Application of imidazole pyridine derivative in preparation of medicine for treating pulmonary fibrosis

Abstract

The invention belongs to the field of biological medicine, and particularly provides novel application of an imidazole pyridine derivative, and the chemical formula of the imidazole pyridine derivative is 4-{5 '-[3'-propyl-2 '-[3' '-(1' '-methyl) indolyl]-imidazole [4, 5-b] pyridine] group} morpholine. The invention provides an application of the imidazole pyridine derivative or the pharmaceutically acceptable salt thereof in preparation of a medicine for treating pulmonary fibrosis. When the compound disclosed by the invention is used for treating the pulmonary fibrosis, after animals with the pulmonary fibrosis are subjected to oral administration and inhalation administration, the lung function, the lung index, the lung tissue lesion and the fibrosis degree of model animals in different administration groups are obviously improved, so that an appropriate dosage of X22 has a treatment effect on the animals with the pulmonary fibrosis; and the pulmonary fibrosis improvement effect of the X22 inhalation way is obviously better than that of the X22 oral administration way and the existing conventional medicine.

Description

Technical Field

[0001] This invention relates to the field of biomedicine, specifically to novel uses of the small molecule compound 4-{5'-[3'-propyl-2'-[3”-(1”-methyl)indolyl]-imidazol[4,5-b]pyridyl]yl}morpholine. Background Technology

[0002] Pulmonary fibrosis is a chronic, diffuse interstitial lung disease clinically characterized by progressive dyspnea. Pathologically, it is characterized by the proliferation and activation of numerous fibroblasts, widespread extracellular collagen deposition, and extensive infiltration of inflammatory cells, leading to the destruction of normal lung tissue structure and abnormal scar hyperplasia. Currently, the choice of drugs for treating pulmonary fibrosis is limited, mainly oral pirfenidone and nintedanib. Although these can improve lung function to some extent and reduce the rate of acute exacerbations, they cannot stop disease progression or reverse the fibrotic areas of the lungs, and are accompanied by systemic side effects such as gastrointestinal discomfort. Inhaled administration allows the drug to directly target the lung lesions, significantly increasing local drug concentration and treatment efficiency, reducing systemic drug exposure, and lowering the incidence of adverse reactions. Therefore, the development of new drugs for treating pulmonary fibrosis via different routes of administration is increasingly urgent.

[0003] The small molecule compound 4-{5'-[3'-propyl-2'-[3”-(1”-methyl)indolyl]-imidazol[4,5-b]pyridyl]yl}morpholine, of formula (I), designated X22, is a previously reported compound. Its main reported functions include: application number 202010059361.9 discloses the use of X22 in the preparation of drugs for the treatment or prevention of kidney-related diseases; application number 2012104847041 discloses the use of X22 in the preparation of drugs for the treatment of inflammation-related diseases; and application number 202311228111.3 discloses its use in the preparation of drugs for the treatment or prevention of myeloproliferative neoplasms. However, no application of X22 in the preparation of drugs for the treatment of pulmonary fibrosis has been reported. Summary of the Invention

[0004] To address the shortcomings of existing technologies, this study reveals the potential application value of X22 in the treatment of pulmonary fibrosis by targeting the core regulatory targets of pulmonary fibrosis, including TGF-β1, MMP2, MMP9, and TIMP-1. It clarifies the target and molecular regulatory mechanism of X22 and provides new uses for it, mainly in the preparation of drugs for the treatment of pulmonary fibrosis.

[0005] The technical solution of this invention is: This invention discloses a novel use of an imidazopyridine derivative, the imidazopyridine derivative having the chemical formula: 4-{5'-[3'-propyl-2'-[3”-(1”-methyl)indolyl]-imidazo[4,5-b]pyridinyl]morpholine; and the use of the imidazopyridine derivative or a pharmaceutically acceptable salt thereof in the preparation of a drug for treating pulmonary fibrosis, wherein the drug is administered via inhalation.

[0006] Further disclosure is made of the use of pharmaceutical compositions containing the imidazopyridine derivative in the preparation of medicaments for treating pulmonary fibrosis.

[0007] Further preferably, the drug is an oral drug or an inhalation drug. In the examples, it is more preferably an inhalation drug.

[0008] Further preferably, the pharmaceutical composition containing the imidazopyridine derivative is formulated into various dosage forms such as tablets, capsules, oral liquids, injections, and powders.

