Pharmaceutical composition for treating cardiomyopathy

Abstract

The invention belongs to the technical field of biological medicines, and particularly relates to a compound concanavaline (L-canavaline) for treating cardiomyopathy, and the cardiomyopathy refers to all diseases causing myocardial structure and dysfunction, including secondary cardiac hypertrophy / heart failure and primary hypertrophic cardiomyopathy. The compound can be prepared into granules, capsules, powder, tablets and injection preparations together with another active ingredient which is different from the compound shown in the formula I and has the functions of enhancing the heart function, expanding peripheral blood vessels, reducing myocardial oxygen consumption and the like, and pharmaceutically acceptable auxiliary materials are added.

Description

Technical Field

[0001] The present invention belongs to the field of biomedical technology, and particularly relates to a therapeutic drug. Background Art

[0002] Myocardium-related diseases refer to all diseases that cause abnormal myocardial structure and function. According to the latest epidemiological survey data, it is estimated that the number of patients with myocardium-related diseases in China can reach up to 10 million, and the global prevalence rate is 1:200. Myocardium-related diseases are no longer rare diseases.

[0003] Myocardium-related diseases are closely related to high-risk outcomes, being the main cause of sudden cardiac death (SCD) and the leading cause of sudden death in young people, and an important cause of heart failure. The heredity of myocardium-related diseases determines their treatment needs. Behind a proband, there is often the treatment need of a family. The treatment need for myocardium-related diseases is gradually increasing. The main symptoms of myocardium-related diseases are myocardial hypertrophy and heart failure.

[0004] The treatment of myocardium-related diseases is divided into medical drug treatment and surgical treatment. Among them, medical drug treatment: in China, there are only beta blockers and non-dihydropyridine calcium channel blockers. At present, the drug treatments in China are all non-specific treatments, which cannot change the natural course of the disease, have limited treatment effects, poor treatment tolerance, and low patient satisfaction. Surgical treatment: includes ventricular septal resection, heart transplantation, etc. If the symptoms are severe, surgical treatment is generally recommended to improve the symptoms, and existing drugs can no longer achieve sufficient effects. Invasive treatments have many complications and are difficult for patients to accept. Moreover, invasive treatments have extremely high requirements for the operator and are difficult to implement in general hospitals. In the United States, Mavacamtem has been approved for the treatment of hypertrophic obstructive myocardium-related diseases, but it has not been introduced in China, and its indications are limited. Thus, it can be seen that the treatment means and drugs for myocardium-related diseases are limited and cannot meet the market demand. Summary of the Invention

[0005] Based on years of research on myocardium-related diseases, the inventor of the present invention has discovered the role of L-canavanine in the treatment of myocardium-related diseases, thus completing the present invention.

[0006] In the first aspect, the present invention provides a compound for treating myocardium-related diseases, and the structure of the compound is as follows:

[0007]

[0008] In the second aspect, the present invention provides the use of the compound of formula I in the preparation of a drug for treating myocardium-related diseases.

[0009] Furthermore, the myocardial-related diseases refer to a group of myocardial organic diseases caused by various reasons, which include abnormal cardiac mechanical activities and / or electrocardiogram dysfunctions. Pathologically, they are manifested as inappropriate ventricular dilation or hypertrophy, and affect the systolic or diastolic function of the heart, ultimately leading to heart failure, atrial or ventricular arrhythmia, or embolism.

[0010] Furthermore, the myocardial-related diseases include: 1) hypertrophic, dilated, arrhythmogenic, restrictive, and unclassified cardiomyopathies; 2) various primary and secondary myocardial hypertrophies; 3) heart failures caused by various reasons; 4) myocardial fibrosis.

[0011] In a third aspect, the present invention provides a pharmaceutical composition for treating myocardial-related diseases, and the pharmaceutical composition contains a compound of formula I.

[0012] Furthermore, the pharmaceutical composition further includes pharmaceutically acceptable excipients.

[0013] Furthermore, the excipients are pharmaceutical excipients that can enhance the efficacy, reduce toxicity, and / or mitigate side effects of the compound of formula I.

[0014] The pharmaceutical composition can be prepared into granule agents, capsule agents, powder agents, tablet agents, injection preparations, etc.

