Application of hydroxyurea in preparation of medicine for preventing and / or treating autoimmune myocarditis

Abstract

The invention discloses an application of hydroxyurea in preparation of a medicine for preventing and / or treating autoimmune myocarditis, hydroxyurea or a pharmaceutically acceptable derivative of hydroxyurea is applied to treatment of autoimmune myocarditis for the first time, and experimental verification of an autoimmune mouse model shows that hydroxyurea or a pharmaceutically acceptable derivative of hydroxyurea can be used for preparing a medicine for preventing and / or treating autoimmune myocarditis. The hydroxyurea can significantly reduce heart loss and inflammation index levels in plasma in an autoimmune myocarditis mouse model, significantly inhibit expression of inflammatory factors and reduce inflammatory cell infiltration of heart tissues, and it is verified that hydroxyurea can effectively improve and treat autoimmune myocarditis. Meanwhile, the safety of the medicine is detected, and obvious hepatorenal toxicity of the hydroxyurea to mice is not found. In addition, experiments also find that the application of hydroxyurea can also effectively improve cardiac hypofunction caused by inflammation. Therefore, the hydroxyurea has a huge application prospect as a medicine for reducing inflammatory response and treating myocarditis.

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to application of hydroxyurea in preparing medicines for preventing and or treating autoimmune myocarditis. Background Art

[0002] Myocarditis is an inflammatory disease of the myocardium caused by a variety of reasons, often accompanied by cardiac dysfunction. Its causes include viral infection, immune response, drug toxicity and other external stimuli. Clinically, myocarditis can manifest as acute chest pain, heart failure, arrhythmia and other symptoms, and severe cases can lead to cardiogenic shock or sudden death. The complications of myocarditis are diverse, the most common of which are acute heart failure, arrhythmia and long-term cardiac dysfunction. Some patients may develop dilated cardiomyopathy and eventually heart failure. In addition, myocarditis can also cause structural and functional changes in the heart, increasing the risk of serious cardiovascular events in patients. Current drug treatments are mainly antiviral therapy, immunomodulatory therapy and supportive therapy. For viral myocarditis, antiviral drugs are often used to inhibit pathogen replication; for immune-mediated myocarditis, immunosuppressants and steroids are often used to reduce inflammatory responses. However, existing treatments still have certain limitations, some patients are ineffective with existing treatments, and the side effects of drugs are large. Therefore, it is of great significance to find new therapeutic drugs.

[0003] Hydroxyurea is a non-alkylated anti-tumor and antiviral drug widely used in clinical practice. It was first used to treat leukemia and sickle cell anemia. Its main mechanism of action is to inhibit the synthesis of nucleotides, interfere with DNA replication, and then inhibit cell proliferation. In addition, hydroxyurea also has an immunomodulatory effect, which can inhibit inflammatory reactions and relieve pain and discomfort caused by splenomegaly, which makes it a potential application prospect in the treatment of various diseases. Studies have shown that hydroxyurea can produce certain therapeutic effects on diseases such as atherosclerosis and diabetic heart disease by regulating the immune system and improving vascular function. However, no relevant experimental studies and clinical applications of hydroxyurea in the prevention and / or treatment of myocarditis have been found in the prior art. Summary of the invention

[0004] Therefore, based on the above background, the present invention provides the use of hydroxyurea in the preparation of a drug for preventing and or treating autoimmune myocarditis, providing a new direction for the treatment of autoimmune myocarditis.

[0005] The technical solution of the present invention is as follows:

[0006] The use of hydroxyurea as shown in structural formula (I), or a pharmaceutically acceptable derivative of hydroxyurea as shown in structural formula (I) in the preparation of a drug for preventing and / or treating autoimmune myocarditis,

[0007] The structural formula (I) is shown below:

[0008]

[0009] Furthermore, the pharmaceutically acceptable derivative of hydroxyurea represented by structural formula (I) is one of the pharmaceutically acceptable salts, hydrates, solvates, enantiomers, stereoisomers or tautomers of hydroxyurea represented by structural formula (I).

[0010] Furthermore, in the above medicine, the hydroxyurea represented by the structural formula (I) or the pharmaceutically acceptable derivative of the hydroxyurea represented by the structural formula (I) is the only active ingredient.

[0011] Based on the same inventive concept, the present invention also provides a drug for preventing and / or treating autoimmune myocarditis, which comprises at least one of hydroxyurea as shown in structural formula (I) and a pharmaceutically acceptable derivative of hydroxyurea as shown in structural formula (I).

