Use of a gcn2 kinase inhibitor in the manufacture of a medicament for treating heart failure with preserved ejection fraction

Abstract

The application belongs to the technical field of biological medicine, and particularly relates to application of a GCN2 kinase inhibitor in preparation of a drug for treating heart failure with preserved ejection fraction. 18 H 12 ClF2N5O3S, the inhibitor can inhibit the activity of GCN2 kinase, and exhibits significant therapeutic effect in a disease model: can obviously improve ventricular diastolic function (E / E' ratio is reduced), reduce the level of heart failure markers, reduce cardiomyocyte hypertrophy and interstitial fibrosis, inhibit the expression of oxidative stress level and inflammatory factors, reduce the accumulation of triglyceride and total cholesterol in myocardium and serum, and improve obesity-related insulin resistance and metabolic disorder. The application provides a novel targeted treatment strategy and candidate drug for heart failure with preserved ejection fraction.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to the application of a GCN2 kinase inhibitor in the preparation of a drug for treating heart failure with preserved ejection fraction. Background Technology

[0002] Heart failure with preserved ejection fraction (HFpEF) has become a significant risk factor for chronic cardiovascular disease worldwide. As a complex clinical syndrome characterized by heart failure symptoms but with a relatively normal left ventricular ejection fraction (LVEF), HFpEF typically progresses from early decline in exercise tolerance to circulatory congestion at rest, and in severe cases can induce multiple organ failure. With the accelerating aging of the global population and the continued rise in the prevalence of metabolic diseases such as obesity, hypertension, and type 2 diabetes, the number of people with HFpEF now accounts for more than half of all heart failure cases. Compared to the general population, patients with HFpEF have a significantly higher risk of cardiovascular death and all-cause hospitalization, and their complex comorbidities lead to generally poorer clinical outcomes, placing a heavy burden on global healthcare systems.

[0003] From a pathogenesis perspective, the pathological process of HFpEF is not limited to the heart itself, but rather involves multi-organ systemic dysfunction driven by a systemic pro-inflammatory state. Comorbid factors such as obesity, hypertension, and metabolic syndrome can induce chronic low-grade inflammation, leading to elevated levels of circulating inflammatory factors (such as TNF-α and IL-6). These inflammatory signals act on coronary artery microvascular endothelial cells, inducing the generation of large amounts of reactive oxygen species (ROS) and reducing the bioavailability of nitric oxide (NO). This, in turn, inhibits the cGMP-PKG signaling pathway, causing increased cardiomyocyte stiffness, cell hypertrophy, and excessive deposition of interstitial collagen fibers (i.e., myocardial fibrosis). In this process, microvascular dysfunction and mitochondrial energy metabolism disorders intertwine, jointly impairing cardiac diastolic compliance, resulting in a sustained increase in left ventricular filling pressure, ultimately driving the pathological evolution from myocardial structural remodeling to clinical heart failure.

[0004] Current clinical treatments for heart failure with reduced ejection fraction (HFpEF) have significant limitations and cannot meet the growing clinical needs. These limitations include: limitations in basic treatments (although managing hypertension and metabolic comorbidities is fundamental, the coexistence of multiple diseases leads to extremely high treatment complexity, and simple lifestyle interventions are insufficient to curb disease progression); limitations in drug therapy (traditional standard treatments for heart failure with reduced ejection fraction (HFrEF) such as beta-blockers and angiotensin-converting enzyme inhibitors have not shown clear survival benefits or significantly reduced mortality in clinical trials for HFpEF patients); and limitations in end-stage treatment (heart transplantation is the final option for refractory HFpEF patients, but its widespread implementation is limited by factors such as extreme donor shortages and high surgical risks).

[0005] In recent years, while SGLT2 inhibitors have made significant progress in reducing the risk of heart failure hospitalization in patients with heart failure pEF, their effects on reversing myocardial remodeling and alleviating myocardial fibrosis remain relatively limited. Furthermore, the HFpEF patient population is highly heterogeneous, making it difficult for existing treatments to achieve individualized coverage, and a significant unmet clinical need remains.