[0015] Furthermore, the pharmaceutical composition further contains another active ingredient different from the compound of formula I.

[0016] Furthermore, the another active ingredient includes, but is not limited to, β-blockers and non-dihydropyridine calcium channel blockers.

[0017] Furthermore, the β-blockers include, but are not limited to, propranolol, atenolol, bisoprolol, metoprolol, etc.

[0018] Furthermore, the non-dihydropyridine calcium channel blockers include, but are not limited to, verapamil, diltiazem, etc.

[0019] Furthermore, the another active ingredient can also be a drug that enhances cardiac function, dilates peripheral blood vessels, and reduces myocardial oxygen consumption. Description of the Drawings

[0020] Figure 1 is myocardial cell staining

[0021] Figure 2 is myocardial hypertrophy and the effects after intervention shown by echocardiogram and gross tissue

[0022] Figure 3 is WGA staining (4 weeks after surgery)

[0023] Figure 4It is Masson staining (4 weeks after surgery)

[0024] Figure 5 It is Masson staining (8 weeks after surgery)

[0025] Figure 6 It is the map of transcriptional changes in mouse myocardial tissue before and after treatment Specific implementation manners

[0026] The specific implementation manners of the present invention will be further described below. It should be noted here that the description of these implementation manners is used to help understand the present invention, but does not constitute a limitation to the present invention. In addition, the technical features involved in the following described implementation manners can be combined with each other as long as they do not conflict with each other.

[0027] The experimental methods in the following examples are all conventional methods unless otherwise specified, and the test materials used in the following examples are all commercially available through conventional commercial channels unless otherwise specified.

[0028] Example 1 Compound screening

[0029] The myocardial septum tissues of 269 HCM patients after Morrow surgery were subjected to RNA-seq sequencing with the myocardial septum tissues of 16 normal donors. The results of the mRNA expression changes of all measured genes were compared with the Connectivity map database, and Compound I was screened out. The gene expression profile changes after acting on the tool cell line were most similar to the gene expression profile changes of HCM patients compared with normal donors and had the possibility of being applied to HCM patients.

[0030] Example 2 In vitro myocardial cell hypertrophy model

[0031] Experimental method:

[0032] 2.1 Isolate primary neonatal rat cardiomyocytes (NRCM)

[0033] Take neonatal rats within 1 day after birth. After disinfecting the skin with 75% ethanol, cut open the chest cavity to take out the heart, remove connective tissues in cold HBSS, and wash until there is no blood residue. Use Pierce TMThe primary cardiomyocyte isolation kit (Thermo Fisher Scientific, USA) was used to digest the heart. After 35 minutes, the digestion was terminated with high-glucose DMEM medium containing 10% FBS (complete medium, the same below). The residual undigested tissue was filtered through a 100-mesh cell strainer, and the filtrate was collected. It was centrifuged at 1000G for 5 minutes at 26°C. After discarding the supernatant, the cells were resuspended in complete medium and transferred to a 10-cm dish for differential adhesion for 1.5 hours. After the adhesion was completed, the supernatant was taken, centrifuged at 1000G for 5 minutes at 26°C. After discarding the supernatant, the cells were resuspended in complete medium and evenly seeded onto a cell plate coated with 0.1% gelatin.

[0034] 2.2 NRCM culture and phenylephrine (PE) stimulation

[0035] After culturing in a 37°C constant temperature incubator for 30 hours, the medium was discarded and replaced with high-glucose DMEM medium without FBS (serum-free medium, the same below) for 16 hours. Then, the cells were divided into three groups for different treatments: the hypertrophy stimulation group, the drug group, and the control group. The hypertrophy stimulation group was replaced with serum-free medium containing 50 nM PE, the drug group was simultaneously replaced with serum-free medium containing 50 nM PE and 100 μM Compound I, and the control group was simultaneously replaced with fresh serum-free medium. After culturing for 24 - 48 hours, subsequent experiments were carried out.

