc., and organic acids that can be used include citric acid, acetic acid, lactic acid, maleic acid, gluconic acid, methanesulfonic acid, succinic acid, 4-toluenesulfonic acid, glutamic acid, or aspartic acid.

[0052]

[0053] In addition, the compounds represented by the above chemical formulae 1 to 3 can be prepared in crystalline or amorphous form, and when prepared in crystalline form, can be optionally hydrated or solvated. The present invention may include not only stoichiometric hydrates of the compounds represented by the above chemical formulae 1 to 3, but also compounds containing various amounts of water. The solvates of the compounds represented by the above chemical formulae 1 to 3 include both stoichiometric solvates and non-stoichiometric solvates.

[0054]

[0055] Meanwhile, examples of the fibrosis include interstitial lung disease (ILD), scleroderma, keloid, hypertrophic scar, non-alcoholic fatty liver disease, primary sclerosing cholangitis (PSC), primary biliary cholangitis (PBC), diabetic retinopathy, age-related macular degeneration (AMD), hypertrophic cardiomyopathy, myocardial infarction, muscular dystrophy, diabetic kidney disease, focal segmental glomerulosclerosis (FSGS), or inflammatory bowel disease. There is idiopathic pulmonary fibrosis (IPF), systemic sclerosis associated interstitial lung disease (SSc-ILD), or chronic fibrosing interstitial lung diseases with a progressive phenotype (PF-ILD).

[0056]

[0057] The term "prevention" of the present invention means any act of inhibiting or delaying the occurrence, spread, and recurrence of the disease by administering the composition of the present invention, and "treatment" means any act of improving or beneficially changing the symptoms of the disease by administering the composition of the present invention.

[0058]

[0059] The pharmaceutical composition of the present invention can be formulated into oral or parenteral dosage forms according to standard pharmaceutical practice. These dosage forms may contain, in addition to the active ingredient, pharmaceutically acceptable additives such as carriers, adjuvants, or diluents.

[0060]

[0061] Suitable carriers include, but are not limited to, saline, polyethylene glycol, ethanol, vegetable oils, and isopropyl myristate, and diluents include, but are not limited to, lactose, dextrose, sucrose, mannitol, sorbitol, cellulose, and / or glycine. In addition, the compounds of the present invention can be dissolved in oils, propylene glycol, or other solvents commonly used in the preparation of injectable solutions. In addition, the compounds of the present invention can be formulated as ointments or creams for topical action.

[0062]

[0063] The pharmaceutical dosage forms of the compounds of the present invention can be used in the form of their pharmaceutically acceptable salts or solvates, and can also be used alone or in combination with other pharmaceutically active compounds as well as in an appropriate combination.

[0064]

[0065] The compounds of the present invention can be formulated as injections by dissolving, suspending, or emulsifying the compounds in water-soluble solvents such as normal saline, 5% dextrose, or in water-insoluble solvents such as synthetic fatty acid glycerides, higher fatty acid esters, or propylene glycol. The formulations of the present invention can include conventional additives such as solubilizers, isotonic agents, suspending agents, emulsifiers, stabilizers, and preservatives.

[0066]

[0067] The preferred dosage of the compound of the present invention varies depending on the patient's condition and body weight, the extent of the disease, the form of the drug, the route of administration, and the duration of administration, but can be appropriately selected by those skilled in the art. However, for a desirable effect, it is recommended to administer the compound of the present invention at 0.0001 to 100 mg / kg (body weight) per day, preferably 0.001 to 100 mg / kg (body weight). Administration can be administered once a day or in divided doses via oral or parenteral routes. Depending on the administration method, the composition can contain 0.001 to 99 wt%, preferably 0.01 to 60 wt%, of the compound of the present invention.

[0068]

[0069] The pharmaceutical composition of the present invention can be administered to mammals, including rats, mice, livestock, and humans, via various routes. Any route of administration is conceivable, including oral, rectal, intravenous, intramuscular, subcutaneous, intrauterine, or intracerebroventricular injection.

[0070] As described above, the pharmaceutical composition according to the present invention can be usefully used for the prevention or treatment of fibrosis by using the first component and the second component together.

