Agent for inhibiting vascular hypertrophy

Abstract

To provide a new therapeutic agent that enables suppression of vascular hypertrophy, exemplified by peripheral pulmonary arterial stenosis in pulmonary hypertension.SOLUTION: An agent for inhibiting vascular hypertrophy comprising a CSF1R inhibitor is provided.SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to a vascular hypertrophy inhibitor and a pharmaceutical composition for preventing, treating, or ameliorating pulmonary arterial hypertension. [Background technology]

[0002] Pulmonary arterial hypertension is a rare disease, and despite recent advances in treatment, many intractable cases remain. Therefore, the development of new therapeutic agents is anticipated. Pulmonary arterial hypertension is a disease in which abnormal proliferation of pulmonary arterial endothelial cells and pulmonary arterial smooth muscle cells leads to peripheral pulmonary arterial stenosis and right heart failure. It is known that the mechanism is the accumulation of inflammatory cells such as macrophages around the pulmonary artery, which exacerbates pulmonary arterial remodeling via cytokines (Non-Patent Document 1), and macrophages play an important role in the pathogenesis of pulmonary hypertension. The present inventors have previously reported that bone marrow-derived inflammatory cells affect the pathology of pulmonary hypertension (Non-Patent Document 2). Therefore, we investigated the effect of treatment targeting macrophages infiltrating around the pulmonary artery on the improvement of pulmonary hypertension. [Prior art documents] [Non-patent literature]

[0003] [Non-Patent Document 1] Circ Res 2014;115:165-175 [Non-patent document 2] Nat Commun 2021;12:6177 Summary of the Invention [Problem to be solved by the invention]

[0004] An objective of the present invention is to provide a novel therapeutic agent capable of suppressing vascular hypertrophy, such as peripheral pulmonary artery stenosis, in pulmonary hypertension. [Means for solving the problem]

[0005] In order to solve the above problems, the present inventors created a rat model of monocrotaline (MCT)-induced pulmonary hypertension and examined the effectiveness of therapeutic agents targeting peripulmonary arterial macrophages. We observed that CD68-positive macrophages and Arginase 1-positive M2 macrophages accumulated around the pulmonary arteries in rat models of pulmonary hypertension compared with control groups. We also found that phosphorylation of CSF1R, a macrophage-specific receptor that regulates macrophage proliferation and differentiation into M2 macrophages, occurred in the pulmonary hypertension model. To clarify the role of CSF1R in pulmonary artery macrophages, we administered the CSF1R inhibitor pexidartinib to rat models of pulmonary hypertension. Surprisingly, pexidartinib significantly suppressed M2 macrophage accumulation and pulmonary artery smooth muscle cell proliferation compared with control groups. Even more surprisingly, CSF1R inhibition improved pulmonary hypertension in rat models of MCT-induced pulmonary hypertension. Thus, the present inventors have newly discovered that activation of CSF1R in macrophages is involved in pulmonary arterial hypertrophy in pulmonary hypertension, and that inhibition of CSF1R activation can improve pulmonary hypertension. The present invention was completed based on the above findings and includes the following aspects: One aspect of the present invention is [1] A vascular hypertrophy inhibitor comprising a CSF1R inhibitor. Here, the vascular hypertrophy inhibitor of the present invention is, in one embodiment, [2] The vascular hypertrophy inhibitor according to [1] above, The CSF1R inhibitor is characterized in that it acts on the accumulation and / or proliferation of M2 macrophages. In one embodiment, the vascular hypertrophy inhibitor of the present invention comprises [3] The vascular hypertrophy inhibitor according to [1] or [2] above, The CSF1R inhibitor is characterized in that it suppresses the proliferation of vascular smooth muscle cells. In one embodiment, the vascular hypertrophy inhibitor of the present invention comprises [4] The vascular hypertrophy inhibitor according to any one of [1] to [3] above, The CSF1R inhibitor is characterized in that it inhibits the kinase activity of CSF1R in macrophages. In one embodiment, the vascular hypertrophy inhibitor of the present invention comprises [5] The vascular hypertrophy inhibitor according to any one of [1] to [4] above, The CSF1R inhibitor is characterized in that it is pexidartinib or a pharmacologically acceptable salt thereof. Another aspect of the present invention is [6] A pharmaceutical composition for preventing, treating, or ameliorating pulmonary arterial hypertension, comprising the vascular hypertrophy inhibitor according to any one of [1] to [5] above.