[0012] Furthermore, the pharmaceutically acceptable derivative of hydroxyurea represented by structural formula (I) is at least one of the pharmaceutically acceptable salts, hydrates, solvates, enantiomers, stereoisomers or tautomers of hydroxyurea represented by structural formula (I).

[0013] Furthermore, the drug also contains pharmaceutically acceptable excipients, which include at least one of a diluent, a binder, an absorbent, a disintegrant, a dispersant, a wetting agent, a solubilizing agent, a buffer, and a surfactant.

[0014] Furthermore, in the above-mentioned drug, the hydroxyurea represented by the structural formula (I) or the pharmaceutically acceptable derivative of the hydroxyurea represented by the structural formula (I) is the only active ingredient.

[0015] Based on the same inventive concept, the present invention also provides the use of hydroxyurea as shown in structural formula (I), or a pharmaceutically acceptable derivative of hydroxyurea as shown in structural formula (I) in the preparation of a drug for preventing and / or treating cardiac dysfunction caused by inflammation.

[0016] Furthermore, the pharmaceutically acceptable derivative of hydroxyurea represented by structural formula (I) is one of the pharmaceutically acceptable salts, hydrates, solvates, enantiomers, stereoisomers or tautomers of hydroxyurea represented by structural formula (I).

[0017] Furthermore, in the above-mentioned drug, the hydroxyurea represented by the structural formula (I) or the pharmaceutically acceptable derivative of the hydroxyurea represented by the structural formula (I) is the only active ingredient.

[0018] Based on the same inventive concept, the present invention also provides a drug for preventing and / or treating cardiac dysfunction caused by inflammation, which comprises at least one of hydroxyurea as shown in structural formula (I) and a pharmaceutically acceptable derivative of hydroxyurea as shown in structural formula (I).

[0019] Furthermore, the pharmaceutically acceptable derivative of hydroxyurea represented by structural formula (I) is at least one of the pharmaceutically acceptable salts, hydrates, solvates, enantiomers, stereoisomers or tautomers of hydroxyurea represented by structural formula (I).

[0020] Furthermore, in the above-mentioned drug, the hydroxyurea represented by the structural formula (I) or the pharmaceutically acceptable derivative of the hydroxyurea represented by the structural formula (I) is the only active ingredient.

[0021] Furthermore, the drug also contains pharmaceutically acceptable excipients, which include at least one of a diluent, a binder, an absorbent, a disintegrant, a dispersant, a wetting agent, a solubilizing agent, a buffer, and a surfactant.

[0022] The beneficial effects achieved by the present invention are as follows:

[0023] The present invention is the first to apply hydroxyurea or a pharmaceutically acceptable derivative of hydroxyurea to the treatment of autoimmune myocarditis. After experimental verification of an autoimmune mouse model, hydroxyurea can significantly reduce the levels of cardiac damage and inflammatory indicators in the plasma of the autoimmune myocarditis mouse model, and significantly inhibit the expression of inflammatory factors, and reduce the infiltration of inflammatory cells in the heart tissue, verifying that hydroxyurea can effectively improve and treat autoimmune myocarditis. At the same time, the safety of the drug was tested, and no obvious liver and kidney toxicity of hydroxyurea to mice was found.

[0024] In addition, experiments have also found that the use of hydroxyurea can effectively improve the heart function impairment caused by inflammation. Therefore, as a drug for reducing inflammatory response and treating myocarditis, hydroxyurea shows great application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Attached Figure 1 The regulatory effect of hydroxyurea on aspartate aminotransferase (AST), an indicator of heart damage in the blood of mice model of autoimmune myocarditis;

[0026] Attached Figure 2 The regulatory effect of hydroxyurea on the inflammatory factor interleukin-1β (IL-1β) in the blood of a mouse model of autoimmune myocarditis;

[0027] Attached Figure 3 The regulatory effect of hydroxyurea on the inflammatory factor interleukin-18 (IL-18) in the blood of a mouse model of autoimmune myocarditis;

[0028] Attached Figure 4 The regulatory effect of hydroxyurea on the inflammatory factor tumor necrosis factor-α (TNF-α) in the blood of a mouse model of autoimmune myocarditis;

[0029] Attached Figure 5 The regulatory effect of hydroxyurea on the inflammatory factor interleukin-10 (IL-10) in the blood of a mouse model of autoimmune myocarditis;

[0030] Attached Figure 6 Hydroxyurea regulates the transcriptional level of inflammatory factor IL-1β in cardiac tissue of autoimmune myocarditis mouse model;

[0031] Attached Figure 7 Hydroxyurea regulates the transcriptional level of inflammatory factor IL-18 in cardiac tissue of autoimmune myocarditis mouse model;

[0032] Attached Figure 8 Hydroxyurea regulates the transcriptional level of inflammatory factor TNF-α in cardiac tissue of autoimmune myocarditis mouse model;

[0033] Attached Fig. 9 Observation of cardiac tissue pathology of autoimmune myocarditis mouse model treated with hydroxyurea;

[0034] Attached Fig.10 The regulatory effect of hydroxyurea on left ventricular ejection fraction (LVEF) in the color Doppler ultrasound of the heart in the autoimmune myocarditis mouse model;

[0035] Attached Fig.11 The regulatory effect of hydroxyurea on the left ventricular short-axis contraction rate (LVFS) in the autoimmune myocarditis mouse model.