[0006] Therefore, developing novel therapeutic drugs with high safety, clear efficacy, and the ability to provide effective intervention for the highly heterogeneous HFpEF patient population is a technical problem that this invention urgently needs to solve. Summary of the Invention

[0007] The purpose of this invention is to provide an application of a GCN2 kinase inhibitor in the preparation of drugs for the treatment or prevention of heart failure with preserved ejection fraction. The inhibitor provided by this invention can significantly improve ventricular diastolic function (decreased E / E' ratio) by efficiently inhibiting the activity of GCN2 kinase, reduce cardiomyocyte hypertrophy and interstitial fibrosis, while inhibiting oxidative stress and inflammatory factor expression, reducing the accumulation of triglycerides and total cholesterol in the myocardium and serum, and improving obesity-related insulin resistance and metabolic disorders.

[0008] To achieve the above objectives, the present invention provides the following technical solution: This invention provides the use of a GCN2 kinase inhibitor in the preparation of a drug for treating or preventing heart failure with preserved ejection fraction, wherein the GCN2 kinase inhibitor is GCN2iB; the chemical formula of GCN2iB is C 18 H 12 ClF₂N₅O₃S, chemical structural formula as shown in Formula 1 As shown.

[0009] Preferably, the GCN2 kinase inhibitor can improve diastolic dysfunction of the heart.

[0010] Preferably, the GCN2 kinase inhibitor can reduce cardiomyocyte hypertrophy.

[0011] Preferably, the GCN2 kinase inhibitor can reduce cardiac interstitial fibrosis.

[0012] Preferably, the GCN2 kinase inhibitor can alleviate cardiac inflammation and / or cardiac oxidative stress.

[0013] Preferably, the GCN2 kinase inhibitor can reduce triglyceride and total cholesterol levels in the myocardium and serum.

[0014] Preferably, the GCN2 kinase inhibitor can improve insulin resistance.

[0015] Preferably, the dosage form of the drug is an oral dosage form or an injectable dosage form.

[0016] Preferably, the unit dose of GCN2iB in the drug is 3 mg / kg body weight, and the dosing frequency is once every 3 days.

[0017] Compared with the prior art, the beneficial effects of the present invention are as follows: (1) Clear mechanism of action and high specificity: As an ATP competitive inhibitor, GCN2iB works by directly binding to the ATP binding site of GCN2 kinase, thereby specifically and efficiently blocking its kinase activity. This is beneficial for drug optimization and subsequent research on the mechanism of action.

[0018] (2) High safety: The present invention has been verified by animal experiments. At the dose that produces significant therapeutic effect, no obvious toxic side effects were observed, demonstrating good biosafety and tolerability. This key advantage lays a solid foundation for its subsequent clinical development and application.

[0019] (3) The inhibitor provided by this invention, when used in the preparation of drugs for the treatment or prevention of heart failure with preserved ejection fraction, does not act through a single pathway, but rather intervenes in the disease process simultaneously from multiple key pathological links. Specifically, it can effectively reverse myocardial structural and metabolic remodeling, directly reduce the abnormal accumulation of triglycerides and total cholesterol in myocardial tissue and serum, significantly alleviate myocardial cell hypertrophy and interstitial fibrosis, and fundamentally alleviate the core pathological feature of increased ventricular stiffness in HFpEF; comprehensively improve cardiac function and metabolic homeostasis, significantly enhance ventricular diastolic function (echocardiography shows a significant decrease in the E / E' ratio), effectively restore cardiac compliance, and improve obesity-related insulin resistance and systemic metabolic disorders; alleviate oxidative stress and systemic inflammation: inhibit the level of oxidative stress in the cardiovascular system, downregulate the expression of key inflammatory factors, and block the inflammatory cascade response triggered by microvascular endothelial dysfunction, thereby delaying or preventing the malignant progression of the disease to decompensated heart failure and end-stage stages.

