Use of a substituted benzamide in the preparation of a medicament for the prevention or treatment of non-alcoholic fatty liver disease

Abstract

The application relates to application of a substituted benzamide in preparation of a medicine for preventing or treating non-alcoholic fatty liver, characterized in that the benzamide structural formula is shown in formula 1: formula 1, wherein R1, R2 and R3 are the same or different and are selected from one of C1-C4 alkyl groups. The substituted benzamide, especially NGI-1, significantly improves the pathological characteristics of non-alcoholic fatty liver disease, significantly alleviates liver steatosis, reduces neutral fat deposition, improves hepatocyte structure and reduces mild liver fibrosis in a KIF13B MKO mouse model fed with CDAHFD, and indicates that the substituted benzamide, especially NGI-1, has a good effect in the treatment of non-alcoholic fatty liver (NAFLD). The application first proposes that the substituted benzamide, especially NGI-1, has potential application value in the treatment of NAFLD, and provides a new strategy and a candidate drug for early intervention of non-alcoholic fatty liver.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to the application of a substituted benzamide in the preparation of drugs for the prevention or treatment of non-alcoholic fatty liver disease. Background Technology

[0002] Nonalcoholic fatty liver disease (NAFLD) is a chronic liver disease caused by metabolic disorders, and its incidence is rising globally. NAFLD can progress from simple fatty liver (NAFL) to nonalcoholic steatohepatitis (NASH), liver fibrosis, cirrhosis, and may eventually develop into hepatocellular carcinoma. Its pathogenesis involves multiple aspects, including lipid metabolism disorders, oxidative stress, inflammatory responses, insulin resistance, and gut microbiota imbalance.

[0003] Currently, two specific drugs are approved for the treatment of NAFLD: semaglutide and resmetirom; however, in clinical practice, lifestyle interventions (such as diet control and weight loss through exercise) remain the foundation of treatment. While some small molecule drugs in the research and development stage (such as PPAR agonists, FXR agonists, and GLP-1 analogs) have shown some efficacy in certain populations, they still have the following major limitations:

[0004] 1. Unstable efficacy or single mechanism of action: The mechanisms of multi-target metabolic diseases are difficult to completely reverse through intervention of a single pathway, thus limiting the therapeutic effect;

[0005] 2. Significant side effects: Some investigational drugs have caused adverse reactions such as high cholesterol, elevated liver enzymes, or gastrointestinal reactions;

[0006] 3. Narrow patient population: Some drugs are only suitable for NAFLD patients with specific metabolic abnormalities (such as diabetes, high cholesterol);

[0007] 4. Lack of early intervention tools: Existing research focuses on mid-to-late stage NASH and fibrosis, and there is a lack of safe and effective small molecule drugs that can be used in the early stages of the disease.

[0008] Therefore, there is an urgent need to develop a novel drug candidate with a novel mechanism, good safety profile, and the ability to be used for early intervention, in order to fill the current gap in the treatment of non-alcoholic fatty liver disease. Summary of the Invention

[0009] To address the aforementioned problems, the present invention provides the use of a substituted benzamide in the preparation of a drug for the prevention or treatment of non-alcoholic fatty liver disease, the structural formula of which is shown in Formula 1:

[0010] Formula 1,

[0011] R1, R2, and R3 may be the same or different and are selected from C1-C4 alkyl groups.

[0012] Furthermore, R1, R2, and R3 may be the same or different, and are selected from one of methyl, ethyl, n-propyl, and n-butyl.

[0013] Furthermore, R1, R2, and R3 may be the same or different, and are selected from methyl or ethyl.

[0014] Furthermore, R1, R2, and R3 are selected from methyl groups.

[0015] NGI-1 is a small molecule oligosaccharide transferase inhibitor, chemically named 5-[(dimethylamino)sulfonyl]-N-(5-methyl-2-thiazolyl)-2-(1-pyrrolidinyl)-benzamide, with the following structural formula: In NGI-1, R1, R2, and R3 are selected from methyl-substituted benzamides, as shown in Formula 1.

[0016] The above applications can improve steatosis in liver tissue, reduce serum liver damage indicators, reduce triglyceride accumulation, and alleviate early signs of liver fibrosis.

[0017] Furthermore, liver damage markers include ALT and AST.

[0018] Furthermore, the dosage of the substituted benzamide is 5-30 mg / kg.

[0019] Furthermore, the dosage of the substituted benzamide is 10 mg / kg.

[0020] This invention constructs a KIF13B macrophage-specific knockout mouse (KIF13B MKO) and induces a non-alcoholic fatty liver model by feeding the mice with a choline-deficient amino acid high-fat diet (hereinafter referred to as CDAHFD). It was found that substituted benzamide, especially NGI-1, can significantly improve liver tissue steatosis, reduce serum liver injury indicators (ALT, AST), reduce triglyceride accumulation, and alleviate early liver fibrosis.

