Use of a compound RKC-B1 in inhibiting neuroinflammation

Abstract

The present invention discloses the use of a compound of formula (I), RKC-B1, in inhibiting neuroinflammation, belonging to the field of pharmaceutical technology. Pharmacological experiments have shown that RKC-B1 can significantly inhibit the release of LPS-induced inflammatory factors such as interleukin-1β (IL-1β), interleukin-6 (IL-6), monocyte chemoattractant MCP-1, and intercellular adhesion molecule-1 (ICAM-1) in the mouse cerebral cortex, hippocampus, and striatum, and has significant inhibitory activity against neuroinflammation. Therefore, the compound can be used to prepare drugs for preventing and / or treating neuroinflammation and related central nervous system diseases such as traumatic brain injury, stroke, Alzheimer's disease, Parkinson's disease, multiple sclerosis, and Huntington's disease, and has high clinical application value and development prospects. #imgabs0#

Description

Technical Field

[0001] The present invention belongs to the field of medical technology and relates to the use of RKC-B1 in the treatment of central nervous system diseases such as neuroinflammation and related traumatic brain injury, stroke, Alzheimer's disease, Parkinson's disease, multiple sclerosis and Huntington's disease. Background Art

[0002] The neuroinflammatory response is a complex, progressive cascade characterized by the activation and proliferation of glial cells, the infiltration of peripheral inflammatory cells, and the expression of related inflammatory cytokines. Neuroinflammatory responses play a key role in the pathogenesis of central nervous system diseases, including traumatic brain injury, stroke, Alzheimer's disease, Parkinson's disease, and Huntington's disease. Microglia are immune cells of the central nervous system. Brain injury triggers microglial activation and the release of inflammatory factors and adhesion molecules, such as interleukin-1β (IL-1β), interleukin-6 (IL-6), monocytic chemoattractant protein 1 (MCP-1), and intercellular adhesion molecule-1 (ICAM-1). These factors interact and regulate each other, leading to neuronal damage, degeneration, and even death. Controlling neuroinflammation is a breakthrough in delaying or treating these neurological diseases.

[0003] LPS, a major component of the cell wall of Gram-negative bacteria, is a potent inducer of inflammatory responses. Animal studies have shown that intraperitoneal administration of LPS to mice can induce astrocyte and microglial activation and promote the expression of inflammatory cytokines in the brain. LPS-induced inflammatory responses have become a classic reagent for establishing animal models of neuroinflammation and are widely used in the screening, evaluation, and mechanism studies of drugs with anti-inflammatory activity.

[0004] Compounds derived from marine microorganisms not only have a biodiverse origin, but also have complex and varied metabolite structures, embodying an endless amount of structural diversity. Obtaining compounds that can inhibit neuroinflammation from marine natural products is an important approach for drug development in this field. RKC-B1 is derived from the marine micromonospora FIM02-523, and B1 is produced by single-component fermentation through high-yield strain selection and fermentation optimization. Currently, there are few reports on the pharmacological activity of RKC-B1, and there are no reports on RKC-B1 being used to inhibit neuroinflammation and related neurological diseases. Summary of the Invention

[0005] The present invention evaluated the anti-neuroinflammatory activity of RKC-B1 using a neuroinflammatory mouse model induced by intraperitoneal injection of LPS. The structural formula of the compound RKC-B1 of the present invention is shown in Formula I:

[0006]

[0007] One of the technical problems to be solved by the present invention is to provide the use of RKC-B1 represented by formula (I) in drugs and other products related to inhibiting neuroinflammation.

[0008] The second technical problem to be solved by the present invention is to provide the use of RKC-B1 shown in (I) in preventing and / or treating central nervous system diseases such as traumatic brain injury, cerebral stroke, Alzheimer's disease, Parkinson's disease, Huntington's disease, etc. caused by neuroinflammation.

[0009] The compound RKC-B1 of the present invention can significantly reduce the increased levels of inflammatory response-related factors IL-1β, IL-6, MCP-1, and ICAM-1 in the cerebral cortex, hippocampus, and striatum of mice induced by LPS, indicating that RKC-B1 has significant anti-neuroinflammatory activity.

[0010] In addition, drugs and other products prepared with RKC-B1 as an active ingredient for preventing and / or treating central nervous system diseases such as neuroinflammation and related traumatic brain injury, stroke, Alzheimer's disease, Parkinson's disease, multiple sclerosis and Huntington's disease also fall within the scope of protection of the present invention.