[0071] Figures 1 and 2 show the results of expression of genes involved in collagen synthesis in Experimental Example 1 of the present invention.

[0072] Figure 3 shows the results of lung function evaluation of Experimental Example 2-1 of the present invention.

[0073] Figure 4 shows the results of histopathological analysis of Experimental Example 2-3 of the present invention.

[0074] Figure 5 is a microscopic photograph of lung tissue of a vehicle object in Experimental Example 2-3 of the present invention.

[0075] Figure 6 is a microscopic photograph of lung tissue of an individual administered with the first component and the second component (PID) in Experimental Example 2-3 of the present invention.

[0076] Figure 7 shows the results of the inflammatory cell infiltration analysis of Experimental Example 2-4 of the present invention.

[0077] Figure 8 shows the results of measuring body weight change in Experimental Example 2-5 of the present invention.

[0078] Figure 9 shows the change in blood concentration of the active ingredient of Experimental Example 3 of the present invention.

[0079] Figure 10 shows the change in plasma concentration for a single administration of Experimental Example 4 of the present invention.

[0080] Figure 11 shows the change in plasma concentration for multiple administrations of Experimental Example 4 of the present invention.

[0081] Figure 12 is a graph showing the correlation between AUC and the administered dose in Experimental Example 4.

[0082] Hereinafter, the present invention will be described in more detail with reference to the following examples. However, the following examples are intended only to illustrate the present invention and the scope of the present invention is not limited to these examples.

[0083]

[0084] Manufacturing Example 1: First Component

[0085] The following compound was prepared in the same manner as Example 40 of Korean Patent Registration No. 10-2084772, and was hereinafter named 'first component' or 'Example'.

[0086]

[0087] 1 H NMR (500 MHz, MeOD): δ 9.67 (s, 1H). 8.02 (d, 1H), 7.82 (d, 1H), 4.62 (m, 2H), 3.60 (m, 1H), 3.28 (m, 1H), 2.99 (m, 2H), 2.25 (m, 2H), 2.08 (m, 2H), 1.99 (m, 1H), 1.78 (m, 2H), 1.54 (m, 1H)

[0088]

[0089] Manufacturing Example 2: Second Component

[0090] Nintedanib (hereinafter referred to as the “second ingredient (NIN)”) and pirfenidone (hereinafter referred to as the “second ingredient (PID)”) were purchased and used as commercial products, and are specifically as follows.

[0091] FunctionControl(or reference) ArticleControl(or reference) ArticleCompound NameNintedanibPirfenidoneSalt formFreeFreeManufacturerU chemCombi-BlocksSupplierU chemCombi-Blocks

[0092]

[0093] Experimental Example 1: Nonclinical Antifibrotic Efficacy Evaluation

[0094] The first component (10 mM) was diluted in DMSO to 100 uM. The dilutions were further diluted with DMSO to 10, 3, and 1 uM, and the second component (NIN) (10 mM) was diluted with DMSO to 50 nM.

[0095]

[0096] DHLF cells (FGMTM-2 Bullet KitTM, 10% FBS), a lung fibrosis cell line, were prepared and cultured in FBM medium in a T75 Easy Flask Filter at 37°C and 5% CO2. The cultured DHLF cells were treated with 10 ng / mL of TGF-β and the test drugs alone or in combination, and cultured for 72 hours. The media was removed, proteins were extracted, and quantified using the BCA Protein Assay Kit. Based on the quantitative value of each extracted protein, 10 to 20 μg of protein was subjected to Western blotting. After running, the proteins were transferred to PVDF membranes, and after three TBS-T washes, 1 mL of ECL solution was treated to each PVDF membrane, and the protein expression level was measured using an AI680 imager. The values ​​of each band were normalized to β-actin using ImageJ, and are shown in Tables 2 and 3 and Figures 1 and 2.

