Application of N-phenylpiperazine derivative in preparation of medicine for treating neuroinflammation

Abstract

The invention belongs to the field of new application of medicines, and particularly relates to application of N-phenylpiperazine derivatives in preparation of medicines for treating neuroinflammation. It is found for the first time that the N-phenylpiperazine derivative AD-X can improve the neuroinflammation of mice by activating microglia in hippocampus and prefrontal cortex of the mice, and has a certain function of treating the neuroinflammation. The invention provides a novel lead compound for screening medicines for treating neuroinflammation.

Description

Technical Field

[0001] The present invention belongs to the field of new uses of drugs, and more specifically, relates to the application of N-phenylpiperazine derivatives in the preparation of drugs for treating neuroinflammation. Background Art

[0002] Neuroinflammation is a defensive response of the nervous system to infection, injury, or autoimmune reactions. Essentially, it is the abnormal activation of immune cells, such as microglia and lymphocytes, in neural tissues, accompanied by the release of inflammatory factors such as IL-6 and TNF-α, leading to neuronal damage or dysfunction. There is a complex interaction between neuroinflammation and dopamine D3 receptors. In the state of neuroinflammation, microglia are activated and release pro-inflammatory cytokines, which may affect the function of dopamine D3 receptors through multiple pathways. Recent studies have shown that in a mouse model of neuroinflammation induced by lipopolysaccharide injection into the nucleus accumbens, the expression of dopamine D3 receptors is significantly decreased, while microglia are activated and show a pro-inflammatory state. Further studies have shown that the downregulation of dopamine D3 receptors can promote the pro-inflammatory response of microglia through the Akt signaling pathway.

[0003] N-phenylpiperazine is an important drug intermediate in organic drug synthesis, and its derivatives have unique drug activities and applications in the directions of antiviral, antibacterial, and anticancer. In order to further explore the application scope and mechanism of action of N-phenylpiperazine derivatives, further research is still needed. Summary of the Invention

[0004] The purpose of the present invention is to provide the application of N-phenylpiperazine derivatives in the preparation of drugs for treating neuroinflammation to solve the above technical problems.

[0005] To achieve the above purpose, the technical scheme adopted by the present invention is as follows: The present invention provides the application of N-phenylpiperazine derivative AD-X in the preparation of drugs for treating neuroinflammation, and the N-phenylpiperazine derivative is compound AD-X.

[0006] The present invention discovers for the first time that N-phenylpiperazine derivative AD-X can improve the activation of microglia in the hippocampus and prefrontal cortex of mice, thereby improving the neuroinflammation of mice, indicating that it has the function of treating neuroinflammation. It provides a new idea for the treatment of neuroinflammation.

[0007] Furthermore, the N-phenylpiperazine derivative AD-X in the drug is used as the only active ingredient.

[0008] Furthermore, the drug also includes pharmaceutically acceptable excipients.

[0009] Furthermore, the adjuvant includes any one or more of non-toxic fillers, stabilizers, diluents, and adjuvants.

[0010] Furthermore, the dosage form of the drug is a solid dosage form or a solution dosage form. The solid dosage forms include granule agents, tablets, capsules, pills, and dripping pills. The solution dosage forms include oral liquid preparations, gavage agents, and injection dosage forms.

[0011] Furthermore, the solution is a solution composed of oil and the compound AD-X, and the mass-volume ratio of the compound AD-X to the oil is 0.625 mg: 1 ml.

[0012] Based on the same inventive concept, the present invention also provides a drug for treating neuroinflammation, which is prepared by mixing the N-phenylpiperazine derivative AD-X and an adjuvant. The content of the N-phenylpiperazine derivative AD-X in the drug is 0.1 wt% to 99 wt%.

[0013] Furthermore, the dosage form of the drug is a solid dosage form or a solution dosage form. The solid dosage forms include granule agents, tablets, capsules, pills, and dripping pills. The solution dosage forms include oral liquid preparations, gavage agents, and injection dosage forms.

[0014] Furthermore, the solution dosage form is a solution composed of sesame oil and the N-phenylpiperazine derivative AD-X.