[0020] (4) Provides a new approach for drug screening: The causal relationship between GCN2 inhibition and the treatment of heart failure with preserved ejection fraction established by this invention provides a clear target and effective technical path for high-throughput screening and discovery of more candidate drugs for the treatment of this disease. Attached Figure Description

[0021] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0022] Figure 1 This diagram illustrates cardiac function in mice with heart failure with preserved ejection fraction (HFpEF) induced by GCN2iB treatment combined with a high-fat diet and L-NAME. A represents the experimental modeling and drug administration flowchart; B shows representative echocardiographic images of each group of mice, including M-mode, mitral valve flow velocity, and tissue Doppler imaging; CF represents quantitative statistical graphs for detecting left ventricular ejection fraction, fractional shortening, E / E' ratio, and E / A ratio in each group of mice; and G represents a statistical graph for detecting serum NT-proBNP levels.

[0023] Figure 2This diagram illustrates how GCN2iB treatment improves HFpEF-induced cardiac pathological remodeling and oxidative stress in mice. A represents representative stained images of cardiac tissue sections from each group of mice (WGA, Masson Trichrome, HE, and DHE staining, respectively). B and D represent quantitative analyses of the cross-sectional area of ​​cardiomyocytes, the percentage of myocardial fibrosis, and the DHE fluorescence intensity in each group of mice.

[0024] Figure 3 GCN2iB treatment improved HFpEF-induced cardiac and systemic lipid metabolism disorders and insulin resistance in mice. Figures A and B show the analysis of triglyceride (TG) and total cholesterol (TC) levels in cardiac tissue of each group of mice, respectively; figures C and D show the analysis of serum triglyceride (TG) and total cholesterol (TC) levels in each group of mice, respectively; figures E and F show the blood glucose curves (E) and their corresponding area under the curve (AUC) statistics (F) of each group of mice in the oral glucose tolerance test (OGTT); figures G and H show the blood glucose curves (G) and their corresponding area under the curve (AUC) statistics (H) of each group of mice in the insulin tolerance test (ITT). Five mice were included in each group; * represents p<0.05, ** represents p<0.01, and *** represents p<0.001. Detailed Implementation

[0025] To further illustrate the present invention, the technical solutions provided by the present invention will be described in detail below with reference to the accompanying drawings and embodiments, but these should not be construed as limiting the scope of protection of the present invention.

[0026] Unless otherwise specified, the production processes, experimental methods, or testing methods involved in the embodiments of this invention are all conventional methods in the prior art, and their names and / or abbreviations are all conventional names in the field, which are very clear and distinct in the relevant application areas. Those skilled in the art can understand the conventional process steps based on the names and apply the corresponding equipment, and implement them according to conventional conditions or the conditions recommended by the manufacturer.

[0027] The various instruments, equipment, raw materials or reagents used in the embodiments of this invention are not subject to any special restrictions on their source. They are all conventional products that can be purchased through regular commercial channels and can be prepared according to conventional methods known to those skilled in the art.

[0028] Example 1 1.1 Laboratory Animals and Grouping Wild-type C57BL / 6J mice aged 8-10 weeks were randomly divided into 3 groups (n=5 in each group, with no significant difference in body weight), as follows: The control group (WT-CTRL) was fed a normal diet and drank pure water throughout the entire period; Model group (WT-HFpEF): fed a high-fat diet and fed L-NAME (0.5 g / L) in drinking water for 7 weeks; thereafter, injected olive oil solvent (containing 0 mg / kg GCN2iB) intraperitoneally every 3 days for 11 consecutive times, while continuing to feed a high-fat diet and maintain L-NAME in drinking water during the administration period; The treatment group (GCN2iB-HFpEF) was fed a high-fat diet and had L-NAME (0.5 g / L) added to their drinking water for 7 weeks. After that, they were given an intraperitoneal injection of 3 mg / kg GCN2iB in olive oil solution every 3 days for 11 consecutive times. During the treatment period, they were fed a high-fat diet and had L-NAME in their drinking water.