[0006] Another aspect of the present invention is [7] A method for preventing, treating, or ameliorating pulmonary arterial hypertension, comprising: The present invention relates to a method comprising the step of administering a pharmaceutical composition containing a CSF1R inhibitor to a subject in need of prevention, treatment, or amelioration. In one embodiment, the method for treating pulmonary arterial hypertension of the present invention comprises: [8] The composition for treating pulmonary arterial hypertension according to [7] above, The CSF1R inhibitor is characterized in that it is pexidartinib or a pharmacologically acceptable salt thereof. Another aspect of the present invention is [9] A CSF1R inhibitor for use in the treatment of pulmonary arterial hypertension. In one embodiment, the CSF1R inhibitor of the present invention comprises

[10] The CSF1R inhibitor according to [9] above, The CSF1R inhibitor is characterized in that it is pexidartinib or a pharmacologically acceptable salt thereof. Another aspect of the present invention is

[11] The use of CSF1R inhibitors in the treatment of pulmonary arterial hypertension. In one embodiment, the use of the CSF1R inhibitor of the present invention comprises:

[12] Use of the CSF1R inhibitor according to

[11] above, The CSF1R inhibitor is characterized in that it is pexidartinib or a pharmacologically acceptable salt thereof. [Effects of the Invention]

[0007] The vascular hypertrophy inhibitor according to the present invention can inhibit the proliferation of smooth muscle cells in blood vessels, thereby inhibiting vascular hypertrophy. [Brief explanation of the drawings]

[0008] [Figure 1] FIG. 1 is a schematic diagram showing the schedule for producing a rat model of MCT-induced pulmonary hypertension and right heart catheterization performed in Example 1 below. [Figure 2] Figure 2 is a graph showing the results of right heart catheterization and measurement of right ventricular hypertrophy in the MCT-induced pulmonary hypertension model rats prepared in Example 1 below. The graph on the left of Figure 2 shows right ventricular pressure (mmHg). The graph on the right of Figure 2 shows the weight ratio of the right ventricle to the left ventricle and septum. [Figure 3] Figure 3 shows images of immunohistochemical staining (left panel) and immunofluorescent staining (center panel: anti-αSMA antibody, anti-CD68 antibody, DAPI; right panel: anti-αSMA antibody, anti-Arginase 1 antibody, DAPI) of lung tissue from a rat model of MCT-induced pulmonary hypertension prepared in Example 1 below. [Figure 4] FIG. 4 is an image showing the results of Western blotting of an extract from lung tissue of a rat model of MCT-induced pulmonary hypertension prepared in Example 1 below, using an anti-P-CSF1R antibody or an anti-CSF1R antibody and an anti-GAPDH antibody. [Figure 5] Figure 5 is an image showing the results of Western blotting performed in Example 2 below. The results are from Western blotting performed using an anti-P-CSF1R antibody or anti-CSF1R antibody and an anti-GAPDH antibody on an extract of RAW264.7 cells obtained by culturing in the presence of pexidartinib and colony-stimulating factor 1 (CSF1). [Figure 6] 6 shows images of immunofluorescence staining performed in Example 2 below. The images show RAW264.7 cells obtained by culturing in the presence of pexidartinib and colony-stimulating factor 1 (CSF1) stained with an anti-Ki67 antibody and DAPI. [Figure 7] FIG. 7 is a graph showing the ratio of the number of Ki67-positive cells obtained from the results of immunostaining performed in Example 2 below (the ratio to the number of Ki67-positive cells in the CSF1-free and pexidartinib-free group (Veh)). [Figure 8] FIG. 8 is a schematic diagram showing the schedule for administering a CSF1R inhibitor to a rat model of MCT-induced pulmonary hypertension performed in Example 3 below. [Figure 9] Figure 9 is a graph showing the results of right heart catheterization and right ventricular hypertrophy measurements in MCT-induced pulmonary hypertension model rats administered a CSF1R inhibitor, as performed in Example 3 below. The graph on the left in Figure 8 shows right ventricular pressure (mmHg). The graph on the right in Figure 8 shows the weight ratio of the right ventricle to the left ventricle and septum. [Figure 10] Figure 10 shows images illustrating the results of immunofluorescence staining in Example 3 below. The images show lung tissue from an MCT-induced pulmonary hypertension model rat administered with a CSF1R inhibitor, stained with an anti-αSMA antibody, an anti-CD68 antibody, and DAPI (top row), or stained with an anti-αSMA antibody, an anti-Arginase 1 antibody, and DAPI (bottom row). [Figure 11] 11 is an image showing the results of Western blotting in Example 3 below. The image shows the results of Western blotting, performed using an anti-P-CSF1R antibody or an anti-CSF1R antibody and an anti-GAPDH antibody, on an extract derived from lung tissue of an MCT-induced pulmonary hypertension model rat to which a CSF1R inhibitor had been administered. DETAILED DESCRIPTION OF THE INVENTION