[0097] TGF-β 2nd component (NIN) 1st component No. Relative value (control standard) COL1A1 SMA-α--control 41.00 1.00 10 ng / ml vehicle 46.25 3.78 50 ng / ml-44.60 2.63 1 ug / ml 44.31 2.403 ug / ml 43.09 1.56 10 ug / ml 42.14 1.05-10 ug / ml 43.26 1.66

[0098]

[0099] TGF-β 1st component 2nd component (NIN) No. Relative value (control standard) COL1A1SMA-α-control-21.001.0010 ng / ml vehicle-24.836.735 ug / ml-23.843.5325 ng / ml24.113.5850 ng / ml23.122.10100 ng / ml21.810.77-100 ng / ml22.812.99

[0100]

[0101] Through the above results, it was confirmed that the effect of reducing gene expression involved in collagen synthesis was more excellent in combination than in single use, and specifically, it was confirmed that the effect of inhibiting fibrosis factors was more excellent in combination use at half the concentration (Nintedanib 50 nM, first component 5 ug / ml) of the existing effective concentration (Nintedanib 100 nM, first component 10 ug / ml) than in the effective concentration of each compound.

[0102]

[0103] Experimental Example 2: Antifibrotic Efficacy Animal Experiment

[0104] In experimental animals acclimatized for more than 5 days, 70 to 100 uL of BLM solution (bleomycin 1 to 3 mg / kg) was administered into the lungs using a catheter. Depending on the purpose of the test, the test drug was administered 7 days after BLM administration, and the drug was administered orally for 2 weeks until the 21st day of BLM administration. The composition of the experimental groups is as shown in Table 4 below, and the anti-fibrotic efficacy was measured by measuring body weight, SpO2, hydroxyproline, and inflammatory cell count.

[0105] GroupArticleNo. of Animals (Male)Dose 1Dose 2VolumeRoute of administration(mg / kg)(mg / kg) (SoC)(uL)G1(NC)Saline9N / AN / A100POG2(PC)Saline9N / AN / A100POG3(Test)First ingredient910N / A100POG4(Test)Second ingredient(NIN)9N / A60100POG5(Test)Second ingredient(PID)9N / A200100POG6(Test)First ingredient+Second ingredient(NIN)91060100POG7(Test)First ingredient+Second ingredient(PID)910200100PO

[0106]

[0107] Experimental Example 2-1: Lung Function Evaluation

[0108] This lung function evaluation is the most direct and important evaluation index that has the greatest impact on the symptoms and quality of life of patients with pulmonary fibrosis. It is an experiment that can most directly show the effect of improving lung function in an animal model of pulmonary fibrosis by measuring the oxygen concentration in the body.

[0109]

[0110] On the 21st day, SpO2 was measured through the abdomen of the mouse using a device (Berry, Veterinary Pulse Oximeter). The results are shown in Table 5 and Fig. 3.

[0111] NormalvehicleFirst component (10 mg / kg)Second component (NIN)(60 mg / kg)First component (10 mg / kg) + Second component (NIN)(60 mg / kg)Second component (PID)(200 mg / kg)First component (10 mg / kg) + Second component (PID)(200 mg / kg)Average value99.076.079.179.882.281.785.1Difference0.03.13.86.25.79.1Increase rate0%4%5%8%7%12%

[0112]

[0113] We attempted to determine the oxygen permeability of the lungs by measuring mouse abdominal SpO2, and confirmed that combined administration (PID and the first component, 12%) improved by more than 50% compared to single administration (PID, 7%). This result confirms that the direct improvement effect of combined administration on lung function is enhanced by increasing the oxygen permeability that is reduced during the pulmonary fibrosis process.

[0114]

[0115] Experimental Example 2-2: Measurement of Total Collagen in Lung

[0116] Pulmonary fibrosis is a condition in which collagen accumulates in the lungs, causing them to harden. The main cause of pulmonary fibrosis is the accumulation of collagen, and the degree of fibrosis progression can be predicted by measuring the collagen content inside the lung tissue.

[0117]

[0118] In this experimental example, the INSOLUBLE Collagen Assay (Biocolor, S2000) was used for analysis. After sacrifice on day 21, frozen lung tissue was pulverized with 100 μL of Fragmentation Reagent. Then, 100 μL of 37% HCl was added and incubated at 65°C for 3 hours. The tube contents were shaken every 30 minutes to promote tissue disintegration. After centrifugation, the concentration was adjusted to 100 μL and collagen staining was performed to prepare the sample. The absorbance was measured at 560 nm. The hydroxyproline value was measured as a ratio value compared to the normal group, and the results are shown in Table 6.