[0009] Further preferably, the pharmaceutical composition containing the imidazopyridine derivative is formulated into various dosage forms such as aerosol, dry powder inhaler, and soft fog inhaler.

[0010] The imidazopyridine derivatives are shown in formula (I):

[0011] The pharmaceutical compositions used in this invention typically also contain a pharmaceutically acceptable carrier. A pharmaceutically acceptable carrier refers to a non-toxic filler, stabilizer, diluent, adjuvant, or other pharmaceutical excipient. Furthermore, the pharmaceutical compositions of this invention may further contain other excipients, such as flavoring agents, sweeteners, etc.

[0012] Pharmaceutically acceptable salts include those formed with inorganic acids, such as hydrochloric acid, hydrobromic acid, sulfuric acid, and phosphoric acid; those formed with organic acids, such as acetic acid, oxalic acid, tartaric acid, maleic acid, citric acid, and ascorbic acid; and those formed by elemental anions, such as chlorine, bromine, and iodine. "Pharmaceutical excipients" refers to conventional pharmaceutical carriers, including but not limited to any adjuvants, carriers, excipients, glidants, sweeteners, diluents, preservatives, dyes / colorants, flavor enhancers, surfactants, wetting agents, dispersants, suspending agents, stabilizers, isotonants, solvents, or emulsifiers approved by the U.S. Food and Drug Administration for acceptable use in humans or livestock.

[0013] Transforming growth factor β1 (TGF-β1) is a recognized key regulator in organ fibrosis. Furthermore, the sudden and excessive degradation and accumulation of the lung extracellular matrix (ECM) can lead to pulmonary fibrosis. MMP2, MMP9, and TIMP-1 are a group of specific enzymes that degrade the ECM, and an imbalance in their ratio is crucial for the occurrence and development of pulmonary fibrosis. Increased levels of MMP2 and MMP9 enhance ECM degradation, and this pathological degradation process triggers pulmonary fibrosis. This study investigated the effects of oral and inhaled X22 on a mouse model of pulmonary fibrosis, using nintedanib as a positive control. By observing serum TGF-β1 levels, lung function, lung index, lung tissue lesions and fibrosis severity, and the expression of MMP2, MMP9, and TIMP-1 proteins in lung tissue, the effects of oral and inhaled X22 on the treatment of pulmonary fibrosis were compared, and the mechanism of action of X22 in treating pulmonary fibrosis was explored.

[0014] This invention discloses the application of an imidazolpyridine derivative in the preparation of a drug for treating pulmonary fibrosis, (1) the effect of X22 on TGF-β1 in the serum of pulmonary fibrosis mice; (2) the effect of X22 on lung function of pulmonary fibrosis mice; (3) the effect of X22 on lung index of pulmonary fibrosis mice; (4) the effect of X22 on pathological changes in lung tissue of pulmonary fibrosis mice; (5) the effect of X22 on the level of TGF-β1 in the serum of pulmonary fibrosis mice; and (6) the effect of X22 on the expression levels of MMP13, MMP9, and TIMP-1 proteins in lung tissue of pulmonary fibrosis mice.

[0015] Compared with the prior art, the beneficial effects of the present invention are: 1. In the treatment of pulmonary fibrosis, the compounds of the present invention showed significant improvement in lung function, lung index, lung tissue lesions and degree of fibrosis in animal models after oral and inhalation administration via different routes of administration, suggesting that appropriate doses of X22 have a therapeutic effect on animals with pulmonary fibrosis.

[0016] 2. In the treatment of pulmonary fibrosis, the compounds of the present invention, when inhaled, show significantly better improvement in pulmonary fibrosis than when taken orally and with existing conventional drugs.

[0017] 3. In the treatment of pulmonary fibrosis, the compounds of this invention, X22, target the core regulatory network of pulmonary fibrosis through a dual mechanism: a. inhibiting the TGF-β1 signaling axis, directly blocking the excessive activation of the key pro-fibrotic factor TGF-β1, reducing fibroblast proliferation and collagen deposition, and inhibiting the fibrosis process; b. restoring the dynamic balance of ECM: simultaneously downregulating MMP2 / MMP9 and upregulating TIMP-1, changing the imbalance of ECM degradation-deposition, and preventing lung tissue structural destruction and abnormal scar hyperplasia.