[0009] 1. Vascular hypertrophy inhibitor One aspect of the present invention provides a vascular hypertrophy suppressant comprising a CSF1R inhibitor. The vascular hypertrophy inhibitor of the present invention can suppress vascular hypertrophy observed in patients with pulmonary arterial hypertension, etc. CSF1R inhibitors can suppress the accumulation and / or proliferation of M2 macrophages around blood vessels such as pulmonary arteries. Therefore, when a CSF1R inhibitor is administered to a patient with pulmonary arterial hypertension, the above-mentioned effects can suppress pulmonary artery hypertrophy and also right ventricular hypertrophy.

[0010] CSF1R is a receptor for colony-stimulating factor 1 (CSF1), which is essential for the differentiation of monocytic cells, and is a receptor tyrosine kinase present on the cell surface (colony-stimulating factor-1 receptor; CSF-1R). In the present invention, CSF1R particularly refers to CSF ​​expressed on the cell surface of macrophages. Upon binding of CSF1 to CSF1R, receptor molecules form dimers and the tyrosine residues between the receptors are autophosphorylated. This phosphorylation activates the tyrosine kinase activity of CSF1R, which activates intracellular signaling pathways. Activation of CSF1R in macrophages promotes macrophage differentiation and proliferation.

[0011] A CSF1R inhibitor refers to a substance that inhibits a function resulting from activation of CSF1R on the cell surface. Preferably, the inhibition of a function resulting from CSF1R activation is a temporary effect. Herein, the function resulting from CSF1R activation particularly refers to the function of promoting differentiation into M2 macrophages and the proliferation of macrophages. The inhibitor of a function resulting from CSF1R activation may be any substance that can inhibit the function in question, and may be any substance that can inhibit activation by CSF1R phosphorylation. Specifically, inhibitors include substances that bind to CSF1R and cause competitive inhibition or allosteric inhibition of CSF1, suppression of CSF1R expression, and inhibition of CSF1 production. CSF1R activation and its inhibition can be confirmed by measuring the phosphorylation state of CSF1R using techniques such as Western blotting. In another embodiment, inhibition of CSF1R activation involves suppressing the expression of CSF1R itself.

[0012] Examples of CSF1R inhibitors include antagonists that can inhibit CSF1R activation and siRNAs that suppress CSF1R expression. Antagonists include small molecule compounds, antibodies, lectins, etc. Suitable small molecule compounds include, but are not limited to, pexidartinib. In one embodiment, the CSF1R inhibitor is a drug that specifically inhibits CSF1R tyrosine kinase. Such a CSF1R inhibitor may also have the effect of inhibiting kinases other than CSF1R tyrosine kinase. The CSF1R inhibitor that can be used in this embodiment is not particularly limited as long as it is a drug that has the above-mentioned properties, but preferred examples include pexidartinib (Clin Cancer Res 2014, 20 (12): 3146-58) (CAS: 1029044-16-3), GW2580 (CAS 870483-87-7), and pharmacologically acceptable salts thereof.

[0013] A preferred pharmacologically acceptable salt is a hydrochloride salt (eg, pexidartinib hydrochloride).

[0014] In this specification, the term "blood vessel" is not limited to any particular blood vessel in a living body that may be hypertrophied. In a preferred embodiment, the blood vessel is an artery, such as a pulmonary artery, aorta, cerebral artery, coronary artery, or systemic artery.