[0119] Day / nPBSBLMEngine 1 (10 mg / kg)Ingredient 2 (NIN) (60 mg / kg)Ingredient 2 (PID) (200 mg / kg)Ingredient 1 (10 mg / kg) + Ingredient 2 (NIN) (60 mg / kg)Ingredient 1 (10 mg / kg) + Ingredient 2 (PID) (200 mg / kg)21 / 988246228192156183120

[0120]

[0121] Through this experiment, it was confirmed that the content of collagen inside the lung was reduced when PID and the first component were used together compared to each other, and this indicates that the degree of pulmonary fibrosis was alleviated.

[0122]

[0123] Experimental Example 2-3: Histopathological (Ashcroft Score) Analysis

[0124] The degree of fibrosis and inflammation in lung tissue was visually observed under a microscope, and the degree of fibrosis in lung tissue was measured using the Fibrotic Index (FI), based on normalized criteria. A higher FI value indicates a more severe degree of fibrosis and disease, while a lower value indicates a more alleviated condition.

[0125]

[0126] On the 21st day, lung tissue was isolated and stained using H&E and MT stain, observed under a microscope at 200X magnification, and Ashcroft scoring (Hubner et al., 2008) was performed. The fibrosis index was calculated by dividing the sum of the modified Ashcroft field scores by the number of examined fields, and is shown in Table 7 and Figure 4.

[0127] Group Number of individuals Fibrotic Index (±SEM) Normal 90.5 (±0.4) Vehicle 95.5 (±0.6) Component 1 (10 mg / kg) 94.6 (±0.6) Component 2 (PID) (200 mg / kg) 93.7 (±0.7) Component 1 (10 mg / kg) + Component 2 (PID) (200 mg / kg) 91.9 (±0.5)

[0128]

[0129] The degree of fibrosis and inflammation of lung tissue was visually observed through a microscope, and the degree of fibrosis of lung tissue was measured using the Fibrotic Index according to normalized standards. As a result, it was confirmed that the combined administration of PID and the first component had a significantly improved effect compared to single administration.

[0130]

[0131] In addition, when comparing the microscopic image of the lung tissue of the vehicle in FIG. 5 with the microscopic image of the lung tissue of the subject administered the first and second components (PID) in FIG. 6, it can be confirmed that the lung tissue of the subject administered the combination has significantly improved inflammation and fibrosis in the lung tissue, even when seen with the naked eye, and is almost similar to normal lung tissue.

[0132]

[0133] Experimental Example 2-4: Inflammation cell count analysis

[0134] Since fibrosis is a chronic inflammatory disease with excessive collagen deposition, the infiltration of inflammatory cells was analyzed to determine the degree of inflammation in lung tissue.

[0135]

[0136] On the 21st day of administration, BALF (Bronchoalveolar lavage fluid) cells obtained through airway lavage of the mouse at sacrifice were diluted with 1.05 X PBS and attached to a slide. The slide was then dipped and removed in the order of 1, 2, and 3 diff quick stain solutions for 30 seconds each and stained. Counting was performed based on 500 cells. Macrophages are the largest in size and have mononuclei that stain blue. Neutrophils and eosinophils form multinucleated cells, but eosinophils are distinguished from neutrophils by their red eosin staining. Lymphocytes are small in size and mononucleated with very little cytoplasm. The total cells were counted and converted to a percentage, which is shown in Table 8 and Figure 7.

[0137] GroupnMacrophageEosinophilNeutrophilLymphocytetotal cell (10^4) PBS930003BLM9660221198Ingredient 1 (10 mg / kg)960017784Ingredient 2 (NIN) (60 mg / kg)953010569Ingredient 1 (10 mg / kg) + Ingredient 2 (NIN) (60 mg / kg)93006339Ingredient 2 (PID) (200 mg / kg)955012674Ingredient 1 (10 mg / kg) + Ingredient 2 (PID) (200 mg / kg)93808450

[0138]

[0139] To determine the extent of inflammation in lung tissue, an inflammatory cell analysis was performed. Compared to single-agent treatment, combined treatment showed a more than 50% improvement in total cell counts, indicating a significant improvement in lung tissue inflammation. Neutrophils, a key inflammatory cell associated with pulmonary fibrosis, were significantly reduced in number by more than 50%, demonstrating a significant improvement in the proportion of key inflammatory cells.