[0029] 1.2 Experimental Methods Mice in each group were fed according to the above-described grouping protocol. Daily consumption of feed and water was recorded, and mice were weighed weekly to monitor their growth. After the experiment (cardiac function echocardiography), whole blood was collected using the ocular blood sampling method and placed in centrifuge tubes. The supernatant serum was separated for serum marker detection. After blood collection, mice were euthanized, and heart tissue was separated. The heart tissue was rinsed with pre-cooled physiological saline to remove surface blood, blotted dry with filter paper, and a portion of the heart tissue was fixed in 4% paraformaldehyde fixative for subsequent histological analysis. Another portion of the heart tissue was rapidly frozen in liquid nitrogen and then transferred to a -80°C freezer for subsequent biochemical marker detection. The experimental procedure is as follows: Figure 1 As shown in Figure A.

[0030] Example 2: Drug GCN2iB improves ventricular diastolic dysfunction induced by a high-fat diet combined with L-NAME in mice. Echocardiography was used to detect cardiac function parameters in mice of each group. Left ventricular ejection fraction (EF), left ventricular fractional shortening (FS), E / E' ratio, and E / A ratio were recorded and analyzed. Simultaneously, mouse serum was collected to detect the level of N-terminal pro-brain natriuretic peptide (NT-proBNP), a biomarker of heart failure. Statistical analysis: Data are expressed as mean ± standard deviation (x ± s). One-way ANOVA was performed using SPSS 22.0 software. LSD-t test was used for pairwise comparisons between groups. *P < 0.05, **P < 0.01, and ***P < 0.001 were considered statistically significant.

[0031] Cardiac function ultrasound images and quantitative test results as follows Figure 1 As shown in BG, according to Figure 1 The experimental data from CD showed that there were no significant differences in left ventricular ejection fraction (EF) and left ventricular fractional shortening (FS) among the groups of mice, maintaining normal levels, consistent with the characteristics of ejection fraction preservation; according to Figure 1 The experimental results of EG showed that, compared with the control group, the E / E' ratio and E / A ratio of mice in the HFpEF model group (injected with olive oil only) were significantly increased, and the serum NT-proBNP level was significantly increased.

[0032] This indicates that a high-fat diet combined with L-NAME exposure successfully induced significant ventricular diastolic dysfunction and heart failure in mice. After treatment with GCN2iB, compared with the HFpEF model group, the E / E' ratio and E / A ratio of the mouse heart were significantly reduced, diastolic function indicators were significantly reversed, and the serum NT-proBNP level also decreased significantly.

[0033] The results of this embodiment confirm that the drug GCN2iB can effectively improve diastolic dysfunction in mice with high-fat diet combined with L-NAME-induced heart failure with preserved ejection fraction (HFpEF) and significantly reduce the levels of heart failure markers.

[0034] Example 3: Drug GCN2iB improves HFpEF-induced cardiac pathological remodeling and oxidative stress in mice. Mouse heart tissue samples were fixed in 4% paraformaldehyde fixative, dehydrated, embedded in paraffin, and sectioned. H&E staining was used to observe tissue morphological damage, Masson staining to assess the degree of interstitial collagen fibrosis, and WGA staining to measure the cross-sectional area of ​​cardiomyocytes. Simultaneously, fresh heart tissue was prepared into frozen sections using OCT embedding and stained with DHE fluorescent probes to assess the accumulation level of reactive oxygen species (ROS) in the myocardial tissue. All stained sections were imaged under a microscope and quantitatively analyzed.

[0035] WGA staining results are as follows Figure 2 As shown in Figures AB, the volume of cardiomyocytes in the HFpEF model group mice was significantly increased; Masson staining results are as follows. Figure 2 A, Figure 2 As shown in Figure C, a large amount of blue-stained collagen fibers were deposited in the myocardial interstitium of the model group; H&E staining results are as follows. Figure 2 As shown in Figure A, the myocardial tissue of the model group showed increased inflammatory infiltration ( Figure 2 A) DHE staining results are as follows Figure 2 A, Figure 2As shown in Figure D, the myocardial tissue in the model group exhibited abundant bright red fluorescence, indicating a large accumulation of reactive oxygen species (ROS). After treatment with GCN2iB, cardiomyocyte hypertrophy was significantly improved. Figure 2 A, Figure 2 B), interstitial fibrosis deposition was significantly reduced ( Figure 2 A, Figure 2 C), reduced inflammatory infiltration of myocardial tissue ( Figure 2 A), and the ROS fluorescence intensity was significantly reduced ( Figure 2 A, Figure 3 D).