[0036] Attached Fig.12 The regulatory effect of hydroxyurea on the left ventricular internal systolic diameter (LVIDs) in the color Doppler ultrasound index of the heart in the autoimmune myocarditis mouse model. DETAILED DESCRIPTION

[0037] The description of the following embodiments is only used to help understand the method of the present invention and its core idea. Except for the definitions, the technical terms used in the following embodiments have the same meanings as those generally understood by the technicians in the field to which the invention belongs. The test reagents used in the following embodiments are all conventional biochemical reagents unless otherwise specified; the experimental methods are all conventional methods unless otherwise specified. Obviously, the described embodiments are only some embodiments of the present invention, not all embodiments. Based on the embodiments in the present invention, all other embodiments obtained by ordinary technicians in the field without making creative work belong to the scope of protection of the present invention.

[0038] The present invention provides a new class of drugs for preventing and / or treating autoimmune myocarditis

[0039] The levels of inflammatory factors in heart tissue and blood, the infiltration of inflammatory cells in heart tissue and cardiac color indexes are one of the important indicators for evaluating the therapeutic effect of mice with autoimmune myocarditis.

[0040] Example

[0041] The therapeutic effect of hydroxyurea on the mouse model of autoimmune myocarditis

[0042] 1. Experimental animals, instruments, and drugs and reagents

[0043] BALB / c mice (6-8 weeks old) were purchased from Peking University Health Science Center Biotechnology Co., Ltd. The animals were housed in an SPF environment (21±2°C, 12-h light cycle) and had free access to food and water during the experiment. Mouse-derived α cardiac myosin heavy chain polypeptide (purity>95%, Gill Biotech Co., Ltd., Shanghai) and complete Freund's adjuvant (CFA, Sigma-Aldrich, USA) were used. Hydroxyurea and dexamethasone were purchased from Dalian Meilun Biotechnology Co., Ltd. Mouse IL-1β, IL-10, IL-18, and TNF-α activity test kits were purchased from Jiangsu Enzyme Immunity Industry Co., Ltd. Color ultrasound diagnostic instrument (VINNOX6, Feiyinuo, Suzhou), biological signal acquisition and processing system (MadLab-4C / 501H, Beijing Zhongshidichuang, Beijing), anesthesia machine (701001, EZVET, Beijing), isoflurane (709014, EZVET, Shenzhen), fluorescence quantitative qPCR instrument (ABI7500, AppliedBiosystems), PCR instrument (ETC811, Dongsheng), spectrophotometer (K5600, Beijing Kaiao Technology Development Co., Ltd.), horizontal electrophoresis tank (Sub-Cell GT-1704486, BIO-RAD) gel imaging system (Tanon 1600, Shanghai Tianneng Technology Co., Ltd.), ultrapure total RNA extraction kit (CW0581S, Kangwei Century Biotechnology Co., Ltd.), HiScript III 1st Strand cDNA Synthesis Kit (+gDNAwiper) (R312-01, Nanjing Novozyme Biotechnology Co., Ltd.), UltraSYBR Mixture (CW0957, Kangwei Century), DM2000 DNA Marker (CW0623S, Kangwei Century Biotechnology Co., Ltd.), PCR primers (IPAGE purification, Shanghai Bioengineering), fluorescence quantitative PCR 96-well plate (PCR-96-FLT-C, Axygen).

[0044] 2. Experimental Design and Animal Grouping

[0045] Twenty-five male BALB / c mice weighing about 20-30 g and aged 6-8 weeks were randomly divided into 5 groups, 5 mice in each group, namely, the model group (Model group), the positive drug administration group (DEX, dexamethasone group), the hydroxyurea low-dose group (HU-L group), and the hydroxyurea high-dose administration group (HU-H group). In addition, 5 BALB / c mice were used as the normal control group (Control group).