[0011] The present invention also relates to pharmaceutical compositions of the compounds of the present invention and conventional pharmaceutical excipients or adjuvants.

[0012] The pharmaceutical composition of the compounds of this invention can be prepared according to methods well known in the art. When used for this purpose, if necessary, the compounds of this invention can be combined with one or more solid or liquid pharmaceutical excipients and / or adjuvants to prepare suitable administration forms or dosage forms that can be used as human or veterinary medicines.

[0013] The compound of the present invention or the pharmaceutical composition containing the same can be administered in a unit dosage form, and the administration route can be enteral or parenteral, such as oral, intramuscular, subcutaneous, nasal, oral mucosa, skin, peritoneum or rectum.

[0014] The compound of the present invention or the pharmaceutical composition containing the same can be administered by injection, including intravenous injection, intramuscular injection, subcutaneous injection, intradermal injection and acupuncture injection.

[0015] The dosage form can be a liquid or solid dosage form. For example, liquid dosage forms can be true solutions, colloids, microparticles, emulsions, or suspensions. Other dosage forms include tablets, capsules, pellets, aerosols, pills, powders, solutions, suspensions, emulsions, granules, suppositories, and freeze-dried powder injections.

[0016] The compound of the present invention can be prepared into common preparations, sustained-release preparations, controlled-release preparations, targeted preparations and various microparticle delivery systems.

[0017] Beneficial technical effects: This compound has outstanding pharmacological activity and is suitable for use in treating various diseases of the nervous system. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 Effect of RKC-B1 on IL-1β levels in the cerebral cortex of mice induced by LPS. Data are expressed as mean ± SD. ##P < 0.01 compared with the control group; **P < 0.01 compared with the LPS model group.

[0019] Figure 2 Effect of RKC-B1 on IL-6 levels in the cerebral cortex of mice induced by LPS. Data are expressed as mean ± SD. Compared with the control group, ##P < 0.01; compared with the LPS model group, *P < 0.05, **P < 0.01.

[0020] Figure 3 Effect of RKC-B1 on MCP-1 levels in the cerebral cortex of mice induced by LPS. Data are expressed as mean ± SD. ##P < 0.01 compared with the control group; **P < 0.01 compared with the LPS model group.

[0021] Figure 4 Effect of RKC-B1 on ICAM-1 levels in the cerebral cortex of mice induced by LPS. Data are expressed as mean ± SD. ##P < 0.01 compared with the control group; **P < 0.01 compared with the LPS model group.

[0022] Figure 5 Effect of RKC-B1 on IL-1β levels in the hippocampus of mice induced by LPS. Data are expressed as mean ± SD. ##P < 0.01 compared with the control group; **P < 0.01 compared with the LPS model group.

[0023] Figure 6 Effect of RKC-B1 on IL-6 levels in the hippocampus of mice induced by LPS. Data are expressed as mean ± SD. Compared with the control group, ##P < 0.01; compared with the LPS model group, *P < 0.05, **P < 0.01.

[0024] Figure 7 Effect of RKC-B1 on MCP-1 levels in the hippocampus of mice induced by LPS. Data are expressed as mean ± SD. ##P < 0.01 compared with the control group; **P < 0.01 compared with the LPS model group.

[0025] Figure 8Effect of RKC-B1 on IL-1β levels in the striatum of LPS-induced mice. Data are expressed as mean ± SD. ##P < 0.01 compared with the control group; **P < 0.01 compared with the LPS model group.

[0026] Figure 9 Effect of RKC-B1 on IL-6 levels in the striatum of LPS-induced mice. Data are expressed as mean ± SD. ##P < 0.01 compared with the control group; **P < 0.01 compared with the LPS model group.

[0027] Figure 10 Effect of RKC-B1 on MCP-1 levels in the striatum of LPS-induced mice. Data are expressed as mean ± SD. ##P < 0.01 compared with the control group; **P < 0.01 compared with the LPS model group.

[0028] Figure 11 Effect of RKC-B1 on ICAM-1 levels in the striatum of mice induced by LPS. Data are expressed as mean ± SD. ##P < 0.01 compared with the control group; **P < 0.01 compared with the LPS model group. DETAILED DESCRIPTION

[0029] Effects of RKC-B1 on LPS-induced neuroinflammation in the cerebral cortex, hippocampus and striatum of mice

[0030] 1. Experimental Methods

[0031] 1.1 Animal grouping and drug administration

[0032] Reagents: LPS (Sigma); IL-1β, IL-6, MCP-1, ICAM-1 detection ELISA kits (Jitai Yikesai Biotechnology Co., Ltd.).

