Use of conessine in the preparation of a medicine for resisting influenza a virus infection

Abstract

The application provides application of Conessine in preparation of a medicine for resisting influenza A virus infection, and belongs to the technical field of biological medicine. 50＝ 0.45 muM, the value is low, and the inhibition efficiency is high. 50 The application also detects the toxicity of Conessine to cells, and finds that the toxicity of Conessine to cells is also low, and the CC 50 The safety index (SI) is much higher than the safety range, and the safety and effectiveness of Conessine as an IAV influenza A virus inhibitor are more guaranteed. Conessine can be used for treating respiratory system infection, pneumonia, encephalitis and other diseases caused by influenza A virus, and provides a new effective treatment medicine for global influenza epidemic.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, and in particular to the application of Conessine in the preparation of drugs for treating influenza A virus infection. Background Technology

[0002] Influenza A virus (IAV) is a human respiratory pathogen that is widespread globally. It is highly contagious and pathogenic, causing severe respiratory infections, pneumonia, encephalitis, and in some cases, death. IAV belongs to the Orthomyxoviridae family and is an enveloped, negative-sense RNA virus. Based on the antigenic properties of its surface hemagglutinin protein (HA) and neuraminidase protein (NA), it can be divided into several subtypes, such as H1N1 and H3N2.

[0003] Influenza A virus (IAV) is a multi-host zoonotic pathogen that can spread between species, leading to reclassification and the emergence of different viral types, which in turn can cause new influenza epidemics and widespread outbreaks. The Spanish flu of 1918-1919 was the most severe, causing over 40 million deaths. Currently, there are few effective treatments for IAV, making drugs that inhibit IAV a key focus for researchers.

[0004] The method of using natural chemical components extracted from typical Chinese medicines to inhibit IAV has strong medical value and clinical potential, and it is in line with the general research direction of integrating Chinese and Western medicine. Compared with the artificially synthesized small molecules of pharmaceutical chemicals used to inhibit IAV in the past, the natural chemical molecules extracted from plant components can more effectively avoid the possible side effects of chemically synthesized molecules on humans, and are more guaranteed in terms of safety and social promotion.

[0005] Conessine, a small-molecule compound from the natural Chinese herbal medicine *Holarrhena flobunda*, is a steroidal alkaloid isolated from the plant and is one of its important natural active ingredients. *Holarrhena flobunda* belongs to the genus *Holarrhena* in the family Apocynaceae. In traditional Chinese medicine, it is mainly used to treat dysentery and flatulence, and has antidiarrheal and antipyretic effects. Its functions include clearing heat and relieving cough, detoxifying and killing parasites, astringing the intestines and stopping diarrhea, cooling the blood and stopping dysentery, and promoting diuresis and relieving pain. It is also effective in treating wind-heat colds, liver and gallbladder diseases, gastrointestinal diseases, and diarrhea.

[0006] Previous studies have shown that Conessine, a small molecule compound derived from natural Chinese medicine, is a highly effective and selective H3 receptor antagonist. Current research has revealed that it primarily possesses antimalarial, anti-dysentery, and antibacterial activities, including inhibition of the Pseudomonas aeruginosa efflux pump. At the cellular level, Conessine has been found to regulate autophagy, inhibiting muscle cell death caused by excessive activation of the hydrogen peroxide-induced autophagy pathway. As a mature small molecule drug derived from natural plants, it possesses the safety profile to be a suitable inhibitor of influenza A virus.

[0007] However, there are few reports on the antiviral effects of the small molecule compound Conessine. Therefore, further research on the antiviral effects of Conessine is of great significance for solving the global influenza problem. Summary of the Invention

[0008] The purpose of this invention is to provide the application of Conessine in the preparation of drugs for treating influenza A virus infection. Conessine is used to treat influenza A virus infection, and the treatment has a good inhibitory effect and high safety.

[0009] To achieve the above-mentioned objectives, the present invention provides the following technical solution:

[0010] This invention provides the application of Conessine in the preparation of drugs for treating influenza A virus infection.