[0046] A total of 5 groups (n=5) were set up in the experiment:

[0047] Animal Grouping:

[0048] Normal control group: normal healthy BALB / c mice, given 0.2 mL of sodium carboxymethylcellulose daily;

[0049] Model group: mice with autoimmune myocarditis were given 0.2 mL of sodium carboxymethyl cellulose by gavage daily;

[0050] (3) Positive drug administration group (0.5 mg / kg): EAM mice were given dexamethasone 0.5 mg / kg by gavage daily;

[0051] (4) Hydroxyurea low-dose group (100 mg / kg): EAM mice were given 100 mg / kg hydroxyurea by gavage daily;

[0052] (5) High-dose hydroxyurea group (200 mg / kg): EAM mice were given 200 mg / kg hydroxyurea by gavage daily;

[0053] The model group and the drug administration group were subcutaneously injected with 0.2 ml of emulsifier (containing 200 μg of polypeptide) on the 0th and 7th day of the experiment, respectively, and the injection sites were the inguinal and axillary regions. The mice in the normal control group were subcutaneously injected with 0.2 mL of 0.9% saline at the same time point and the same injection site. After the last modeling, the mice in the drug administration group were orally administered for 14 consecutive days. Two hours after the last administration, all experimental animals were subjected to cardiac color ultrasound examination. After color ultrasound measurement, 0.5 mL of 20% urethane was anesthetized and killed. Serum was collected for the determination of heart damage and inflammatory factors; at the same time, the upper part of the heart tissue was collected and directly stored in a -80° refrigerator for the determination of the transcription level of inflammatory factors in the heart tissue. The lower part of the heart tissue was fixed in 4% paraformaldehyde fixative, dehydrated and transparent, and then embedded in wax for sectioning and observed by hematoxylin-eosin staining.

[0054] 3. Analytical methods

[0055] The collected whole blood was placed at room temperature for about half an hour, centrifuged at 3500 rpm and 4°C for 10-15 minutes, and the supernatant was taken as serum for subsequent indicator determination.

[0056] The determination of inflammatory factors in serum uses IL-1β, IL-10, IL-18, and TNF-α activity detection kits. For specific methods, please refer to the instructions. The determination of heart damage indicators in serum uses AST activity detection kits. For specific methods, please refer to the instructions. The qPCR determination of cardiac tissue inflammatory factors IL-1β, IL-18, and TNF-α uses a fluorescent quantitative qPCR instrument. For specific methods, please refer to the instructions. The heart color indicators LVEF, LVFS, and LVIDs are determined using a color ultrasound diagnostic instrument. For specific methods, please refer to the instructions.

[0057] Prism 5.0 software was used for statistical analysis. Data were expressed as mean ± standard deviation, and one-way analysis of variance was used for comparison among groups.

[0058] 4. Results

[0059] The therapeutic effects of hydroxyurea or its active metabolites on the autoimmune myocarditis mouse model Figure 1 As shown in the figure, the level of AST, an indicator of heart damage in the model group, was significantly higher than that in the normal control group, indicating that the myocardial inflammation model was successful. Similarly, the level of heart damage indicators in the positive drug group and the high-dose hydroxyurea tablets group after modeling was significantly reduced.

[0060] like Figure 2 As shown in the figure, the level of inflammatory factor IL-1β in the blood of the model group was significantly higher than that of the normal control group, indicating that the myocardial inflammation model was successful. Similarly, the levels of inflammatory factors in the positive drug group, low-dose hydroxyurea group and high-dose hydroxyurea group were significantly reduced after modeling.

[0061] like Figure 3 As shown in the figure, the level of inflammatory factor IL-18 in the blood of the model group was significantly higher than that of the normal control group, indicating that the myocardial inflammation model was successful. Similarly, after modeling, the levels of inflammatory factors in the positive drug group, the low-dose hydroxyurea group, and the high-dose hydroxyurea group were significantly reduced.

[0062] like Figure 4 As shown in the figure, the level of inflammatory factor TNF-α in the blood of the model group was significantly higher than that of the normal control group, indicating that the myocardial inflammation model was successful. Similarly, the levels of inflammatory factors in the positive drug group, the low-dose hydroxyurea group and the high-dose hydroxyurea group were significantly reduced after modeling.

[0063] like Figure 5 As shown, the level of anti-inflammatory factor IL-10 in the blood of the model group was significantly lower than that of the normal control group, and the levels of anti-inflammatory factors in the positive drug group and the high-dose hydroxyurea administration group were significantly increased after modeling.

[0064] like Figure 6As shown, the transcription level of inflammatory factor IL-1β in cardiac tissue of the model group was significantly higher than that of the normal control group, and the level in the positive drug group, low-dose hydroxyurea group and high-dose hydroxyurea tablet group was significantly reduced.