[0021] The choice of this model is based on clear pathological and mechanistic evidence: KIF13B is an important molecule related to cell membrane transport and signal regulation, and its knockout in macrophages has been found to exacerbate metabolic disorders and inflammatory responses, which are closely related to the pathogenesis of NAFLD. On the other hand, CDAHFD is a classic dietary model that can induce hepatic steatosis, inflammatory responses, and mild fibrosis in mice within a short period of time, simulating the early course of human non-alcoholic steatohepatitis (NASH). Therefore, the combined establishment of KIF13B MKO and CDAHFD forms a highly sensitive NAFLD model with simultaneous metabolic, inflammatory, and tissue damage manifestations, suitable for screening small molecule drug candidates with early intervention potential.

[0022] This invention is the first to propose that substituted benzamides, especially NGI-1, have potential application value in the treatment of NAFLD, providing a new strategy and candidate drug for the early intervention of non-alcoholic fatty liver disease.

[0023] The beneficial technical effects of this invention include:

[0024] 1. Substituted benzamides, especially NGI-1, may exert their effects through interventions in unknown cellular or molecular mechanisms, the effects of which require further investigation.

[0025] 2. Significantly improves pathological features of non-alcoholic fatty liver disease. Substituted benzamides, especially NGI-1, significantly alleviated hepatic steatosis, reduced neutral fat deposition, improved hepatocyte structure, and reduced mild liver fibrosis in a CDAHFD-fed KIF13B MKO mouse model, indicating their good efficacy in the treatment of non-alcoholic fatty liver disease (NAFLD).

[0026] 3. Reduction of liver function damage indicators: The plasma ALT and AST levels of mice treated with substituted benzamide, especially NGI-1, decreased significantly, suggesting that it has a protective effect against hepatocellular damage.

[0027] 4. Improvement of systemic and intrahepatic lipid metabolism disorders: Substituted benzamides, especially NGI-1, significantly reduce triglyceride (TG) levels in liver tissue and plasma, helping to restore hepatic lipid metabolism homeostasis.

[0028] 5. Dual validation by histological and biochemical indicators: HE, Oil Red O, and Sirius Red staining all showed that substituted benzamide, especially NGI-1, had significant tissue protective effects; at the same time, the improvement in serum and liver metabolic indicators further supported its effectiveness. Attached Figure Description

[0029] Figure 1 NGI-1 treatment significantly improved liver morphology in CDAHFD diet-induced KIF13B MKO mice.

[0030] Figure 2 NGI-1 alleviates hepatic steatosis in CDAHFD-fed KIF13B MKO mice.

[0031] Figure 3 NGI-1 reduces neutral fat deposition in the liver tissue of CDAHFD-fed KIF13B MKO mice.

[0032] Figure 4 NGI-1 significantly reduced hepatic triglyceride accumulation in CDAHFD-fed KIF13B MKO mice.

[0033] Figure 5 NGI-1 reduces plasma triglyceride levels in CDAHFD-fed KIF13B MKO mice.

[0034] Figure 6 NGI-1 significantly reduced plasma ALT levels in CDAHFD-fed KIF13B MKO mice.

[0035] Figure 7 NGI-1 significantly reduced plasma AST levels in CDAHFD-fed KIF13B MKO mice. Detailed Implementation

[0036] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0037] To verify the therapeutic effect of NGI-1 on non-alcoholic fatty liver disease (NAFLD), a NAFLD model was established by feeding KIF13B liver-specific knockout mice (KIF13B MKO) with CDAHFD. The experimental mice were fed CDAHFD for 4 weeks starting at 8 weeks of age, and were divided into the following groups:

[0038] Control group (Solvent): Administered an equal volume of solvent via gavage daily;

[0039] Experimental group (NGI-1): NGI-1 was administered by gavage daily (dose was 10 mg / kg / day).

[0040] After the treatment, blood and liver tissue were collected for histological and biochemical analysis.

[0041] Serological markers: ALT and AST levels were significantly decreased, suggesting reduced liver damage;

[0042] Lipid indicators: Liver and plasma triglyceride (TG) levels decreased significantly;

[0043] Histological results: HE staining showed reduced fatty degeneration, Oil Red O staining showed reduced lipid droplet area, and PSR staining showed reduced collagen fiber deposition.

[0044] Example 1: The effect of NGI-1 on improving non-alcoholic fatty liver induced by CDAHFD in KIF13B MKO mice

[0045] Animal model establishment: Eight-week-old male KIF13B macrophage-specific knockout mice (KIF13B MKO) were selected and divided into a control group (Solvent) and an experimental group (NGI-1). All mice were fed a choline-deficient amino acid high-fat diet (CDAHFD) for 4 weeks to induce a non-alcoholic fatty liver disease model.

[0046] Drug treatment: Mice in the experimental group were given NGI-1, 10 mg / kg, intraperitoneally, daily during the modeling period; the control group was given an equal volume of solvent.

