WO2022068075A1 - Application of compound zju-37 in preparation of drug for preventing and/or treating liver diseases - Google Patents

Abstract

The present invention relates to the field of medicines. Specifically disclosed are an application of a compound ZJU-37 of formula I in the preparation of a drug for preventing and/or treating liver diseases. After extensive and in-depth research, a use of the compound ZJU-37 in the preparation of a drug for treating liver diseases was found, expanding the application field of the compound ZJU-37. Moreover, the present invention provides treatment methods and treatment drugs for major liver diseases such as severe hepatitis and liver failure, has obvious liver protection and treatment effects, and has significant social benefits.

Description

Application of compound ZJU-37 in the preparation of medicaments for the prevention and/or treatment of liver diseases technical field

The invention relates to the field of medicine, and specifically discloses the application of compound ZJU-37 in the preparation of medicaments for the prevention and/or treatment of liver diseases.

Background technique

Acute and chronic liver injury is a common clinical disease. If it is not controlled and treated in time, it will develop into irreversible diseases such as severe hepatitis and liver failure. The fatality rate of acute liver failure is as high as 80% to 97%. %. At present, there is still no effective clinical treatment and drugs for severe hepatitis and liver failure. Therefore, it is urgent to develop new therapeutic drugs and develop new treatment methods and methods to improve the survival rate of severe hepatitis. Studies have shown that receptor-interacting protein kinase RIP1 is involved in various liver diseases. The use of RIP1 protein kinase inhibitors can reduce cell death and alleviate various inflammatory responses, but the specific mechanism is unclear. Currently in clinical stage RIP1 kinase inhibition is mainly aimed at neurodegenerative diseases and autoimmune diseases, including ulcerative colitis, psoriasis, rheumatoid arthritis and so on. RIP1-involved cell death can promote tissue inflammation and damage, if not inhibit or even amplify tissue damage. Therefore, the development of targeting RIP1 kinase activity has clinical needs and broad application prospects.

SUMMARY OF THE INVENTION

The purpose of the present invention is to overcome the defects of the prior art and provide a new use of the compound ZJU-37.

The first aspect of the present invention provides the use of the compound ZJU-37 of formula I in the preparation of medicaments for the prevention and/or treatment of liver diseases.

Preferably, the liver disease is liver injury, hepatitis, liver failure, liver tissue necrosis/apoptosis.

Wherein, the liver injury may be acute liver injury. The hepatitis may be severe hepatitis. The liver failure may be acute liver failure.

The second aspect of the present invention provides new uses of the compound ZJU-37 of formula I, including the use of any one or more of the following medicaments.

(1) Drugs that can lower ALT levels

(2) Drugs that can reduce AST levels

(3) Drugs that can reduce the degree of liver damage

(4) Drugs that can relieve liver inflammation

(5) Drugs that can reduce the degree of necrosis of liver tissue

(6) Drugs that reduce inflammatory cell infiltration

(7) Drugs that inhibit the secretion of inflammatory factors

(8) Drugs that inhibit apoptosis and programmed necrosis of liver tissue.

Compared with the prior art, the beneficial effects of the present invention are:

After extensive and in-depth research, the present invention discovers for the first time the application of compound ZJU-37 in the preparation of medicaments for the treatment of liver diseases, and opens up the application field of compound ZJU-37. At the same time, the present invention provides a new treatment mode and a treatment drug for serious liver diseases such as severe hepatitis and liver failure, and has obvious liver protection and treatment effects and obvious social benefits.