[0140]

[0141] Experimental Example 2-5: Measuring Changes in Body Weight

[0142] Body weight is an indirect indicator of the degree of improvement in the overall body condition of an animal model. If the degree of weight loss is small, it can be assumed that the overall body condition or symptoms of the disease have improved.

[0143]

[0144] Body weight was measured on days 0, 7, 14, 17, and 21 for three weeks, and then every three days starting on day 14, when the drug effect began to become apparent. The results are shown in Table 9 and Figure 8.

[0145] Day / nPBSBL Component 1 (10 mg / kg) Component 2 (NIN) (60 mg / kg) Component 2 (PID) (200 mg / kg) Component 1 (10 mg / kg) + Component 2 (NIN) (60 mg / kg) Component 1 (10 mg / kg) + Component 2 (PID) (200 mg / kg)00.00.00.00.00.00.00.073.2-10.1-10.1-10.1-10.2-12.1-13.0146.1-22.5-22.7-22.6-2 2.7-23.8-18.7179.3-24.8-22.2-20.0-18.1-21.5-17.12113.3-23.0-20.5-18.2-16.5-16.0-12.2

[0146]

[0147] It can be confirmed that the overall symptoms of the combination treatment group improved as the degree of weight loss was improved in the combination treatment group compared to the single treatment group.

[0148]

[0149] Experimental Example 3: Animal Pharmacokinetic / Pharmacodynamic Analysis

[0150] After a single oral administration of the first component in ICR mice, a kinetic study was conducted as shown in Table 10. The blood drug concentration over time obtained through LC-MS / MS was used to calculate pharmacokinetic parameters using Excel® and WinNonlin6.1 software, and the changes in the blood concentration of the first component are shown in Table 11 and Figure 9.

[0151] Experimental animal: ICR mice (male) Administration route: Oral Number of administrations: Single dose (mg / kg) 10 Administration amount (mL / kg) 10 Blood collection time (hr) 0.083, 0.25, 0.5, 1, 2, 4, 6, 8, 24 Excipient: Saline Number of test animals: 6 (n=3, cross-blood collection)

[0152]

[0153] Test substance 1st ingredient administration dose (mg / kg) 10t 1 / 2 (hr)2.50 ± 2.48T max (hr)2.67 ± 1.15C max (μg / mL)0.263 ± 0.032AUC 0-t (hr·μg / mL)1.07 ± 0.21AUCinf (hr·ng / mL)1190

[0154]

[0155] Manufacturing Example 1: Method for manufacturing an enteric-coated capsule containing the first ingredient

[0156] The main ingredient, in the hydrochloride form of the first ingredient, was filled into capsules according to dosage using Vcaps® enteric-coated capsules without any additional excipients.

[0157]

[0158] Manufacturing Example 2: Method for manufacturing an enteric-coated tablet containing the first ingredient

[0159] Microcrystalline cellulose, lactose monohydrate, crospovidone, and magnesium stearate were mixed with the hydrochloride form of the first component, and the mixture was compressed into a plate shape using a dry granulator, and then ground with an oscillator to produce dry granules. Microcrystalline cellulose, lactose monohydrate, and magnesium stearate were further mixed into the granules, and compression molded to produce tablets, which were then subjected to enteric coating to complete the process.

[0160]

[0161] Experimental Example 4: Human Pharmacokinetic / Pharmacodynamic Analysis

[0162] To confirm the safety / tolerance / pK in humans, a randomized, double-blind, placebo-controlled clinical trial was designed with 72 healthy adults as shown in Table 12, consisting of a single dose and a stepwise increase in dose. The formulation used in the clinical trial was prepared using the same method as in Manufacturing Examples 1 and 2 above.