[0036] 2.3 Total RNA extraction from adherent cells and real-time PCR verification

[0037] After discarding the medium from the adherently cultured NRCM, the cells were gently washed twice with PBS. An appropriate amount of Trizol (Thermo Fisher Scientific, USA) was added to the wells of the cell plate to digest the heart, and the cells were lysed on ice for 10 minutes. Then, after pipetting with a 1-ml pipette, the liquid was transferred into a 1.5-ml EP tube, shaken vigorously for 15 seconds, and then left standing for 5 minutes. Then, 200 μl of pre-cooled chloroform was added, stirred well, and placed on ice for 30 minutes. After centrifuging the supernatant at 15000G for 30 minutes at 4°C, it was carefully transferred to a new EP tube. After adding an equal volume of pre-cooled isopropanol, it was placed at -20°C overnight. The next day, it was centrifuged at 15000G for 30 minutes at 4°C, the supernatant was discarded, and the total RNA precipitate was obtained after washing twice with 75% ethanol. An appropriate amount of RNASE-free water was added to dissolve the RNA precipitate, and the concentration of RNA was measured using a Nanodrop 2000 spectrophotometer. Real-time PCR detection was carried out using a TB-Green PCR kit. The relative expression levels of the cardiomyocyte injury markers ANP and BNP in the three groups were normalized by the expression level of GAPDH, and the 2 -ΔΔ Ct method was used for calculation.

[0038] 2.4 Phalloidin cardiomyocyte staining

[0039] NRCM was cultured in a cell plate with cell culture inserts. After discarding the culture medium of adherent - cultured NRCM, the cells were gently washed twice with PBS and fixed with 4% paraformaldehyde at room temperature for 10 minutes. The cells were thoroughly washed 3 times with PBS, 5 minutes each time. The cells were permeabilized with 0.1% Triton X - 100 for 15 minutes. The cells were thoroughly washed 3 times with PBS, 5 minutes each time. The cells were stained with Alexa Fluor TM 488 phalloidin staining solution (Thermo Fisher Scientific, USA), and incubated in the dark at room temperature for 20 minutes. The cells were thoroughly washed 3 times with PBS, 5 minutes each time. The cell culture inserts were taken out, and a mounting medium containing DAPI was dropped on the glass slide, then the cell culture inserts were covered on the glass slide and stained for 5 minutes. Cell photos were taken using a laser confocal microscope, and the area of all cells in 10 random fields in each well was analyzed using Image J software to compare the cell areas of the three groups.

[0040] Experimental results:

[0041] Through the total RNA extraction of adherent cells and real - time PCR verification experiments, it was found that the expression levels of ANP and BNP in NRCM increased after PE stimulation, while Compound I could effectively reduce the increase of ANP and BNP in NRCM after PE stimulation, suggesting the alleviation of cardiomyocyte injury.

[0042] Through the phalloidin cardiomyocyte staining experiment, it was found that the cross - sectional area of NRCM cells increased after PE stimulation, while Compound I could effectively reduce the cross - sectional area of NRCM cells after PE stimulation, suggesting the therapeutic effect of the compound on cardiomyocyte hypertrophy, as Figure 1 shown.

[0043] Example 3 Mouse models of hypertrophic and myocardial - related diseases

[0044] Experimental method:

[0045] 3.1 Construction of mouse models of hypertrophic and myocardial - related diseases and drug intervention

[0046] Using the Crisper-Cas9 technology, the MYH6 R454C mutation and TNNT2 R127W mutation were introduced into mouse fertilized eggs to construct a Knock-in mouse model carrying pathogenic mutations related to human hypertrophic and myocardial diseases. Model (HET, the same below) mice and wild (WT) mice were raised until 12 weeks old before starting the experiment. The experiment was divided into four groups: the WT group, the HET + solvent group, the HET + Compound I group, and the HET + positive control group. Among them, the solvent was a 2% sodium carboxymethylcellulose solution, the concentration of Compound I was 200 mg / kg / d, and the positive control was Mavacamten (2.5 mg / kg / d). The administration method was gavage with a 12G needle. All interventions were performed once a day at a fixed time every day for a total of 4 weeks.