[0015] As used herein, "vascular hypertrophy" refers to an increase in the thickness or diameter of a blood vessel wall, which is composed of the intima, media, and adventitia, compared to a normal state. The present invention is particularly directed to vascular hypertrophy accompanied by proliferation of smooth muscle cells present in the media.

[0016] "Suppressing vascular hypertrophy" means reducing the thickness or diameter of a blood vessel whose thickness or diameter has become larger than normal, or suppressing the thickness or diameter of the blood vessel wall from becoming larger than normal, or suppressing the degree of hypertrophy.

[0017] The CSF1R inhibitor used in the present invention has the effect of particularly suppressing the number of proliferated smooth muscle cells in enlarged blood vessels, or suppressing the abnormal proliferation of smooth muscle cells in enlarged blood vessels.

[0018] In one embodiment of the vascular hypertrophy inhibitor according to the present invention, the CSF1R inhibitor acts on the accumulation and / or proliferation of M2 macrophages. The CSF1R inhibitor used in the present invention can suppress the activation of CSF1R present on the cell surface of macrophages, thereby suppressing differentiation into M2 macrophages and the proliferation and / or accumulation of macrophages around blood vessels. Perivascular macrophages can be detected by immunofluorescent staining of tissue sections containing blood vessels using anti-CD68 antibodies, and M2 macrophages can be detected by immunofluorescent staining of tissue sections containing blood vessels using anti-Arginase 1 antibodies.

[0019] There are two types of macrophages: M1 and M2. M1 macrophages, also known as inflammatory macrophages, are involved in immune responses against pathogens such as bacteria and viruses. They secrete cytokines and inflammatory molecules to activate immune responses. On the other hand, M2 macrophages, also known as anti-inflammatory macrophages, suppress inflammation and promote tissue repair. M1 macrophages are known to differentiate from precursor cells or existing macrophages upon stimulation with inflammatory cytokines, while M2 macrophages are known to differentiate upon stimulation with anti-inflammatory cytokines or growth factors (e.g., CSF1).

[0020] In one embodiment of the vascular hypertrophy inhibitor according to the present invention, the CSF1R inhibitor inhibits the proliferation of vascular smooth muscle cells. The vascular hypertrophy inhibitor of the present invention can inhibit the proliferation of vascular smooth muscle cells and suppress vascular hypertrophy. The present inventors have found that a CSF1R inhibitor can inhibit the accumulation and / or proliferation of M2 macrophages around blood vessels and further inhibit the proliferation of vascular smooth muscle cells.

[0021] Vascular smooth muscle cells are smooth muscle cells present in the tunica media of blood vessel walls and play roles in vasoconstriction, vasodilation, and vascular repair. For example, abnormal proliferation of smooth muscle cells is observed in enlarged pulmonary arteries in patients with pulmonary arterial hypertension. As used herein, "inhibiting the proliferation of vascular smooth muscle cells" means inhibiting the abnormal proliferation of vascular smooth muscle cells to inhibit blood vessel hypertrophy, or inhibiting the number of proliferated vascular smooth muscle cells in hypertrophied blood vessels. Vascular smooth muscle cells can be evaluated by preparing tissue sections containing vascular regions and performing immunohistochemical staining or immunofluorescence staining using anti-αSMA antibodies.

[0022] 2. Pharmaceutical composition for pulmonary arterial hypertension In another aspect, the present invention provides a pharmaceutical composition comprising a vascular hypertrophy suppressant consisting of a CSF1R inhibitor, the pharmaceutical composition being for preventing, treating, or ameliorating pulmonary arterial hypertension. The pharmaceutical composition of the present invention has the effect of inhibiting abnormal proliferation of vascular smooth muscle cells and has the effect of preventing, treating, or ameliorating diseases in which abnormal proliferation of vascular smooth muscle cells is observed. In a preferred embodiment, the pharmaceutical composition of the present invention inhibits proliferation of vascular smooth muscle cells caused by M2 macrophages that accumulate or proliferate around blood vessels. An example of a disease in which abnormal proliferation of vascular smooth muscle cells is observed is pulmonary arterial hypertension.

[0023] As used herein, "prevention" refers to preventing or delaying the onset of a disease or condition in an individual, or reducing an individual's risk of developing a disease or condition. Additionally, "improvement" refers to improvement of a disease, symptom or condition, prevention or delay of worsening of a disease, symptom or condition, or reversal, prevention or delay of progression of a disease or symptom. Furthermore, "treatment" includes not only complete cure of a disease but also amelioration of symptoms.