[0163] Classification GroupActive ingredientDosage formAdministration methodSingle doseSAD1100 mgEnteric-coated capsuleSingle oral administration on an empty stomachSAD2300 mgSAD3600 mgSAD4500 mgEnteric-coated tabletMultiple doseMAD125 mgOral administration twice a day on an empty stomach for 13 consecutive days, then single administration on the morning of the 14th dayMAD250 mgMAD3100 mgMAD4200 mg

[0164]

[0165] The above clinical results did not reveal any significant abnormalities or side effects.

[0166]

[0167] As a result of confirming pK for a single administration, as can be seen in Table 13 and Figure 10 below, the maximum plasma concentration was recorded within 1.5 to 8 hours after administration, the geometric mean half-life was in the range of 6.91 to 9.90 hours, and it was confirmed that the plasma concentration increased in proportion to the dose.

[0168] Item (GCV%)Active ingredient 100 mg (N=6)Active ingredient 300 mg (N=6)Active ingredient 600 mg (N=6)Active ingredient 500 mg (N=6)C max (ng / mL)123.96 (37.72)276.80 (66.21)213.43 (433.75)513.56 (41.27)t max (h)*3.00 (1.50, 4.00)2.50 (1.50, 8.00)2.00 (1.50, 5.00)3.01 (0.50, 5.00)AUC 0-last (h*ng / mL)594.10 (61.23)1478.84 (48.07)1008.65 (625.00)2282.37 (54.92)AUC 0-∞ (h*ng / mL)639.07 (58.10)1525.99 (47.06)1922.64 (222.08)2339.20 (53.27)T 1 / 2 (h)6.91 (39.98)8.33 (24.57)9.90 (71.50)7.80 (13.49)Vz / F (L)1560.44 (34.98)2361.87 (41.96)4459.07 (84.14)2406.62 (48.04)CL / F (L / h)156.48 (58.10)196.59 (47.06)312.07 (222.08)213.75 (53.27)MRT (h)9.26 (33.69)9.75 (21.76)9.79 (25.60)9.57 (27.28)GCV% = Geometric coefficient of variationNotes: t max represented as median; min, max.

[0169]

[0170] In addition, as can be seen in Table 14 and Figure 11, the pK confirmation results on the 14th day for multiple administrations showed that the maximum plasma concentration was recorded within 1.5 to 5 hours at steady state after administration, the geometric mean half-life was in the range of 4.4 to 12.35 hours, and the geometric mean dosing interval period in vivo exposure (AUCτ) was in the range of 183.32-3012.35 h·ng / mL. It was confirmed that the plasma concentration had a linear relationship with the dose within the dose range of 25 mg to 200 mg of the active ingredient.

[0171] Item (GCV%) Active ingredient 25 mg T (N=6) Active ingredient 50 mg T (N=6) Active ingredient 100 mg T (N=6) Active ingredient 200 mg T (N=6)Day 14C max, ss (ng / mL)36.50 (66.19)102.61 (67.99)151.27 (65.36)595.35 (36.32)t max, ss (h)*2.00 (1.50, 4.00)3.00 (3.00, 5.00)3.00 (1.50, 4.00)2.00 (1.50, 5.00)AUCτ (h*ng / mL)183.32 (76.35)429.09 (128.63)617.40 (60.53)3012.35 (48.42)AUC 0-∞ (h*ng / mL)300.73 (102.79)565.34 (232.53)887.97 (60.55)4747.29 (64.90)T 1 / 2 (h)7.63 (46.84)4.40 (101.38)9.26 (22.69)12.35 (34.54)Vz / F ss (L)1433.79 (61.75)738.97 (34.90)2163.04 (63.58)1182.53 (48.47)CL / F ss(L / h)122.65 (79.49)116.53 (128.63)161.97 (60.53)66.39 (48.42)MRT (h)10.53 (31.65)8.44 (68.43)10.17 (9.35)12.07 (26.90)Accumulation ratio based on AUCτ1.94 (26.34)2.33 (31.42)1.88 (35.07)3.75 (36.01)Abbreviations: GCV% = Geometric coefficient of variation; n = Number of subjects; SD = Standard Deviation; T = Tablet.Notes: * t max, ss represented as median; min, max. MRT, AUCτ, CL / F ss , and Vz / F ss are considering a 12 hour dosing interval.