[0047] 3.2 Echocardiogram

[0048] At 0 weeks (before intervention), 1 week, 2 weeks, and 4 weeks (after the end of intervention), echocardiogram examinations were performed on all experimental mice. The chest and upper abdomen of the mice were depilated to fully expose the skin. After anesthesia with 3% isoflurane and maintenance with 2% isoflurane, the mice were fixed in the supine position on a platform. The chest was coated with coupling agent, and ultrasonic detection was performed under the MS400 probe. In the parasternal long-axis section, B-mode ultrasonic images were recorded; the probe was rotated clockwise by 90 degrees, that is, the left ventricular short-axis section, and the left ventricular motion was recorded with M-mode ultrasound. After the examination, the mice were returned to the cage. Using VIVO LAB software, the left ventricular end-diastolic anterior wall thickness, left ventricular end-systolic anterior wall thickness, left ventricular end-diastolic posterior wall thickness, left ventricular end-systolic posterior wall thickness, left ventricular end-diastolic internal diameter, and left ventricular end-systolic internal diameter were measured on the M-mode image. And the left ventricular ejection fraction was calculated by the software. When measuring each parameter, the values at 3 - 5 positions not affected by respiration were continuously measured. The differences in each parameter at each time point among the four groups were compared. Among them, after the echocardiogram at 0 weeks was completed, all HET mice were divided into the HET + solvent group, the HET + Compound I group, and the HET + positive control group according to the average left ventricular end-systolic posterior wall thickness.

[0049] 3.3 Mouse euthanasia and sample collection

[0050] After 4 weeks of intervention, the mice were euthanized, weighed, and the body weight was recorded. After removing the heart, it was placed in cold PBS, and the blood, excess adipose tissue, and connective tissue were removed. After slightly blotting the liquid on the surface of the heart, it was weighed, and the heart weight was recorded. Subsequently, it was photographed under a stereomicroscope. Three mice were randomly selected from each group, and the largest cross-sectional part containing the left and right ventricles was cut and fixed in 4% paraformaldehyde solution for subsequent pathological experiments. For the other heart tissues, the left ventricle was taken, longitudinally and evenly cut into four pieces, and then frozen in liquid nitrogen. The tibia of the mouse was cut, and after stripping the muscle tissue, the tibia length was measured with a vernier caliper, and the heart weight / body weight ratio and heart weight / tibia length ratio were calculated.

[0051] 3.4 Total RNA extraction from cardiac tissue and verification by real-time PCR

[0052] Take out 20 mg of cardiac tissue from liquid nitrogen, place it in a Roche centrifuge tube containing magnetic beads, and add 500 μl of Trizol (Thermo Fisher Scientific, USA). Grind it at 6500 rpm in a Roche tissue homogenizer for 15 seconds, cool it on an ice plate for 30 seconds, and repeat 2 - 3 times until no tissue chunks can be seen with the naked eye. Invert it up and down to mix well, and let it stand at room temperature for 15 minutes to fully lyse the tissue. Add chloroform and the subsequent steps are the same as described in 2.3. The real-time PCR steps are the same as described in 2.3.

[0053] 3.5 Paraffin sections of cardiac tissue

[0054] The fixed cardiac tissue after sampling is fixed in 4% paraformaldehyde solution for 48 h, then put into a tissue embedding cassette, marked, and rinsed with tap water for about 1 hour. Dehydrate it in an automatic dehydrator, and after completion, embed it in paraffin directionally and cool and solidify it on an ice table. Section it continuously with a microtome at a thickness of 5 μm, unfold and pick up the sections in water at 45 °C. After the water on the sections dries, store them at room temperature for standby.

[0055] 3.6 WGA staining

[0056] Bake the paraffin sections at 68 °C for 45 minutes, dewax them in xylene in an automatic stainer; hydrate them with gradient ethanol and place them in pure water. Pour 1000 ml of citrate antigen retrieval solution into a pressure cooker, boil it at 2400 W, place the tissue flat in the pot, cover the lid, boil it at 1000 W, release the gas smoothly and then boil for another 2 minutes. After completion, rinse the lid with tap water, open the lid after the pressure drops, cool it with tap water for 20 minutes, then take out the sections and place them in PBS. Block them with working sheep serum solution at room temperature for 1 hour. Wash them 3 times by horizontal shaking with PBS, 5 minutes each time. Dilute WGA with PBS at a ratio of 1:500, add 100 μl to each section, and incubate them at room temperature in a humidified box for 2 h. Wash them 3 times by horizontal shaking with PBS, 5 minutes each time, mount them with a mounting medium containing DAPI, and take pictures with a laser scanning confocal microscope. Use Image J software to analyze the area of all cardiomyocytes in 5 randomly selected fields on each section.