[0033] Adult male BALB / c mice weighing 20–22 g were obtained from Beijing Sibeifu Biotechnology Co., Ltd. (animal quality certificate number: SYXK(Beijing)2016-0002). They were housed under normal conditions: a temperature of 25 ± 1°C, a relative humidity of 55%–65%, and a 12-h photoperiod. Food and water were available ad libitum.

[0034] Animal Grouping and Dosing: After 3 days of acclimatization, animals were randomly divided into the following groups: vehicle control group (Vehicle), LPS model group (LPS), RKC-B1 5 mg / kg group (LPS + RKC-5 mg / kg), and RKC-B1 10 mg / kg group (LPS + RKC-10 mg / kg). The vehicle control and LPS model groups were intraperitoneally injected with an equal volume of vehicle (0.1% Tween 80-containing saline) for 7 consecutive days; the RKC-B1 5 and 10 mg / kg groups were intraperitoneally injected for 7 consecutive days. After the last dose, the LPS model group and the RKC-B1 5 and 10 mg / kg groups were intraperitoneally injected with 5 mg / kg of LPS; the vehicle control group was intraperitoneally injected with an equal volume of saline. Six hours after LPS injection, tissue samples were collected from 8 animals in each group. Cortical, hippocampal, and striatal tissues were isolated and stored at -80°C for subsequent ELISA analysis.

[0035] 1.2 ELISA test

[0036] Tissues from specific mouse sites were weighed and 1 mL of pre-chilled saline was added per 100 mg of tissue. The homogenate was ground on ice using a handheld homogenizer and centrifuged at 4500 rpm at 4°C for 15 minutes. The supernatant was collected. Protein content was determined by the BCA assay, and IL-1β, IL-6, MCP-1, and ICAM-1 levels were measured by ELISA kits. The procedure was performed according to the manufacturer's instructions. The following steps were briefly described: The sample or a standard solution with varying concentrations was added to a 96-well ELISA plate. The biotinylated antibody working solution was then added and incubated at room temperature for 120 minutes. After thorough washing, the sample and biotinylated antibody working solution were discarded. HRP-conjugated secondary antibody was added to all wells except the blank well, and the plate was incubated at room temperature for 60 minutes. After discarding the secondary antibody solution and washing thoroughly, a chromogenic substrate was added and incubated at room temperature for 15 minutes in the dark. Within 10 minutes after adding the stop solution, the absorbance at 450 nm was measured. The levels of inflammatory factors were calculated by plotting a standard curve.

[0037] Experimental results:

[0038] Compared with the normal control group, the levels of inflammatory response-related factors IL-1β, IL-6, MCP-1, and ICAM-1 in the cortical tissue of mice in the LPS model group were significantly increased; while RKC-B1 5 and 10 mg·kg -1 The drug-treated groups could significantly reduce the expression of IL-1β, a inflammatory response-related factor in the cortical tissue of mice induced by LPS (Table 1, Figure 1 ), IL-6 (Table 2, Figure 2 ), MCP-1 (Table 3, Figure 3 ), ICAM-1 (Table 4, Figure 4 ) increase.

[0039] Table 1 Effects of RKC-B1 on IL-1β levels in LPS-induced mouse cerebral cortex

[0040] Group IL-1β (pg / mg protein) Normal control group 31.68±6.30 LPS model group 182.50±67.82## LPS+RKC-B1 5mg / kg group 68.99±29.81** LPS+RKC-B1 10 mg / kg group 58.02±29.85**

[0041] Table 2 Effects of RKC-B1 on IL-6 levels in LPS-induced mouse cerebral cortex

[0042] Group IL-6 (pg / mg protein) Normal control group 2.30±0.53 LPS model group 22.46±11.83## LPS+RKC-B1 5mg / kg group 4.81±1.91** LPS+RKC-B1 10 mg / kg group 12.26±8.62*

[0043] Table 3 Effects of RKC-B1 on MCP-1 levels in LPS-induced mouse cerebral cortex

[0044] Group MCP-1 (pg / mg protein) Normal control group 3.66±1.35 LPS model group 1607.18±741.02## LPS+RKC-B1 5mg / kg group 393.32±216.90** LPS+RKC-B1 10 mg / kg group 404.56±236.50**