[0011] Furthermore, the structure of Conessine is shown in formula (I):

[0012]

[0013] Furthermore, Conessine can inhibit the infection and replication of influenza A virus.

[0014] Furthermore, the anti-influenza A virus infection drug may be added with a pharmaceutically acceptable carrier.

[0015] Furthermore, the dosage form of the anti-influenza A virus infection drug is one of the following: powder, solution, capsule, granule, tablet, emulsion, or suspension.

[0016] The beneficial effects of the application of Conessine described in this invention in the preparation of drugs against influenza A virus infection compared with existing technologies are as follows:

[0017] (1) This invention utilizes the small molecule Conessine to inhibit influenza A virus and detects the IC50 value of Conessine's inhibitory effect. The results show that Conessine has a strong inhibitory effect on influenza A virus, with an IC50 of 2.07 μM, which is relatively low and indicates high inhibitory efficiency. The cytotoxicity of the small molecule Conessine was also detected, and it was found to have low cytotoxicity, with an IC50 of 34.11 μM, which is relatively high. The selectivity index (SI) is much higher than the safety range, which further ensures the safety and efficacy of Conessine as an inhibitor of influenza A virus.

[0018] (2) This invention utilizes a natural chemical component extracted from plants—small molecule Conessine—to inhibit influenza A virus, effectively avoiding the side effects of chemically synthesized molecules and exhibiting high safety. Furthermore, small molecule Conessine has a high inhibitory efficiency against influenza A virus. It can be used to treat respiratory infections, pneumonia, encephalitis, and other diseases caused by influenza A virus, providing a new and effective treatment for influenza worldwide. Attached Figure Description

[0019] 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.

[0020] Figure 1 The figures show the cytotoxicity curve of Conessine and the inhibition rate curve of IAV in Experiment Example 1. Detailed Implementation

[0021] The embodiments of the present invention are described in detail below. These embodiments are intended to explain the present invention and should not be construed as limiting the present invention. Where specific techniques or conditions are not specified in the embodiments, they are performed according to the techniques or conditions described in the literature in the art or according to the product instructions. Reagents or instruments used, unless otherwise specified, are all commercially available conventional products.

[0022] The small molecule Conessine (also known as Conessinum, Wrightine, Roquessine, Neriine, Konessin) used in the following embodiments of the present invention is manufactured by TargetMo, product number T20712 CAS 546-06-5.

[0023] The small molecule Conessine of this invention belongs to the steroidal alkaloid family and has the molecular formula: C 24 H40 N2, molecular weight: 356.59, structure shown in formula (I):

[0024]

[0025] Example 1

[0026] Example 1 of this invention involved screening drugs to inhibit influenza A virus, specifically including the following steps:

[0027] Preliminary screening of small molecules with IAV inhibitory activity was conducted in a small molecule drug library.

[0028] The drug molecules in the small molecule library are all at a concentration of 10 mM (in DMSO). The screened IAVs are linked to a gene expressing Gaussia Luciferase (Gluc), and the viral infection and replication status in the host cell can be obtained by detecting the luciferase value after cell infection.

[0029] Conessine, a small molecule from the small molecule library, was diluted 1000-fold to 10 μM in Mock DMEM, and then the IAVA / PR / 8 / 34(PR8)-Gluc strain was added to it, with the original venom titer being 1.0 × 10⁻⁶. 5 The final venom concentration was then diluted 100-fold and mixed with MDCK cells for infection. After 24 hours, the cells were subjected to luciferin detection (Gaussian-LumiNova). TM The Gaussian luciferase reporter gene assay kit (Beyotime Biotechnology) was used to detect luciferin levels in both positive and negative control groups, including those infected with IAV virus only and those without virus-free pure cells. The results showed that Conessine's luciferin value was close to that of the pure cell control group, while the luciferin values ​​of most other small molecules were close to those of the IAV-infected cell control group. Therefore, Conessine exhibits a near 100% inhibition rate against IAV and can be used to prepare drugs for treating influenza A virus infection.