[0057] 3.7 Masson staining

[0058] The paraffin section was processed in the same way as in 3.5. After the dewaxing and hydration procedures were completed, it was soaked in potassium dichromate solution for 18 hours. Rinse with running water for 5 minutes, stain with hematoxylin solution for 5 minutes, and rinse with running water. Differentiate with 1% hydrochloric acid-ethanol solution for 2 s, and adjust the differentiation time accordingly according to the color of the cell nucleus under the microscope. Rinse with warm water to turn blue, and stain with ponceau for 5 minutes. Discard the ponceau, rinse with running water, and dry the water as much as possible. Stain with phosphomolybdic acid, and it can be replaced in the middle until the collagen appears pink under the microscope. Discard the liquid, wash with water, dry the water, stain with aniline blue for 16 - 20 s, and adjust the time specifically based on the change of the vascular intima to blue. Rinse with running water, dehydrate with absolute ethanol, and clear with xylene. Seal the section with neutral quick-drying glue in an automatic cover slip machine, and scan the whole section with a full-automatic digital section scanning system after drying in a fume hood. Randomly select 5 fields of view for each section, and use Image J software to analyze the fibrotic area.

[0059] Experimental results:

[0060] Through echocardiogram experiments, it was found that after 4 weeks of intervention with the compound of formula I, the degree of myocardial hypertrophy (determined by the left ventricular posterior wall thickness at the end of systole) in HET mice was reduced, significantly lower than that of the solvent group, and the therapeutic effect was similar to that of the positive control group, as Figure 2 shown.

[0061] After 4 weeks of intervention with the compound of formula I, through physiological indicators, it was found that the heart weight / body weight ratio and heart weight / tibia length ratio of HET mice were significantly lower than those of the solvent group, indicating a reduction in the degree of cardiac hypertrophy, and the effect was similar to that of the positive control group.

[0062] Through the experiments of total RNA extraction from heart tissues and real-time PCR verification, it was found that the expressions of myocardial ANP and BNP in HET mice were increased (compared with WT mice), and after the intervention with the compound of formula I, the expressions of myocardial ANP and BNP decreased, indicating a reduction in heart damage, and the effect was similar to that of the positive control group.

[0063] Through WGA staining experiments, it was found that the cross-sectional area of myocardial cells in HET mice was increased (compared with WT mice), and after the intervention with the compound of formula I, the cross-sectional area of myocardial cells decreased, indicating the therapeutic effect of the compound on myocardial cell hypertrophy, and the effect was similar to that of the positive control group, as Figure 3 shown.

[0064] Through Masson staining experiments, it was found that the cardiac fibrosis in HET mice was aggravated (compared with WT mice), and after the intervention with the compound of formula I, the fibrosis decreased, indicating the improvement effect of the compound on myocardial fibrosis, and the effect was similar to that of the positive control group, as Figure 4 shown.

[0065] Example 4 Secondary myocardial hypertrophy mouse model

[0066] Experimental method:

[0067] 4.1 Construction of a secondary myocardial hypertrophy mouse model and drug intervention

[0068] Eight-week-old mice were selected for aortic arch constriction (TAC) surgery. After hair removal, they were anesthetized with tribromoethanol and fixed on a thermostatic blanket. The neck skin was incised, the trachea was exposed, the muscle tissues on both sides of the trachea were lifted, and the incision was made along the sternum to the position of the 1st - 2nd ribs. The mouse was fixed with a chest spreader to expose the aortic arch as much as possible. A self-made pick needle with a 7G suture was used to pick through the aortic arch, the needle was removed leaving the thread, and a 25G needle was placed as a pad for ligation. After the operation, the incision was sutured and penicillin was administered. The mouse was rewarmed and revived on an electric blanket. After the operation, echocardiography was used to confirm the success of ligation. In the sham operation group, only the aortic arch was stirred without threading and ligation. The experiment was divided into four groups: the sham group, the TAC + solvent group, the TAC + compound I pre-administration group, and the TAC + compound I treatment group. Among them, the solvent was a 2% sodium carboxymethyl cellulose solution, the concentration of compound I was 200 mg / kg / d, and the administration method was gavage with a 12G needle. All interventions were performed once a day at a fixed time every day. The TAC + compound I pre-administration group started the intervention 1 week before TAC, resumed the intervention 3 days after the operation, and the postoperative intervention lasted for 8 weeks, with a total of 9 weeks of intervention. The TAC + compound I treatment group started the intervention 3 days after the operation, and the postoperative intervention lasted for 8 weeks. The intervention time lines of the sham group and the TAC + solvent group were the same as those of the TAC + compound I pre-administration group. Among them, 4 weeks after the operation was the detection point for myocardial hypertrophy phenotype, and 8 weeks after the operation was the detection point for heart failure phenotype.