[0015] Beneficial effects: The present invention provides the use of the N-phenylpiperazine derivative AD-X in the preparation of a drug for anti-neuroinflammation, and provides a new method for improving neuroinflammation. By intraperitoneally injecting the N-phenylpiperazine derivative AD-X into a mouse model of neuroinflammation, it is found that it can improve the activation of microglia in the hippocampus and prefrontal cortex of the mouse, thereby improving mouse neuroinflammation, indicating that it has the efficacy of inhibiting neuroinflammation. Description of the drawings

[0016] Figure 1 It is a structural diagram of the compound AD-X.

[0017] Figure 2 It is a statistical chart of the iba1 immunoblotting results in the mouse hippocampus. Among them, * indicates p <0.05, ** indicates p <0.01.

[0018] Figure 3 It is a statistical chart of the iba1 immunoblotting results in the mouse prefrontal cortex.

[0019] Figure 4 It is a statistical chart of the gfap immunoblotting results in the mouse hippocampus.

[0020] Figure 5 Statistical chart of gfap immunoblotting results in the prefrontal cortex of mice. Detailed implementation manners

[0021] The present invention will be described in detail below with reference to the accompanying drawings and specific embodiments, but it should not be construed as a limitation of the present invention. Unless otherwise specified, the technical means used in the following embodiments are conventional means well-known to those skilled in the art. The materials, reagents, etc. used in the following embodiments can be obtained from commercial channels unless otherwise specified.

[0022] The compound AD-X used in the present invention is an N-phenylpiperazine derivative, and its structural formula is as Figure 1 shown. The compound AD-X used in the present invention was independently synthesized in this laboratory, and the specific synthesis process can be found in the Chinese patent with the application number: CN201810040879.0.

[0023] Example 1: Effect of compound AD-X on neuroinflammation in mice I. Establishment of a mouse neuroinflammation model.

[0024] The mice were SPF-grade 8-week-old male wild-type C57 / BL6J mice. After being purchased, the animals were first raised in the laboratory environment for one week, with free access to water and food, and a 12-hour light / dark cycle environment. The temperature of the animal room was 22 °C, and the humidity was 58%. Lipopolysaccharide was intraperitoneally injected into male wild-type C57 / BL6J mice to prepare neuroinflammatory mice. Lipopolysaccharide was intraperitoneally injected once at a volume of 10 ml / kg, and the injection dose of lipopolysaccharide was 1 mg / kg.

[0025] II. Compound AD-X improves lipopolysaccharide-induced neuroinflammation in mice.

[0026] Compound AD-X was dissolved in sesame oil at a concentration of 0.625 mg / ml. An experimental group, a model group, a blank group, and a control group were set up, and the specific grouping is as follows:

[0027] Blank group: Wild-type C57 / BL6J mice were intraperitoneally injected with sterile physiological saline and detected 24 hours after injection.

[0028] Model group: Neuroinflammatory model mice were intraperitoneally injected with sterile physiological saline and detected 24 hours after injection.

[0029] Control group: Five days before the injection of lipopolysaccharide, mice were intraperitoneally injected with the solvent control sesame oil, 0.1 ml for every 10 g body weight of the mice. Samples were taken for experiments on the second day of neuroinflammation modeling.

[0030] Experimental group: Five days before injecting lipopolysaccharide, mice with neuroinflammation model were intraperitoneally injected with compound AD-X solution at a concentration of 0.625 mg / ml, and 0.1 ml was injected for each mouse weighing 10 g. Samples were taken for experiments on the second day of neuroinflammation modeling.