[0024] As described above, the pharmaceutical composition of the present invention is a pharmaceutical composition for preventing, treating, or ameliorating pulmonary arterial hypertension. When the pharmaceutical composition of the present invention is used to treat pulmonary arterial hypertension, it can suppress pulmonary arterial hypertrophy and right ventricular hypertrophy in a subject. Right ventricular hypertrophy can be assessed by right heart catheterization and weight ratio. Right heart catheterization can be performed according to standard procedures, inserting a micrometer catheter into the right ventricle through the right external jugular vein and measuring right ventricular systolic pressure. Right ventricular mass can be assessed by calculating the weight ratio between the right ventricle and the left ventricle and septum.

[0025] The vascular hypertrophy inhibitor, pharmaceutical composition, and treatment method of the present invention can be applied to mammals including humans, such as mice, rats, guinea pigs, hamsters, rabbits, cats, dogs, sheep, pigs, cows, horses, goats, monkeys, and humans, and is preferably used in humans.

[0026] The vascular hypertrophy inhibitor and pharmaceutical composition of the present invention can be administered containing one or more pharmaceutically compatible ingredients. The pharmaceutically compatible ingredients can be appropriately selected from formulation additives and other ingredients commonly used in the field of drug or pharmaceutical composition production, depending on the dosage and administration concentration of the CSF1R inhibitor of the present invention. They can further contain appropriate carriers, excipients, and diluents commonly used in the production of drugs or pharmaceutical compositions, and can be used in dosage forms appropriate for the route of administration, such as powders, granules, tablets, pills, capsules, solutions, oils, suspensions, emulsions, syrups, and injections, according to conventional methods. Carriers, excipients, and diluents may include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, amorphous cellulose, polyvinylpyrrolidone, water, methyl hydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearate, and mineral oil.

[0027] The vascular hypertrophy inhibitor and pharmaceutical composition of the present invention may be administered via various routes, including oral, transdermal, subcutaneous, intradermal, intramuscular, intravenous, or intraperitoneal. A preferred dosage can be determined appropriately by those skilled in the art, taking into account various relevant factors such as the patient's age, sex, and weight, health condition, and severity of the disease.

[0028] The CSF1R inhibitor of the present invention can be orally administered to a human adult (60 kg) once or twice every 1 to 7 days, preferably once a day or twice a day. The kinase inhibitor used in the present invention can be orally administered at a single dose of 0.1 mg to 500 mg, preferably once a day. The kinase inhibitor of the present invention can be administered intravenously to a human once every 1 to 180 days, preferably once every 1, 2, 3, or 4 weeks. The CSF1R inhibitor used in the present invention can be administered intravenously at a single dose of 0.1 mg to 500 mg, preferably once a day.

[0029] When the CSF1R inhibitor used in the present invention is pexidartinib or a pharmacologically acceptable salt thereof, it can be orally administered at a dose of 62.5 mg, 125 mg, 250 mg, 1000 mg, 1500 mg, or 2000 mg per dose, preferably once every two days.

[0030] The present invention will be specifically described with reference to the following examples, but the present invention is not limited to these examples. Furthermore, these examples should not be construed as limiting in any sense. [Example]

[0031] Example 1. MCT-induced pulmonary hypertension model rats In this example, a rat model of MCT-induced pulmonary hypertension was prepared, and cardiac hypertrophy, pulmonary arterial wall hypertrophy, and CSF1R phosphorylation status of the model rats were evaluated.

[0032] 1-1 Creation of a rat model of MCT-induced pulmonary hypertension In this example, male Sprague-Dawley rats were given a single subcutaneous injection of MCT (60 mg / kg, Crotaline, C2401, Sigma-Aldrich) to create an MCT-induced pulmonary hypertension model rat. As a control, SD rats were administered saline instead of MCT. Right heart catheterization of the pulmonary hypertension model rats was performed 4 weeks after MCT administration (Figure 1).