Description of drawings

Figure 1 shows the comparison of the therapeutic effects of compound ZJU-37 and Nec-1s on liver injury and systemic inflammation in a mouse model of acute liver failure; Figure A and Figure B show the comparison of serum liver injury indexes ALT and AST levels in the LPS/GalN model group The normal control group was significantly increased, and the serum ALT and AST levels in the treatment group were significantly decreased. Compound ZJU-37 could significantly improve ALT and AST compared with commercial Nec-1s. Serum ALT and AST were reduced after Nec-1s treatment (LPS/GalN vs. Nec-1s: 1053±107.7 vs. 724.8±119.5). And ZJU-37 can significantly reduce ALT (LPS/GalN vs. ZJU-37:2110±307.2vs.166±31.11) and AST (LPS/GalN vs. ZJU-37:21053±107.7vs.206.5±21.16)( Figures A, B). We used CBA to detect serum cytokine levels in each group. The results showed that the compound ZJU-37 could reduce the production of TNFα, IL-6, IFN-γ, and MCP-1 better than Nec-1s (Figure C-F). At the same time, ZJU-37 can prolong the survival time of mice compared with Nec-1s (Figure G). The same dose (5 mg/kg) of compound ZJU-37 or commercial Nec-1s was administered intraperitoneally 30 minutes before the LPS/GalN modeling. Compared with commercial Nec-1s, compound ZJU-37 could significantly improve liver injury and liver injury. Systemic inflammation.

Figure 2 shows the comparison of compound ZJU-37 and Nec-1s on liver pathological improvement and neutrophil infiltration in a mouse model of acute liver failure; Figures A and B represent the LPS/GalN model group, and HE staining of liver tissue shows the liver Tissue necrosis and inflammatory cell infiltration were evident. Compared with the model group, the compound ZJU-37 treatment group and the Nec-1s treatment group had significantly reduced liver tissue necrosis and inflammatory cell infiltration (Figures C, D). Histopathological sections showed little necrosis of liver tissue and less inflammatory cell infiltration. The immunohistochemistry of hepatic neutrophils by using Ly6G was consistent with HE staining. Compared with the model group, the neutrophil infiltration in the liver tissue of the compound ZJU-37 treatment group and Nec-1s treatment group was significantly reduced; especially Compound ZJU-37 has obvious curative effect, and histopathological sections of mouse liver tissue showed that there was almost no neutrophil infiltration in liver tissue (Figure E-H). Scale bar 50uM.

Figure 3 shows the comparison of compound ZJU-37 and Nec-1s on liver cell death in a mouse model of acute liver failure; TUNEL-positive cells in the LPS/D-GalN model group were significantly increased compared with the normal control group (Panels A, B), while There was almost no TUNEL positive signal in the normal control group, indicating that a large number of liver cells underwent apoptosis in the model group. The expression of TUNEL was significantly down-regulated in the treatment group; especially the compound ZJU-37 had the best effect, and histopathological sections of mouse liver tissue showed that the TUNEL signal was almost undetectable in liver tissue (Figures C, D).

Detailed ways

Before further describing the specific embodiments of the present invention, it should be understood that the protection scope of the present invention is not limited to the following specific specific embodiments; it should also be understood that the terms used in the examples of the present invention are for describing specific specific embodiments, It is not intended to limit the protection scope of the present invention.

Unless otherwise defined, all technical and scientific terms used in the present invention have the same meaning as commonly understood by one of ordinary skill in the art. In addition to the specific methods, equipment and materials used in the embodiments, according to the mastery of the prior art by those skilled in the art and the description of the present invention, the methods, equipment and materials described in the embodiments of the present invention can also be used Any methods, devices and materials similar or equivalent to those of the prior art can be used to implement the present invention.

The compound ZJU-37 of the present invention belongs to the prior art, and can be prepared by the existing method.

The preparation of embodiment 1 compound ZJU-37 solution

Experimental method: Compound ZJU-37, the stock solution of compound ZJU-37 prepared by using DMSO to dissolve, was diluted with PBS to prepare the solvent of compound ZJU-37 before injection.