[0018] 4. In the treatment of pulmonary fibrosis, the compounds of the present invention, X22, can exert a therapeutic effect on pulmonary fibrosis by regulating the levels of TGF-β1, MMP2, MMP9 and TIMP-1, and the inhalation route is more effective.

[0019] The detailed structure of the present invention will be further described below with reference to the accompanying drawings and specific embodiments. Attached Figure Description

[0020] Figure 1 A comparative graph showing the effect of X22 on serum FGF-B1 levels; Figure 2 A comparison chart showing the impact of X22 on FVC levels; Figure 3 A comparison chart showing the impact of X22 on FEV20 levels; Figure 4 A comparative graph showing the effect of X22 on the lung index; Figure 5 The images show pathological sections of lung tissue (HE, x200). Note: A. Normal control group; B. Model control group; C. Nintedanib group; D. X22 oral administration group; E. X22 inhalation administration group. Figure 6 This is a comparison chart showing the impact of X22 on pathology scores; Figure 7 The images show pathological sections of lung tissue (MASSON, X200). Note: A. Normal control group; B. Model control group; C. Nintedanib group; D. X22 oral administration group; E. X22 inhalation administration group. Figure 8 The graph shows the effect of X22 on the pathological score (MASSON). Figure 9 This is a comparative graph showing the effects of X22 on the expression levels of MMP13, MMP9, and TIMP-1 proteins in lung tissue. Detailed Implementation

[0021] The present invention is further illustrated in the following embodiments. These embodiments are merely for illustrating the technical effects of the present invention and are not intended to limit the scope of the invention.

[0022] 1. Materials 1.1 Laboratory Animals

[0023] Fifty SPF-grade C57BL / 6 mice, with 25 males and 25 females, weighing 10 - 12 g, were purchased from Hunan Slack Jingda Experimental Animal Co., Ltd.

SCXK(Xiang)2021 - 0002

SYXK(Xiang)2020 - 0015

[0024] 1.2 Main reagents and instruments

[0025] X22 was provided by the Pharmaceutical Research Department of Hunan Prima Pharmaceutical Research Center Co., Ltd. Nintedanib was purchased from Shanghai Macklin Biochemical Co., Ltd., batch number C13249029, and is applicable to the treatment of idiopathic pulmonary fibrosis (IPF).

[0026] HRH-MNE3026 type small animal single-concentration nasal and oral dynamic exposure inhalation toxicant delivery system (Beijing Huironghe Technology Co., Ltd.); human atomizing cup (Bertrand) PFT pulmonary function detection system (Shanghai Tawang Intelligent Technology Co., Ltd.), B203LED type biological microscope (produced by Chongqing Optoelec Instrument Co., Ltd.), RM2235 type paraffin slicer (produced by Leica, Germany), TP1020 type automatic dehydrator (produced by Leica, Germany), HI1220 type baking machine (produced by Leica, Germany), HI1210 type spreading machine (produced by Leica, Germany), EG1150H+C type tissue embedding machine (produced by Leica, Germany), DFC 420C type pathological imaging system (produced by Leica, Germany), etc.

[0027] 2 Methods 2.1 Preparation of pulmonary fibrosis model and drug treatment

[0028] Fifty qualified SPF-grade C57BL / 6 mice (half male and half female, weighing 10–12 g) were selected and randomly divided into two groups according to body weight: a normal control group (n=10) and a model group (n=40). The model group mice were injected intravenously with 20 mL / kg of bleomycin at a dose of 150 mg / kg to establish a mouse pulmonary fibrosis model. The normal control group mice were injected intravenously with an equal volume of 0.9% sodium chloride solution. Observations were conducted for 7 consecutive days. On day 8 of model establishment, 40 mice from each group were randomly divided into four groups according to sex and body weight: a model control group, a nintedanib group (39 mg / kg), an X22 oral administration group (10 mg / kg), and an X22 inhalation administration group (0.7 mg / kg). Before administration, nintedanib was prepared to the appropriate concentration using 0.5% CMC-Na and administered orally at a dose of 20 mL / kg. Animals in the X22 inhalation administration group inhaled X22 solution (22 μg / mL) continuously for 10 min. Animals in the normal control group and model control group inhaled 0.9% sodium chloride injection via mouth and nose for 10 min. The administration was once a day for 14 consecutive days.