[0033] 1-2. Right heart catheterization and evaluation of right ventricular hypertrophy Right heart catheterization was performed according to the following procedure. A micrometer catheter was inserted into the right ventricle through the right external jugular vein, and right ventricular systolic pressure was measured. The rats were then euthanized, and the heart and lungs were removed. To evaluate right ventricular hypertrophy, the weight ratio of the right ventricle to the left ventricle and septum was calculated. The results are shown in Figure 2. As shown in Figure 2, both the right ventricular pressure and the weight ratio of the right ventricle to the left ventricle and septum were significantly higher in the MCT-administered pulmonary hypertension model rats compared to the control group.

[0034] 1-3. Immunohistochemical and immunofluorescent staining Rat lung tissues were also fixed in 4% paraformaldehyde solution for paraffin embedding and subjected to immunohistochemical and immunofluorescent staining. For immunohistochemistry, slides were deparaffinized, rehydrated, and then antigen retrieval was performed using sodium citrate buffer. Slides were incubated with a primary antibody (anti-αSMA antibody) and then detected using an appropriate secondary antibody conjugated with peroxidase and diaminobenzidine. Samples were counterstained with hematoxylin and mounted in xylene. Elastica-Masson staining was also performed. For immunofluorescent staining, slides were deparaffinized, rehydrated, and then antigen retrieval was performed using sodium citrate buffer. Slides were incubated with a primary antibody (anti-αSMA antibody, anti-CD68 antibody, or anti-Arginase 1 antibody) and the appropriate secondary antibody. Nuclei were stained with DAPI. The results are shown in Figure 3. As shown in Figure 3, hypertrophy of the pulmonary artery wall accompanied by proliferation of smooth muscle cells was observed in the MCT-administered pulmonary hypertension model rat group compared to the control group. Furthermore, infiltration of macrophages around the pulmonary artery was observed in the MCT-administered pulmonary hypertension model rat group, and the macrophages were identified as M2-type macrophages because they were stained with anti-Arginase 1 antibodies.

[0035] Western Blot Western blot analysis was also performed to examine the phosphorylation status of CSF1 in rat lung tissue. Specifically, proteins were extracted from homogenized rat lung tissue. A portion of the proteins was subjected to SDS-polyacrylamide gel electrophoresis and transferred to a nitrocellulose membrane. The nitrocellulose membrane was incubated with a primary antibody (anti-P-CSF1R antibody or anti-CSF1R antibody) and an appropriate secondary antibody. Fluorescent immunoreactive bands were detected using the Odyssey CLX imaging system.

[0036] The results are shown in Figure 4. As shown in Figure 4, CSF1R expression was confirmed in both the MCT-administered pulmonary hypertension model rat group and the control group. Furthermore, an increase in phosphorylated CSF1R was confirmed in the MCT-administered pulmonary hypertension model rat group.

[0037] Example 2. Culture and treatment of RAW264.7 cells In this example, RAW264.7 cells, which are macrophage-like cells, were cultured in the presence of a CSF1R inhibitor and / or colony-stimulating factor, and the effects on CSF1R phosphorylation and cell proliferation activity were examined. 2-1.Cultivation of RAW264.7 cells RAW264.7 cells were cultured in Dulbecco's modified Eagle's medium containing 10% fetal bovine serum. Pexidartinib (colony-stimulating factor 1 inhibitor, HY-16749, MedChemExpress) and colony-stimulating factor 1 (CSF1, 416-ML, R&D Systems) were added to the culture medium. After one day of culture, RAW264.7 cells were subjected to Western blot or immunofluorescence staining.

[0038] Western Blot Western blotting was performed as in Example 1. The results of the Western blotting are shown in Figure 5. As shown in Figure 5, phosphorylation of CSF1R was confirmed in the group without addition of pexidartinib, and the addition of CSF1 increased the amount of phosphorylated CSF1R. On the other hand, in the group with addition of pexidartinib, phosphorylated CSF1R could not be confirmed, regardless of whether CSF1 was added or not.

[0039] 2-3. Immunofluorescence staining Immunofluorescence staining was performed as in Example 1, using anti-Ki67 antibody as the primary antibody. The results of immunofluorescence staining are shown in Figure 6, and the ratio of Ki67-positive cells obtained from the staining results (ratio to the number of Ki67-positive cells in the CSF1-free, pexidartinib-free group (Veh)) is shown in Figure 7. As shown in Figures 6 and 7, the number of Ki67-positive cells, which indicates cell proliferation activity, significantly increased in the CSF1-free, pexidartinib-free group. Furthermore, the number of Ki67-positive cells in the CSF1-free, pexidartinib-added group was not different from the CSF1-free, pexidartinib-free group. On the other hand, the number of Ki67-positive cells in the CSF1-free, pexidartinib-added group was not different from the CSF1-free, pexidartinib-free group. This indicates that pexidartinib significantly inhibited the increase in Ki67-positive cells caused by CSF1 addition.