Example 2 Study on the effects of compounds ZJU-37 and Nec-1s on LPS/GalN-induced acute liver failure, liver injury and systemic inflammation in mice

experimental method:

Animal models are research tools to verify the effectiveness of new therapeutic drugs and new treatments. Currently, the animal models used internationally to simulate liver failure include drug-induced liver failure models (acetaminophen model, LPS/GalN model, sulfur acetamide model, carbon tetrachloride model, etc.), acute hepatic ischemia model, partial hepatectomy model, etc. Among them, the LPS/GalN model is an ideal animal model of liver failure because of its good reproducibility, no obvious extrahepatic toxicity, and the liver injury performance is close to clinical.

Six-week-old male C57BL/6 mice (purchased from Nanjing Institute of Biomedicine) were randomly divided into normal control group, model group (LPS/GalN group), and compound ZJU-37 treatment group (5 mg/kg ZJU-37+LPS/ GalN group), compound Nec-1s treatment group (5mg/kg Nec-1s+LPS/GalN group). Compound ZJU-37 and Nec-1s were injected intraperitoneally, and 30 minutes later, LPS/GalN (40 μg/kg LPS+800 mg/kg D-GalN) was injected intraperitoneally to make a model; the model group was replaced by an equal volume of solvent in compound ZJU-37 ; The normal control group was not treated. The LPS/GalN model was established for 6 hours, and the blood was collected by removing the eyeball for biochemical and CBA detection.

Experimental results:

The experimental results are shown in Figure 1: Figure A and Figure B show that the levels of serum liver injury indexes ALT and AST in the LPS/GalN model group were significantly higher than those in the normal control group, and the serum ALT and AST levels in the treatment group were significantly lower. Compound ZJU‐ 37 Compared with commercial Nec-1s, it can significantly improve ALT, AST. Serum ALT and AST were reduced after Nec-1s treatment (LPS/GalN vs. Nec-1s: 1053±107.7 vs. 724.8±119.5). And ZJU-37 can significantly reduce ALT (LPS/GalN vs. ZJU-37:2110±307.2vs.166±31.11) and AST (LPS/GalN vs. ZJU-37:21053±107.7vs.206.5±21.16)( Figures A, B). We used CBA to detect serum cytokine levels in each group. The results showed that the compound ZJU-37 could reduce the production of TNFα, IL-6, IFN-γ, and MCP-1 better than Nec-1s (Figure C-F). At the same time, ZJU-37 can prolong the survival time of mice compared with Nec-1s (Figure G). The same dose (5 mg/kg) of compound ZJU-37 or commercial Nec-1s was administered intraperitoneally 30 minutes before LPS/GalN modeling. Compared with commercial Nec-1s, compound ZJU-37 could significantly improve liver injury and liver injury. Systemic inflammation.

That is, compared with Nec-1s, compound ZJU-37 can better reduce liver injury and improve systemic inflammation in LPS/GalN-induced acute liver failure in mice at the same dose.

Example 3 Study on liver pathology and neutrophil infiltration of LPS/GalN-induced acute liver failure in mice by compounds ZJU-37 and Nec-1s

experimental method:

Six-week-old male C57BL/6 mice (purchased from Nanjing Institute of Biomedicine) were randomly divided into normal control group, model group (LPS/GalN group), and compound ZJU-37 treatment group (5 mg/kg ZJU-37+LPS/ GalN group), compound Nec-1s treatment group (5mg/kg Nec-1s+LPS/GalN group). Compound ZJU-37 and Nec-1s were injected intraperitoneally, and 30 minutes later, LPS/GalN (40 μg/kg LPS+800 mg/kg D-GalN) was injected intraperitoneally to make a model; the model group was replaced by an equal volume of solvent in compound ZJU-37 ; The normal control group was not treated. Six hours after LPS/GalN modeling, liver samples were collected for HE and immunohistochemical staining.