[0065] like Figure 7 As shown, the transcription level of inflammatory factor IL-18 in cardiac tissue of the model group was significantly higher than that of the normal control group, and the level in the positive drug group, low-dose hydroxyurea group and high-dose hydroxyurea tablet group was significantly reduced.

[0066] like Figure 8 As shown, the transcription level of inflammatory factor TNF-α in cardiac tissue of the model group was significantly higher than that of the normal control group, and the levels in the positive drug group, low-dose hydroxyurea group and high-dose hydroxyurea tablet group were significantly reduced.

[0067] like Fig. 9 As shown, by pathological sectioning of the heart tissue in each group, the normal control group mice had fewer inflammatory cells and tight tissue connections, while the inflammatory cells in the model group increased significantly and the tissue connections were loose. The positive drug group and the hydroxyurea administration group significantly improved the above changes.

[0068] like Fig.10 As shown in the figure, the LVEF level of the model group was significantly higher than that of the normal control group, indicating that the myocardial inflammation model was successful. Similarly, the levels of the LVEF level of the positive drug group and the high-dose hydroxyurea tablets group were significantly increased after modeling.

[0069] like Fig.11 As shown in the figure, the level of LVFS in the model group was significantly higher than that in the normal control group, indicating that the myocardial inflammation model was successful. Similarly, the levels of LVFS in the positive drug group and the high-dose hydroxyurea tablets group were significantly increased after modeling.

[0070] like Fig.12 As shown in the figure, the level of LVIDs in the model group was significantly higher than that in the normal control group, indicating that the myocardial inflammation model was successful. Similarly, the levels of LVIDs in the positive drug group and the high-dose hydroxyurea tablets group were significantly reduced after modeling.

[0071] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. Use of hydroxyurea as shown in structural formula (I), or a pharmaceutically acceptable derivative of hydroxyurea as shown in structural formula (I) in the preparation of a drug for preventing and / or treating autoimmune myocarditis, characterized in that: The structural formula (I) is shown below:

2. The use according to claim 1, characterized in that: The pharmaceutically acceptable derivative of hydroxyurea represented by structural formula (I) is one of the pharmaceutically acceptable salts, hydrates, solvates, enantiomers, stereoisomers or tautomers of hydroxyurea represented by structural formula (I).

3. A drug for preventing and / or treating autoimmune myocarditis, characterized in that: The invention comprises at least one of hydroxyurea as shown in the structural formula (I) and a pharmaceutically acceptable derivative of hydroxyurea as shown in the structural formula (I).

4. A drug for preventing and / or treating autoimmune myocarditis according to claim 3, characterized in that: The pharmaceutically acceptable derivative of hydroxyurea represented by structural formula (I) is at least one of the pharmaceutically acceptable salt, hydrate, solvate, enantiomer, stereoisomer or tautomer of hydroxyurea represented by structural formula (I).

5. A drug for preventing and / or treating autoimmune myocarditis according to claim 3 or 4, characterized in that: The drug further comprises pharmaceutically acceptable excipients, which include at least one of a diluent, a binder, an absorbent, a disintegrant, a dispersant, a wetting agent, a solubilizing agent, a buffer, and a surfactant.

6. Use of hydroxyurea as represented by structural formula (I) or a pharmaceutically acceptable derivative of hydroxyurea as represented by structural formula (I) in the preparation of a drug for preventing and / or treating cardiac dysfunction caused by inflammation.

7. The use according to claim 6, characterized in that: The pharmaceutically acceptable derivative of hydroxyurea represented by structural formula (I) is one of the pharmaceutically acceptable salts, hydrates, solvates, enantiomers, stereoisomers or tautomers of hydroxyurea represented by structural formula (I).

8. A drug for preventing and / or treating cardiac dysfunction caused by inflammation, characterized in that: The invention comprises at least one of hydroxyurea as shown in the structural formula (I) and a pharmaceutically acceptable derivative of hydroxyurea as shown in the structural formula (I).

9. A drug for preventing and / or treating cardiac dysfunction caused by inflammation according to claim 8, characterized in that: The pharmaceutically acceptable derivative of hydroxyurea represented by structural formula (I) is at least one of the pharmaceutically acceptable salt, hydrate, solvate, enantiomer, stereoisomer or tautomer of hydroxyurea represented by structural formula (I).

10. A drug for preventing and / or treating autoimmune myocarditis according to claim 8 or 9, characterized in that: The drug further comprises pharmaceutically acceptable excipients, which include at least one of a diluent, a binder, an absorbent, a disintegrant, a dispersant, a wetting agent, a solubilizing agent, a buffer, and a surfactant.

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