[0030] Example 2

[0031] Example 2 of this invention utilizes Conessine to prepare tablets for treating influenza A virus infection. The specific preparation method is as follows:

[0032] Take Conessine, add it to the conventional excipients for tablet preparation, mix well, and then compress into tablets.

[0033] Example 3

[0034] Example 3 of this invention utilizes Conessine to prepare a powdered anti-influenza A virus infection drug. The specific preparation method is as follows:

[0035] Take Conessine, add it to the conventional excipients for preparing the granules, mix well, and then package.

[0036] Example 4

[0037] Example 4 of this invention utilizes Conessine to prepare an aerosol-based antiviral drug against influenza A virus infection. The specific preparation method is as follows:

[0038] Take Conessine and add it to conventional excipients for preparing aerosols, such as propellants and various additives.

[0039] Example 5

[0040] Example 5 of this invention utilizes Conessine to prepare a capsule-form antiviral drug for influenza A virus infection. The specific preparation method is as follows:

[0041] Take Conessine, add it to the conventional excipients for preparing capsules, mix well, and quantitatively fill it into capsule shells made primarily of gelatin and seal.

[0042] Experimental Example 1

[0043] Experiment 1 of this invention tested the inhibitory effect of Conessine on IAV and its cytotoxicity. The specific method is as follows:

[0044] (1) Inhibition effect detection:

[0045] Conessine was diluted in Mock DMEM to concentrations of 50 μM, 25 μM, 10 μM, 5 μM, 2.5 μM, 1 μM, 0.1 μM, and 0.01 μM. MDCK cells were treated with different concentrations of Conessine, and simultaneously infected with IAV-Glucose. After 24 h of infection, luciferase activity was measured and the values ​​were recorded. Curve analysis was then performed to determine the IC50 value at which the small molecule concentration achieved a 50% inhibition rate. 50 The results are shown Figure 1 .

[0046] (2) Cytotoxicity detection:

[0047] Conessine was diluted in Mock DMEM to concentrations of 1000 μM, 100 μM, 50 μM, 25 μM, 10 μM, 5 μM, 2.5 μM, 1 μM, 0.33 μM, 0.1 μM, and 0.01 μM. Cells were incubated with different concentrations of Conessine, and five groups of cells under the same conditions were set up as an average control. Cytotoxicity was measured using the CCK8 kit (Enhanced Cell Counting Kit-8, Beyotime Biotechnology). Cytotoxicity curves of this small molecule were plotted, and the results were analyzed to finally determine the concentration value (CC) at which the cytotoxicity reached half. 50 The result is as follows Figure 1 As shown.

[0048] Depend on Figure 1 It is known that Conessine has a strong inhibitory effect on influenza A virus, IC50. 50＝ 2.07 μM, a relatively low value. CC 50 =34.11μM, a relatively high value, indicates low cytotoxicity. The selectivity index (SI) is much higher than the safety range, which further ensures the safety and efficacy of Conessine as an inhibitor of IAV influenza A virus.

[0049] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. The application of Conessine in the preparation of drugs against H1N1 influenza virus infection, characterized in that, The structure of Conessine is shown in formula (I): Formula (I).

2. The application of Conessine according to claim 1 in the preparation of drugs against H1N1 influenza virus infection, characterized in that, Conessine can inhibit the infection and replication of the H1N1 influenza virus.

3. The application of Conessine according to claim 2 in the preparation of drugs against H1N1 influenza virus infection, characterized in that, The anti-H1N1 influenza virus infection drug includes a pharmaceutically acceptable carrier.

4. The application of Conessine according to claim 3 in the preparation of drugs against H1N1 influenza virus infection, characterized in that, The dosage form of the drug for treating H1N1 influenza A virus infection is one of the following: powder, solution, capsule, granule, tablet, emulsion, or suspension.

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