[0172]

[0173] Accordingly, the optimal pharmacokinetic model parameters that can explain the concentration changes at all doses observed in the blood drug concentrations obtained in the above human multiple-dose clinical trial were estimated using Excel® and WinNonlin8.1 software, and the exposure levels in the body in humans when various doses are administered orally were predicted. The main results are as shown in Table 15 below.

[0174] Dose (mg)Parameter50100150200T 1 / 2 (h)9.7531959.7531959.7531959.753195T max (h)2.8032.8032.8032.803C max (ng / mL)38.27176.5419114.813153.084CL / F (mL / hr)215856.7215856.7215856.7215856.5Vd / F (mL)3037295303729530372953037293AUC tau(hr*ng / mL)231.6352463.2704694.9055926.5413AUC tau _24hr (hr*ng / mL)463.2704926.54071389.8111853.08256

[0175]

[0176] From the rat experimental model experiment for pulmonary fibrosis of Experimental Example 3 above, 10 mg / kg administered once a day is the effective dose for rats, and the rat plasma AUC at the effective dose inf The value was confirmed to be 1190 hr·ng / mL. Therefore, the AUC is the same level as the effective dose in rats in humans. inf In order to predict the dose expected to show the value, the correlation with AUC according to the administered dose was examined as in Figure 12, and as a result, it was confirmed that 150 mg administration is necessary when administering twice a day to humans.

[0177]

[0178] Experimental Example 5: Confirming Drug Interactions

[0179] To evaluate drug interactions when the first and second components are administered together, a clinical trial was conducted with 48 healthy adults, divided into two groups of 24 each (Part 1 and Part 2). The trial was designed with a fixed sequence and three periods, and one 150 mg enteric-coated tablet of the first component was administered orally.

[0180]

[0181] Subjects enrolled in Part 1 sequentially received a single dose of 600 mg of the second component (PID) in Period 1, a single dose of 150 mg of the first component in Period 2, followed by a 3-day washout, multiple doses of 150 mg of the first component for 3 days, and a single combination of the first and second components (PID) in Period 3. Subjects enrolled in Part 2 sequentially received a single dose of 150 mg of the second component (NIN) in Period 1, multiple doses of 150 mg of the first component for 3 days in Period 2, and a single combination of the first and second components (NIN) in Period 3.

[0182]

[0183] After each administration, pK was observed for 24 hours, and adverse reactions were checked for 13-19 days.

[0184]

[0185] In Part 1, the results of the drug interaction evaluation showed that no change in the exposure of each drug was observed in each administration group, and thus the two drugs were evaluated as not having a significant effect on each other. In Part 2, it was confirmed that there was no clinically significant drug interaction when co-administered with the first component (NIN) compared to when administered alone.

[0186]

[0187] Experimental Example 6: Confirmation of the effect of combining the first component with pirfenidone / nintedanib.

[0188] To confirm the safety and efficacy of the first component in patients with idiopathic pulmonary fibrosis, a randomized, double-blind, placebo-controlled clinical trial will be conducted with either a standard treatment group or a non-administered group, as shown in Table 16. To confirm the safety and tolerability of the first component, the patients will be evaluated in comparison with the placebo after 24 weeks of administration of the first component. To confirm the therapeutic effect of the first component on idiopathic pulmonary fibrosis, the rate of decrease in forced vital capacity (FVC) from baseline for 24 weeks after 24 weeks of administration of the first component will be evaluated. Secondary efficacy endpoints include 1) time to IPF disease progression, including respiratory-related mortality or hospitalization, acute exacerbation of IPF, a relative decrease of ≥10% of the predicted value in FVC, and an absolute decrease of ≥15% of the predicted value in diffusing capacity for carbon monoxide (DLCO) adjusted for Hgb; 2) time to the first unplanned hospitalization for any cause during 24 weeks; 3) Baseline change in functional exercise capacity assessed by 6-minute walk test (6MWT) distance at 24 weeks; 4) Baseline change in DLCO (adjusted for hemoglobin) at 24 weeks; 5) Categorical assessment of absolute change from baseline in percent predicted value of FVC at 24 weeks; 6) Baseline change in quantitative chest high-resolution CT (HRCT) at 24 weeks; 7) Baseline change in patient-reported outcomes (PROs) measured by the St George's Respiratory Questionnaire, SGRQ, and Living with Idiopathic Pulmonary Fibrosis (L-IPF) at 24 weeks. Exploratory efficacy endpoints include 1) Baseline change in IPF-specific biomarkers at 24 weeks; and 2) Baseline change in blood biomarkers at 24 weeks.As safety evaluation variables, 1) incidence of adverse reactions after administration of the active ingredient; 2) physical examination; 3) 12-lead electrocardiogram test; 4) vital signs; 5) clinical laboratory performance tests are evaluated.