[0047] Histological evaluation: HE staining results showed that NGI-1 significantly reduced the degree of macrovesicular steatosis in liver tissue; Oil Red O staining further confirmed that NGI-1 significantly reduced neutral fat deposition.

[0048] Biochemical indicators: Plasma ALT and AST levels in the NGI-1 treatment group were significantly lower than those in the solvent control group, indicating reduced hepatocellular damage. Liver TG and plasma TG levels also decreased significantly, reflecting improved lipid metabolism.

[0049] Statistical analysis: All the above experiments were statistically significant (P < 0.01 or P < 0.001), indicating that NGI-1 has a clear protective effect against NAFLD under this model.

[0050] Figure 1 This image shows a comparison of the gross morphology of the livers of KIF13B MKO mice after 4 weeks of CDAHFD feeding. The left side represents the control group (Solvent), where the liver is significantly enlarged and dull in color, indicating obvious fatty degeneration and pathological changes. The right side represents the NGI-1 treatment group, whose livers are smaller, reddish in color, and smooth in surface, suggesting that NGI-1 can effectively alleviate the phenotypic changes of non-alcoholic fatty liver disease.

[0051] Figure 2The image shows the HE staining results of liver tissue from KIF13B MKO mice after 4 weeks of CDAHFD feeding. The left column shows the original field of view, and the right column shows magnified views of the affected areas. The liver tissue in the solvent group showed extensive macrovesicular steatosis and disordered hepatocyte arrangement; while the NGI-1 treatment group showed a significant reduction in fat vacuoles and improved hepatocyte structure, suggesting that NGI-1 can effectively alleviate hepatic steatosis. The scale bars are shown in the figures (left: 100 μm, right: 50 μm).

[0052] Figure 3 This image shows Oil Red O (ORO) staining results of liver tissue from KIF13B MKO mice after 4 weeks of CDAHFD feeding, used to label neutral fat. The left side is the original field of view, and the right side is a magnified view. Numerous red lipid droplets are visible in the liver tissue of the solvent group, indicating severe lipid accumulation; the NGI-1 treatment group showed a significant reduction in neutral fat, suggesting its role in improving hepatic lipid deposition. Scale bars are shown in the figure (left 100 μm, right 50 μm).

[0053] Figure 4 This shows the difference in liver triglyceride (TG) levels between the NGI-1 treatment group (purple) and the solvent control group (blue) in KIF13B MKO mice after 4 weeks of CDAHFD feeding. NGI-1 significantly reduced liver TG content, suggesting that it can effectively improve hepatic lipid metabolism disorders. The difference between groups was statistically significant (P < 0.001, n = 6). Data are expressed as mean ± SEM.

[0054] Figure 5 The changes in plasma triglyceride (TG) levels in KIF13B MKO mice after 4 weeks of CDAHFD feeding are shown. Compared with the solvent group (blue), the NGI-1 treatment group (purple) showed a significant reduction in plasma TG concentration, suggesting that NGI-1 has an effect on improving systemic lipid metabolism. The differences between groups were highly statistically significant (P < 0.001, n = 6). Data are presented as mean ± SEM.

[0055] Figure 6 In a mouse model fed with CDAHFD for 4 weeks, the NGI-1 treatment group (purple) showed a significant reduction in plasma alanine aminotransferase (ALT) levels compared to the solvent control group (blue), suggesting a less severe degree of hepatocyte damage. Statistical analysis showed a significant difference between the two groups (P < 0.001, n = 6). Data are presented as mean ± SEM.

[0056] Figure 7In a CDAHFD diet-induced KIF13B MKO mouse model, the NGI-1 treatment group (purple) significantly reduced plasma aspartate aminotransferase (AST) levels compared to the solvent control group (blue) (P = 0.004, n = 6). These results suggest that NGI-1 has a significant hepatoprotective effect. Data are presented as mean ± SEM.

Claims

1. The use of a substituted benzamide in the preparation of a drug for the prevention or treatment of non-alcoholic fatty liver disease, characterized in that: The structural formula of the benzamide is shown in Formula 1: Formula 1, R1, R2, and R3 may be the same or different and are selected from one of the C1-C4 alkyl groups.

2. The application according to claim 1, characterized in that: R1, R2, and R3 may be the same or different, and are selected from one of methyl, ethyl, n-propyl, and n-butyl.

3. The application according to claim 1, characterized in that: R1, R2, and R3 may be the same or different, and are selected from methyl or ethyl.

4. The application according to claim 1, characterized in that: R1, R2, and R3 are methyl groups.

5. The application according to any one of claims 1-4, characterized in that: It can improve steatosis in liver tissue, reduce serum liver damage indicators, reduce triglyceride accumulation, and alleviate early signs of liver fibrosis.

6. The application according to claim 5, characterized in that: The indicators of liver damage are ALT and AST.

7. The application according to any one of claims 1-4, characterized in that: The dosage of the substituted benzamide is 5-30 mg / kg.

8. The application according to claim 7, characterized in that: The dosage of the substituted benzamide is 10 mg / kg.

Images