[0031] The experimental operation steps of immunoblotting are as follows: Take out the hippocampus and prefrontal cortex tissues of mice, extract proteins. After fully lysing the tissues with lysis buffer, protein quantification was performed by the BCA method. Subsequently, loading buffer was added to denature the proteins at 100 °C for 10 minutes to form the primary structure, and at the same time, 10% SDS by mass fraction was added to neutralize the charge. Then, electrophoresis was performed using a BIO-RAD electrophoresis apparatus. When electrophoresis was carried out, a voltage of 100 v was used for 1 h. After completion, the gel was cut, and a constant current of 220 mA was used for 1.5 h for transfer to transfer the proteins onto the PVDF membrane. After completion, blocking was carried out using BSA or skim milk powder. After blocking and washing, primary antibodies were incubated, including iba1 recombinant rabbit monoclonal antibody (Huaan Biotech, JM36-62), gfap recombinant rabbit monoclonal antibody (Huaan Biotech, SA03-04), GADPH (Wuhan Sanying Biotechnology Co., Ltd., 60004-1-lg) overnight. The next day, after the PVDF membrane was washed, secondary antibody was added, and HRP-labeled goat anti-mouse IgG (servicebio, GB23301) was incubated at room temperature for 2 h. Ultra-sensitive chemiluminescence solution ECL was used for color development, and imaging was performed by exposure and color development. Quantitative analysis was performed using Image J.

[0032] The experimental results are as Figures 2 - 5 shown: Compared with the mice in the blank group, in the hippocampus and prefrontal cortex of the mice in the model group, the expression levels of ionized calcium-binding adapter molecule 1 (iba1) and glial fibrillary acidic protein (gfap) were significantly increased, indicating that lipopolysaccharide successfully induced neuroinflammation. Compared with the mice in the control group, AD-X could significantly reduce the increase in the expression levels of iba1 and gfap induced by lipopolysaccharide, indicating that injecting compound AD-X could improve the activation of microglia in the hippocampus and prefrontal cortex of mice.

[0033] It should be noted that when the claims of the present invention involve numerical ranges, it should be understood that any value between the two endpoints of each numerical range and the two endpoints can be selected. To avoid repetition, preferred embodiments of the present invention are described.

[0034] Although the preferred embodiments of the present invention have been described, those skilled in the art can make additional changes and modifications to these embodiments once they know the basic creative concept. Therefore, the appended claims are intended to be interpreted to include the preferred embodiments and all changes and modifications falling within the scope of the present invention.

[0035] Obviously, those skilled in the art can make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if these modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalent technologies, the present invention is also intended to include these modifications and variations.

Claims

1. Use of N-phenylpiperazine derivatives in the preparation of drugs for treating neuroinflammation, characterized in that, The N-phenylpiperazine derivative is compound AD-X.

2. Use of the N-phenylpiperazine derivative according to claim 1 in the preparation of a medicament for treating neuroinflammation, characterized in that, Compound AD-X in the drug serves as the sole active ingredient.

3. Use of the N-phenylpiperazine derivative according to claim 1 in the preparation of a medicament for treating neuroinflammation, characterized in that, The drug further comprises pharmaceutically acceptable excipients.

4. Use of the N-phenylpiperazine derivative according to claim 3 in the preparation of a medicament for treating neuroinflammation, characterized in that, The excipients include any one or more of non-toxic fillers, stabilizers, diluents, and adjuvants.

5. Use of the N-phenylpiperazine derivative according to claim 1 in the preparation of a medicament for treating neuroinflammation, characterized in that, The dosage form of the drug is a solid dosage form or a solution dosage form. The solid dosage forms include granule agents, tablets, capsules, pills, and dripping pills. The solution dosage forms include oral liquid preparations, gavage agents, and injection dosage forms.

6. Use of the N-phenylpiperazine derivative according to claim 5 in the preparation of a medicament for treating neuroinflammation, characterized in that, The solution is a solution composed of oil and compound AD-X, and the mass-volume ratio of compound AD-X to oil is 0.625 mg: 1 ml.

7. A drug for treating neuroinflammation, characterized in that, The drug is prepared by mixing compound AD-X described in claim 1 and excipients, and the content of compound AD-X in the drug is 0.1 wt% - 99 wt%.

8. The drug according to claim 7, wherein The dosage form of the drug is a solid dosage form or a solution dosage form. The solid dosage forms include granule agents, tablets, capsules, pills, and dripping pills. The solution dosage forms include oral liquid preparations, gavage agents, and injection dosage forms.

9. The drug according to claim 8, characterized in that, The solution dosage form is a solution composed of sesame oil and compound AD-X.

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