[0069] 4.2 Echocardiography

[0070] Echocardiography was performed on all experimental mice at 1 week before the operation (-1 week), before the operation (0 week), 3 days after the operation, 4 weeks after the operation, and 8 weeks after the operation. Among them, the methods at 1 week before the operation (-1 week), before the operation (0 week), 4 weeks after the operation, and 8 weeks after the operation were the same as those described in 3.2. Three days after the operation, the blood flow velocity of the aortic arch was additionally measured using Doppler blood flow spectrum. A significant acceleration of the blood flow velocity indicated successful ligation. Mice with failed ligation were euthanized, and mice with successful ligation were evenly divided into the TAC + solvent group and the TAC + compound I treatment group according to the average left ventricular posterior wall thickness at the end of systole.

[0071] 4.3 Euthanasia and tissue sampling of mice

[0072] The mice were euthanized 4 weeks and 8 weeks after the intervention, and the tissue sampling and subsequent experimental protocols were the same as those in 3.3. Among them, half of the mice were sacrificed at 4 weeks, and it was necessary to ensure that the parameters of the sampled tissues and the remaining mice in each group were consistent.

[0073] 4.4 Total RNA extraction from heart tissue and real-time PCR verification

[0074] The total RNA extraction and real-time PCR procedures were the same as described in 3.4.

[0075] 4.5 Paraffin sections of heart tissue

[0076] The paraffin section protocol was the same as 3.5.

[0077] 4.6 WGA staining

[0078] The WGA staining protocol was the same as 3.6.

[0079] 4.7 Masson staining

[0080] The Masson staining protocol was the same as 3.7.

[0081] Experimental results:

[0082] Through echocardiogram experiments, it was found that at 4 weeks after surgery (the myocardial hypertrophy phenotype detection point), the degree of myocardial hypertrophy (measured by the left ventricular posterior wall thickness at end-systole) in the TAC + solvent group mice was significantly higher than that in the sham group, while the degree of myocardial hypertrophy in both the TAC + Compound I pre-treatment group and the TAC + Compound I treatment group mice was significantly lower than that in the TAC + solvent group mice, suggesting the preventive and therapeutic effects of the compound on myocardial hypertrophy. In addition, at 8 weeks after surgery (the heart failure phenotype detection point), the cardiac function (measured by the left ventricular ejection fraction) of the TAC + solvent group mice was significantly lower than that in the sham group, while the cardiac function of both the TAC + Compound I pre-treatment group and the TAC + Compound I treatment group mice was significantly better than that in the TAC + solvent group mice, suggesting the preventive effect of the compound on heart failure.

[0083] At 4 weeks after surgery (the myocardial hypertrophy phenotype detection point) and 8 weeks after surgery (the heart failure phenotype detection point), through physiological indicators, it was found that the heart weight / body weight ratio and heart weight / tibia length ratio of the TAC + solvent group mice were significantly higher than those in the sham group, while the heart weight / body weight ratio and heart weight / tibia length ratio of both the TAC + Compound I pre-treatment group and the TAC + Compound I treatment group mice were significantly lower than those in the TAC + solvent group mice, suggesting a reduction in the degree of cardiac hypertrophy and enlargement.

[0084] Through the total RNA extraction and real-time PCR verification experiments of heart tissue, it was found that at 4 weeks after surgery (the myocardial hypertrophy phenotype detection point) and 8 weeks after surgery (the heart failure phenotype detection point), the expression levels of myocardial ANP and BNP in the TAC + solvent group mice were significantly higher than those in the sham group, while the expression levels of myocardial ANP and BNP in both the TAC + Compound I pre-treatment group and the TAC + Compound I treatment group mice were significantly lower than those in the TAC + solvent group mice, suggesting the improvement effect of the compound on myocardial injury.