[0036] The results of this embodiment confirm that the drug GCN2iB can significantly inhibit HFpEF-induced cardiomyocyte hypertrophy, interstitial fibrosis, inflammation, and myocardial oxidative stress damage.

[0037] Example 4: Drug GCN2iB improves glucose and lipid metabolism disorders and insulin resistance in HFpEF mice. Heart tissue homogenates were extracted from each group of mice and serum was separated. Triglyceride (TG) and total cholesterol (TC) levels were measured strictly according to the instructions of the test kit. At the same time, the mice were fasted before the end of the experiment, and oral glucose tolerance test (OGTT) and insulin tolerance test (ITT) were performed. The changes in blood glucose concentration at different time points after administration were recorded, and the corresponding area under the curve (AUC) was calculated.

[0038] The results of the cardiac and serum tests are as follows: Figure 3 AB and Figure 3 As shown in CD, compared with the normal control group, the levels of TG and TC in the heart tissue and serum of mice in the HFpEF model group were significantly increased, indicating that the high-fat diet combined with L-NAME successfully induced abnormal lipid metabolism in the whole body and the heart. After treatment with GCN2iB, the levels of TG and TC in the heart and serum of mice were significantly decreased. This confirms that GCN2iB can effectively reverse lipid deposition in heart tissue caused by HFpEF. The results of glucose and insulin tolerance tests are as follows... Figure 3 EF and ​ As shown in GH, in the OGTT and ITT experiments, compared with the normal control group, the HFpEF model group mice showed a significant decrease in glucose clearance capacity and a significant increase in the area under the curve (AUC), exhibiting severe glucose intolerance and insulin resistance. After GCN2iB treatment, the dynamic curve of glucose metabolism in mice was significantly flattened, and the AUC of both OGTT and ITT was significantly reduced.

[0039] In summary, this invention, through echocardiographic assessment of cardiac function, biochemical analysis of myocardial tissue, and histological examination, confirms that the drug GCN2iB can exert a protective effect against HFpEF myocardial injury by improving ventricular diastolic dysfunction induced by high-fat diet combined with L-NAME, reducing myocardial lipid deposition, inhibiting cardiomyocyte hypertrophy, interstitial fibrosis, inflammation, and myocardial oxidative stress damage, thus providing a new drug option for the treatment of heart failure with preserved ejection fraction.

[0040] Although the above embodiments have provided a detailed description of the present invention, they are only some embodiments of the present invention, and not all embodiments. People can obtain other embodiments based on these embodiments without creative effort, and these embodiments all fall within the protection scope of the present invention.

Claims

1. The use of GCN2 kinase inhibitors in the preparation of drugs for the treatment or prevention of heart failure with preserved ejection fraction, characterized in that, The GCN2 kinase inhibitor is GCN2iB; the chemical formula of GCN2iB is C. 18 H 12 ClF₂N₅O₃S, chemical structural formula as shown in Formula 1 As shown.

2. The application according to claim 1, characterized in that, The GCN2 kinase inhibitor can improve diastolic dysfunction of the heart.

3. The application according to claim 1, characterized in that, The GCN2 kinase inhibitor can reduce cardiomyocyte hypertrophy.

4. The application according to claim 1, characterized in that, The GCN2 kinase inhibitor can reduce cardiac interstitial fibrosis.

5. The application according to claim 1, characterized in that, The GCN2 kinase inhibitor can alleviate cardiac inflammation and / or cardiac oxidative stress.

6. The application according to claim 1, characterized in that, The GCN2 kinase inhibitor can reduce triglyceride and total cholesterol levels in the myocardium and serum.

7. The application according to claim 1, characterized in that, The GCN2 kinase inhibitor can improve insulin resistance.

8. The application according to claim 1, characterized in that, The drug is available in oral or injectable form.

9. The application according to any one of claims 1-8, characterized in that, The unit dose of GCN2iB in the drug is 3 mg / kg body weight, and the dosing frequency is once every 3 days.

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