[0029] 2.2 Detection Indicators (1) General clinical observation: During the administration period, observe the physiological state and mortality of the animals before and after administration.

[0030] (2) Detection of cytokines in serum: On the day after the last administration, 10 mice were taken from the normal control group, model control group, X22 oral administration group and X22 inhalation administration group. 0.5 mL of blood was collected from the jugular vein, centrifuged at 3000 r / min for 10 min, and the supernatant was collected. The content of FGF-β1 in mouse serum was detected by ELISA kit.

[0031] (3) Lung function test: On the day after the last administration, 10 mice from each group were anesthetized with 60 mg / kg of Supra-50, the trachea was surgically exposed, and the mice were fixed in a supine position in a volumetric recording box. A ventilation tube was inserted into the trachea, and the forced vital capacity (FVC) and forced expiratory volume in 20 ms (FEV1) of each group of mice were measured using a small animal lung function tester. 20 ).

[0032] (4) Lung index and lung tissue pathological examination: Ten mice from each group were taken the day after the last administration. After the lung function test, the mice were euthanized by cervical dislocation. The lungs were dissected and weighed to calculate the lung index, which is calculated as: lung wet weight (mg) / body weight (g). After weighing, the lungs were fixed in 10% neutral formalin, embedded in paraffin, sectioned, and stained with HE and MASSON. The pathological changes of the lung tissue were observed under a light microscope, and the degree of inflammation and hemorrhagic exudation, the degree of alveolar epithelial destruction, and the degree of inflammatory cell infiltration were scored. The specific scoring criteria are as follows: no obvious abnormality, scored as "0"; mild, scored as "1"; moderate, scored as "2"; severe, scored as "3".

[0033] (5) Expression levels of MMP13, MMP9, and TIMP-1 proteins in lung tissue: Four mice were taken from the normal control group, model control group, X22 oral administration group, and X22 inhalation administration group on the day after the last administration. 0.025g of lung tissue was taken, washed with ice-cold PBS, and 200μL of RIPA lysis buffer was added to the homogenizer and the tissue was repeatedly ground until no tissue block was visible. After lysis and centrifugation, protein quantification was performed. 50μg of total protein was loaded onto the sample, electrophoresed, transferred to a membrane, and incubated overnight at 4℃ with polyclonal primary antibody on a PVDF membrane. The ECL chemiluminescence method was used to expose and scan the image. The relative expression levels of MMP13, MMP9, and TIMP-1 proteins in the lungs of mice in each group were analyzed using Quantity One software, and the relative ratio of the expression of the corresponding proteins in each group to the internal reference GAPDH was calculated.

[0034] 2.3 Statistical Methods Statistical analysis was performed using SPSS 22.0 software. Quantitative data are expressed as mean ± standard deviation (SD). This indicates that comparisons among multiple samples were performed using one-way ANOVA. Based on the homogeneity of variance test, the LSD method and Tamhane's T2 analysis were used for homogeneous and heterogeneous variances, respectively. Ordinal data were presented as frequencies, and the rank-sum test for ordinal data was used for statistical analysis. P < 0.05 indicates that the difference is significant. P < 0.01 indicates that the difference is highly significant.

[0035] 3 Results

[0036] 3.1 General physical signs observation After modeling, all mice showed reduced spontaneous activity, piloerection, and rapid breathing. The above symptoms were still present in the X22 oral administration group, X22 inhalation administration group, and nintedanib group after administration, but the symptoms of each group were relieved to varying degrees in the later stage of administration.

[0037] 3.2 Effect on serum TGF-β1 levels like Figure 1As shown, compared with the normal control group, the serum TGF-β1 level in the model control group animals was significantly increased ( P <0.01); Compared with the model control group, the serum TGF-β1 level in mice in the X22 oral administration group and the X22 inhalation administration group was significantly reduced ( P < 0.05); Compared with the X22 oral administration group, the serum TGF-β1 level in the X22 inhalation administration group was significantly reduced ( P < 0.05).

[0038] 3.3 Effects on lung function like Figure 2 and Figure 3 As shown, compared with the normal control group, the model control group mice had lower FVC and FEV1. 20 Significantly reduced ( P <0.01); Compared with the model control group, the FVC and FEV of mice in the X22 oral administration group, X22 inhalation administration group and nintedanib group were significantly lower. 20 Significantly increased ( P < 0.01); compared with the nintedanib group and the X22 oral administration group, the FVC and FEV of mice in the X22 inhalation administration group were significantly lower. 20 Significantly increased ( P < 0.05).