[0085] Through WGA staining experiments, it was found that at 4 weeks after surgery (the detection point for myocardial hypertrophy phenotype) and 8 weeks after surgery (the detection point for heart failure phenotype), the cross-sectional area of cardiomyocytes in the TAC + solvent group mice increased (compared with the sham group), while the cross-sectional area of cardiomyocytes in both the TAC + Compound I pre-administration group and the TAC + Compound I treatment group mice was significantly smaller than that in the TAC + solvent group mice, suggesting the alleviating effect of the compound on cardiomyocyte hypertrophy.

[0086] Through Masson staining experiments, it was found that at 4 weeks after surgery (the detection point for myocardial hypertrophy phenotype) and 8 weeks after surgery (the detection point for heart failure phenotype), myocardial fibrosis in the TAC + solvent group mice was aggravated (compared with the sham group), while myocardial fibrosis in both the TAC + Compound I pre-administration group and the TAC + Compound I treatment group mice was significantly less than that in the TAC + solvent group mice, suggesting the improving effect of the compound on myocardial fibrosis, as Figure 5 shown.

[0087] Myocardial Transcriptomic Characterization before and after Treatment in Example 5

[0088] Experimental Method:

[0089] 5.1 RNA-seq Sequencing of Mouse Myocardial Tissue after Treatment

[0090] The mouse myocardial tissue obtained in 4.4 was used for mRNA transcriptomic sequencing. There were 4 cases in each of the three groups: the sham group, the TAC + solvent group, and the TAC + Compound I treatment group, with a total of 12 cases.

[0091] 5.2 Functional Enrichment Analysis of RNA-seq Results

[0092] Transcriptomics defined several genes that were upregulated in the TAC + solvent group compared with the sham group, downregulated in the TAC + Compound I treatment group compared with the TAC + solvent group, or downregulated in the TAC + solvent group compared with the sham group and upregulated in the TAC + Compound I treatment group compared with the TAC + solvent group. Subsequently, GO functional enrichment was performed on the above selected differential genes to obtain the main functional annotations of the differential genes, suggesting the potential therapeutic targets of Compound I.

[0093] Experimental Results:

[0094] Through sequencing and subsequent functional enrichment analysis, we can know that the therapeutic effect of Compound I is mainly reflected in regulating the cell cycle of cardiomyocytes.

Claims

1. A compound for treating cardiomyopathy, characterized in that, The compound is a compound of formula I 2. Use of a compound of formula I in the preparation of a medicament for treating cardiomyopathy, the structure of the compound of formula I being as follows 3. Use of the compound of formula I as claimed in claim 2 in the preparation of a medicament for treating cardiomyopathy, characterized in that, The cardiomyopathy includes dilated cardiomyopathy, hypertrophic cardiomyopathy, restrictive cardiomyopathy, arrhythmogenic cardiomyopathy, undefined cardiomyopathy and cardiac fibrosis.

4. A pharmaceutical composition for treating cardiomyopathy, characterized in that, The pharmaceutical composition contains a compound of formula I.

5. The pharmaceutical composition according to claim 4, wherein The pharmaceutical composition includes pharmaceutically acceptable excipients.

6. The pharmaceutical composition according to claim 5, wherein The excipients are pharmaceutical excipients that enhance the efficacy, reduce the toxicity and / or alleviate the toxic and side effects of the compound of formula I.

7. The pharmaceutical composition according to any one of claims 4 to 6, characterized in that The pharmaceutical composition can be prepared into granules, capsules, powders, tablets or injection preparations.

8. The pharmaceutical composition according to any one of claims 4-7, characterized in that, The pharmaceutical composition further contains another active ingredient different from the compound of formula I.

9. The pharmaceutical composition according to claim 8, characterized in that, The another active ingredient includes, but is not limited to, one or more of β-blockers and non-dihydropyridine calcium channel blockers.

10. The pharmaceutical composition according to claim 8, characterized in that, The another active ingredient can also be a drug that enhances cardiac function, dilates peripheral blood vessels and reduces the oxygen consumption of the myocardium of the pharmaceutical composition.

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