[0040] Example 3: Treatment of pulmonary arterial hypertension animal models with pexidartinib In this example, the therapeutic effect of pexidartinib, a CSF1R inhibitor, was examined by administering it to a rat model of MCT-induced pulmonary hypertension. 3-1. Administration of CSF1R inhibitor to MCT-induced pulmonary hypertension rat model Two weeks after MCT injection, rats were randomly assigned to receive oral administration of pexidartinib (a colony-stimulating factor 1 inhibitor, 30 mg / kg, every 2 days, HY-16749, MedChemExpress) or vehicle (Veh) for two weeks (Figure 8). Right heart catheterization was then performed, and the rat lung tissues were subjected to Western blotting and immunofluorescence staining. Right heart catheterization, Western blotting, and immunofluorescence staining were performed as described in Example 1.

[0041] 3-2. Right heart catheterization and evaluation of right ventricular hypertrophy The results of right heart catheterization are shown in Figure 9. As shown in Figure 9, the right ventricular pressure and the weight ratio of the right ventricle to the left ventricle and septum were significantly higher in the MCT-administered pulmonary hypertension model rat group compared to the control group. Meanwhile, in the MCT-administered pulmonary hypertension model rat group, the right ventricular pressure and the weight ratio of the right ventricle to the left ventricle and septum were significantly reduced in the pexidartinib-administered group compared to the vehicle-administered group. These results indicate that pexidartinib was able to suppress the increase in right ventricular pressure and right ventricular hypertrophy in pulmonary arterial hypertension.

[0042] 3-3. Immunofluorescence staining The results of immunofluorescence staining are shown in Figure 10. As shown in Figure 10, in the vehicle-administered rat group of MCT-administered pulmonary hypertension model rats, pulmonary arterial wall hypertrophy accompanied by proliferation of smooth muscle cells was observed, and macrophage infiltration around the pulmonary arteries was observed. These macrophages were identified as M2-type macrophages because they were stained with anti-Arginase 1 antibodies. On the other hand, in the pexidartinib-administered rat group of MCT-administered pulmonary hypertension model rats, pulmonary arterial wall hypertrophy and macrophage accumulation around the pulmonary arteries were suppressed. These results indicate that pexidartinib has the effect of ameliorating pulmonary arterial hypertension.

[0043] The results of Western blot analysis are shown in Figure 11. As shown in Figure 11, no phosphorylation of CSF1R was observed in either the vehicle- or pexidartinib-treated control group. On the other hand, an increase in phosphorylated CSF1R was observed in the vehicle-treated MCT-induced pulmonary hypertension model rats compared to the control group. Furthermore, the increase in phosphorylated CSF1R was suppressed in the pexidartinib-treated MCT-induced pulmonary hypertension model rats compared to the control group.

Claims

1. A vascular hypertrophy inhibitor consisting of a CSF1R inhibitor.

2. The vascular hypertrophy inhibitor according to claim 1, A vascular hypertrophy inhibitor, wherein the CSF1R inhibitor acts on the accumulation and / or proliferation of macrophages.

3. The vascular hypertrophy inhibitor according to claim 1, The vascular hypertrophy inhibitor, wherein the CSF1R inhibitor inhibits the proliferation of vascular smooth muscle cells.

4. The vascular hypertrophy inhibitor according to claim 1, A vascular hypertrophy suppressant, wherein the CSF1R inhibitor inhibits the kinase activity of M2 macrophages against CSF1R.

5. The vascular hypertrophy inhibitor according to claim 1, A vascular hypertrophy inhibitor, wherein the CSF1R inhibitor is pexidartinib or a pharmacologically acceptable salt thereof.

6. A pharmaceutical composition for preventing, treating, or ameliorating pulmonary arterial hypertension, comprising the vascular hypertrophy inhibitor according to any one of claims 1 to 5.

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