Experimental results:

The experimental results are shown in Figure 2: Figures A and B represent the LPS/GalN model group. HE staining of liver tissue showed that liver tissue necrosis and inflammatory cell infiltration were obvious. Compared with the model group, the compound ZJU-37 treatment group and the Nec-1s treatment group had significantly reduced liver tissue necrosis and inflammatory cell infiltration (Figures C, D). Histopathological sections showed little necrosis of liver tissue and less inflammatory cell infiltration. The immunohistochemistry of hepatic neutrophils by using Ly6G was consistent with HE staining. Compared with the model group, the neutrophil infiltration in the liver tissue of the compound ZJU-37 treatment group and Nec-1s treatment group was significantly reduced; especially Compound ZJU-37 has obvious curative effect, and histopathological sections of mouse liver tissue showed that there was almost no neutrophil infiltration in liver tissue (Figure E-H). Scale bar 50uM.

That is, compound ZJU-37 can better improve the liver pathology and reduce the infiltration of neutrophils in LPS/GalN-induced acute liver failure in mice under the same dose.

Example 4 Compound ZJU-37 inhibits apoptosis of liver tissue in LPS/GalN model mice

experimental method:

Six-week-old male C57BL/6 mice (purchased from Nanjing Institute of Biomedicine) were randomly divided into normal control group, model group (LPS/GalN group), and compound ZJU-37 treatment group (5 mg/kg ZJU-37+LPS/ GalN group), compound Nec-1s treatment group (5mg/kg Nec-1s+LPS/GalN group). Compound ZJU-37 and Nec-1s were injected intraperitoneally, and 30 minutes later, LPS/GalN (40 μg/kg LPS+800 mg/kg D-GalN) was injected intraperitoneally to make a model; the model group was replaced by an equal volume of solvent in compound ZJU-37 ; The normal control group was not treated. After 6 hours of LPS/GalN modeling, liver tissue was collected for TUNEL staining.

Experimental results:

The experimental results are shown in Figure 3: TUNEL-positive cells in the LPS/D-GalN model group were significantly increased compared with the normal control group (Figures A, B), while the normal control group had almost no TUNEL-positive signal, indicating that a large number of liver cells were apoptosis in the model group. Death. The expression of TUNEL was significantly down-regulated in the treatment group; especially the compound ZJU-37 had the best effect, and histopathological sections of mouse liver tissue showed that the TUNEL signal was almost undetectable in liver tissue (Figures C, D).

That is, compound ZJU-37 can better improve liver cell death in LPS/GalN-induced acute liver failure in mice. This may also be a potential mechanism for the difference in the efficacy of the two drugs.

The above examples are intended to illustrate the disclosed embodiments of the present invention, and should not be construed as limiting the present invention. Furthermore, various modifications set forth herein and variations in the methods and compositions of the invention will be apparent to those skilled in the art without departing from the scope and spirit of the invention. Although the present invention has been described in detail in conjunction with various specific preferred embodiments of the present invention, it should be understood that the present invention should not be limited to these specific embodiments. Indeed, various modifications as described above which are obvious to those skilled in the art to obtain the invention are intended to be included within the scope of the present invention.

Claims (10)

1. Application of compound ZJU-37 of formula I in the preparation of medicaments for the prevention and/or treatment of liver diseases

   
2. The application according to claim 1, wherein the liver disease is liver injury, hepatitis, liver failure, or liver tissue necrosis/apoptosis.
3. Application of compound ZJU-37 in the preparation of medicines for reducing ALT levels.
4. Application of compound ZJU-37 in the preparation of medicines for reducing AST levels.
5. Application of compound ZJU-37 in the preparation of medicines for reducing the degree of liver injury.
6. Application of compound ZJU-37 in the preparation of medicines for relieving liver inflammation.
7. The application of compound ZJU-37 in the preparation of medicine for reducing the degree of necrosis of liver tissue.
8. Application of compound ZJU-37 in the preparation of medicines for reducing inflammatory cell infiltration.
9. The application of compound ZJU-37 in the preparation of medicines for inhibiting the secretion of inflammatory factors.
10. Application of compound ZJU-37 in the preparation of medicines for inhibiting apoptosis and programmed necrosis of liver tissue.

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