[0189]

[0190] The target group is divided into patients receiving pirfenidone as the existing standard treatment, patients receiving nintedanib, and patients receiving no other treatment. The first component or placebo is administered to each patient group to determine the efficacy, and the effect of co-administration with the standard treatment is compared to monotherapy. If adverse reactions due to the first component occur, the patient's response is closely monitored and the dosage is reduced.

[0191] Previously administered treatment agent 1 component 150 mg, BID 1 component 100 mg, BID placebo (0 mg), BID pirfenidone (mg, administered 3 times a day) 80 1 80 1 80 1 5 3 4 5 3 4 5 3 4 2 6 7 2 6 7 6 0 0 6 0 0 6 0 0 4 0 0 4 0 0 2 ...

Claims

1. 1) A first component of a compound represented by the following chemical formula 1, or a pharmaceutically acceptable salt thereof, and 2) A second component comprising any one of the following: a compound represented by the following chemical formula 2, a pharmaceutically acceptable salt thereof, a compound represented by the following chemical formula 3, and a pharmaceutically acceptable salt thereof. A pharmaceutical composition for preventing or treating fibrosis, The first component and the second component are administered together in the same formulation or in different formulations. Pharmaceutical composition: [Chemical Formula 1] [Chemical formula 2] [Chemical Formula 3] .

2. In paragraph 1, The above fibrosis may be caused by interstitial lung disease (ILD), scleroderma, keloid, hypertrophic scar, non-alcoholic fatty liver disease, primary sclerosing cholangitis (PSC), primary biliary cholangitis (PBC), diabetic retinopathy, age-related macular degeneration (AMD), hypertrophic cardiomyopathy, myocardial infarction, muscular dystrophy, diabetic kidney disease, focal segmental glomerulosclerosis (FSGS), or inflammatory bowel disease, IBD) Pharmaceutical composition.

3. In paragraph 2, The above interstitial lung disease is idiopathic pulmonary fibrosis (IPF), systemic sclerosis associated interstitial lung disease (SSc-ILD), or chronic fibrosing interstitial lung diseases with a progressive phenotype (PF-ILD). Pharmaceutical composition.

4. In paragraph 1, The weight ratio of the first component and the second component is 1:0.5 to 1:30, Pharmaceutical composition.

5. In paragraph 1, The second component is a compound represented by the chemical formula 2, or a pharmaceutically acceptable salt thereof, The weight ratio of the first component and the second component is 1:2 to 1:25, Pharmaceutical composition.

6. In paragraph 1, The second component is a compound represented by the chemical formula 3, or a pharmaceutically acceptable salt thereof, The weight ratio of the first component and the second component is 1:0.6 to 1:10, Pharmaceutical composition.

7. In paragraph 1, Containing 100 to 150 mg of the first component, Pharmaceutical composition.

8. In paragraph 1, The second component is a compound represented by the chemical formula 2, or a pharmaceutically acceptable salt thereof, Containing 100 to 150 mg of the first ingredient, Containing 200 to 800 mg of the second component, Pharmaceutical composition.

9. In paragraph 1, The second component is a compound represented by the chemical formula 3, or a pharmaceutically acceptable salt thereof, Containing 100 to 150 mg of the first ingredient, Containing 100 to 150 mg of the second component, Pharmaceutical composition.

10. In paragraph 1, The first and second components are each administered twice a day or three times a day, Pharmaceutical composition.

11. In paragraph 1, The first component is administered twice a day and the second component is administered three times a day, or the first component is administered three times a day and the second component is administered twice a day. Pharmaceutical composition.

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