[0039] 3.4 Effect on Lung Index like Figure 4 As shown, compared with the normal control group, the lung index of mice in the model control group was significantly increased ( P < 0.01); Compared with the model control group, the lung index of mice in the X22 oral administration group, X22 inhalation administration group and nintedanib group was significantly reduced ( P <0.05 or P < 0.01); compared with the nintedanib group and the X22 oral administration group, the lung index of mice in the X22 inhalation administration group was significantly reduced ( P < 0.05).

[0040] 3.5 Histopathological examination like Figure 5 As shown, under a microscope, no significant changes were observed in the lung tissue of the normal control group animals; the lung tissue of the model control group mice showed pathological changes such as alveolar wall thickening, inflammatory cell infiltration and alveolar congestion. The lung lesions of mice in the X22 oral administration group, X22 inhalation administration group and nintedanib group were all reduced to varying degrees compared with the model control group.

[0041] like Figure 6 As shown, compared with the normal control group, the lung tissue pathological score of mice in the model control group was significantly increased ( P<0.01); Compared with the model control group, the lung tissue pathological scores of mice in the X22 oral administration group, X22 inhalation administration group and nintedanib group were significantly reduced ( P < 0.01); Compared with the nintedanib group and the X22 oral administration group, the lung tissue pathological score of mice in the X22 inhalation administration group was significantly reduced ( P < 0.05).

[0042] like Figure 7 and Figure 8 As shown, compared with the normal control group, the MASSON score of lung tissue in the model control group mice was significantly increased ( P < 0.01); Compared with the model control group, the MASSON scores of lung tissue in mice in the X22 oral administration group, X22 inhalation administration group and nintedanib group were significantly reduced ( P < 0.01); Compared with the X22 oral administration group and the nintedanib group, the MASSON score of the lung tissue of mice in the X22 inhalation administration group was significantly reduced ( P < 0.05).

[0043] 3.6 Effects on the expression levels of MMP13, MMP9, and TIMP-1 proteins in lung tissue like Figure 9 As shown, compared with the normal control group, the levels of MMP13 and MMP9 in the lung tissue of mice in the model control group were significantly increased ( P < 0.05), TIMP-1 was significantly reduced ( P < 0.01); Compared with the model control group, the levels of MMP13 and MMP9 in the lung tissue of mice in the X22 oral administration group and the X22 inhalation administration group were significantly reduced ( P < 0.05 or P < 0.01), the TIMP-1 level in the lung tissue of mice in the X22 oral administration group and the X22 inhalation administration group was significantly increased ( P < 0.05); Compared with the X22 oral administration group, the levels of MMP13 and MMP9 in the lung tissue of mice in the X22 inhalation administration group were significantly reduced ( P < 0.05), lung tissue TIMP-1 levels were significantly elevated ( P <0.05).

[0044] The above description is a specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any equivalent substitutions or modifications made by those skilled in the art within the technical scope disclosed in the present invention, based on the technical solution and concept of the present invention, should be covered within the scope of protection of the claims of the present invention.

Claims

1. A new use of an imidazopyridine derivative of the formula: 4-{5'-[3'-propyl-2'-[3"-(l"-methyl)indolyl]-imidazo[4,5-b]pyridine]yl}morpholine; characterized in that, Use of the imidazopyridine derivative or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating pulmonary fibrosis.

2. The novel use according to claim 1, characterized in that, Use of a pharmaceutical composition containing the imidazopyridine derivative in the manufacture of a medicament for treating pulmonary fibrosis.

3. The use according to claim 1 or 2, wherein the compound is a compound of formula (I) ###0001### (I) or a pharmaceutically acceptable salt thereof. The medicament is a medicament for oral administration or a medicament for inhalation administration.

4. The novel use according to claim 1 or 2, characterized in that, The pharmaceutical composition containing the imidazopyridine derivative is prepared into tablets, capsules, oral solutions, injections, powders.

5. The use according to claim 1 or 2, wherein the compound is ###0002### The pharmaceutical composition containing the imidazopyridine derivative is prepared into various dosage forms such as aerosols, dry powder inhalants, soft mist inhalants, etc.

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