[0045] Table 4 Effects of RKC-B1 on ICAM-1 levels in LPS-induced mouse cerebral cortex

[0046] Group ICAM-1 (pg / mg protein) Normal control group 59.76±8.23 LPS model group 267.89±77.82## LPS+RKC-B1 5mg / kg group 161.76±67.94** LPS+RKC-B1 10 mg / kg group 159.21±56.24**

[0047] Compared with the normal control group, the levels of inflammatory response-related factors IL-1β, IL-6, MCP-1, and ICAM-1 in the hippocampus of mice in the LPS model group were significantly increased; while RKC-B1 5 and 10 mg·kg -1 The drug-treated groups could significantly reduce the expression of IL-1β, a inflammatory response-related factor in the hippocampus of mice induced by LPS (Table 5, Figure 5 ), IL-6 (Table 6, Figure 6 ), MCP-1 (Table 7, Figure 7 ) increase.

[0048] Table 5 Effects of RKC-B1 on IL-1β levels in the hippocampus of mice induced by LPS

[0049] Group IL-1β (pg / mg protein) Normal control group 24.77±6.56 LPS model group 126.35±31.08## LPS+RKC-B1 5mg / kg group 77.30±31.89** LPS+RKC-B1 10 mg / kg group 57.73±30.21**

[0050] Table 6 Effects of RKC-B1 on IL-6 levels in the hippocampus of mice induced by LPS

[0051]

[0052]

[0053] Table 7 Effects of RKC-B1 on MCP-1 levels in the hippocampus of mice induced by LPS

[0054] Group MCP-1 (pg / mg protein) Normal control group 22.42±5.39 LPS model group 697.08±386.51## LPS+RKC-B1 5mg / kg group 265.51±179.16** LPS+RKC-B1 10 mg / kg group 209.84±128.55**

[0055] Compared with the normal control group, the levels of proinflammatory cytokines IL-1β, IL-6, MCP-1, and ICAM-1 in the striatum of mice in the LPS model group were significantly increased; while RKC-B1 5 and 10 mg·kg -1 The drug-treated groups could significantly reduce the expression of IL-1β, an inflammatory factor, in the striatum tissue of mice induced by LPS (Table 8, Figure 8 ), IL-6 (Table 9, Figure 9 ), MCP-1 (Table 10, Figure 10 ), ICAM-1 (Table 11, Figure 11 ) increase.

[0056] Table 8 Effects of RKC-B1 on IL-1β levels in LPS-induced mouse striatum

[0057] Group IL-1β (pg / mg protein) Normal control group 36.30±7.37 LPS model group 109.16±27.42## LPS+RKC-B1 5mg / kg group 58.96±12.06** LPS+RKC-B1 10 mg / kg group 64.60±16.72**

[0058] Table 9 Effects of RKC-B1 on IL-6 levels in LPS-induced mouse striatum

[0059] Group IL-6 (pg / mg protein) Normal control group 4.40±0.69 LPS model group 23.35±5.00## LPS+RKC-B1 5mg / kg group 7.20±0.75** LPS+RKC-B1 10 mg / kg group 9.05±2.35**

[0060] Table 10 Effects of RKC-B1 on MCP-1 levels in the striatum of mice induced by LPS

[0061] Group MCP-1 (pg / mg protein) Normal control group 2.38±0.82 LPS model group 547.14±244.91## LPS+RKC-B1 5mg / kg group 196.41±86.39** LPS+RKC-B1 10 mg / kg group 175.52±142.81**

[0062] Table 11 Effects of RKC-B1 on ICAM-1 levels in LPS-induced mouse striatum

[0063] Group ICAM-1 (pg / mg protein) Normal control group 139.55±11.21 LPS model group 326.92±40.64## LPS+RKC-B1 5mg / kg group 233.62±55.06** LPS+RKC-B1 10 mg / kg group 221.37±44.43** .

Claims

1. Use of the compound RKC-B1 represented by general formula (I) or a pharmaceutically acceptable salt thereof in the preparation of a drug for inhibiting neuroinflammation, 2. Use of a pharmaceutical composition in the preparation of a drug for inhibiting neuroinflammation, characterized in that: The pharmaceutical composition comprises the compound RKC-B1 represented by general formula (I) or a pharmaceutically acceptable salt thereof, and at least one pharmaceutically acceptable carrier.

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