Application of morusin M in preparation of anti-HSV-1 drugs

Abstract

The invention discloses application of morusin M in preparation of anti-HSV-1 drugs, and belongs to the technical field of biological medicines. Based on the discovery that morusin M has anti-HSV-1 activity, a new candidate medicine is provided for treating HSV-1 infection, meanwhile, the morusin M is very low in cytotoxicity, has a remarkable inhibition effect on HSV-1 even at a relatively low concentration, and can inhibit nerve cell infection caused by HSV-1, so that the morusin M has an application prospect in the field of treatment of HSV-1 infection.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of biological medicine, in particular to application of moracin M in preparation of anti-HSV-1 drugs. BACKGROUND

[0002] Herpes simplex virus 1 (HSV-1) and herpes simplex virus 2 (HSV-2) belong to the herpesviridae family, HSV-1 and HSV-2 are highly prevalent in humans, according to the statistics of the World Health Organization, the global infection rate is 70%. HSV-1 and HSV-2 can cause serious diseases in immunocompetent adults and neonates, such as herpes encephalitis caused by HSV-1, although antiviral therapy is performed, but still have sequelae. HSV-1 can also cause eye infections, leading to visual impairment, and currently HSV-1 is the main cause of infectious blindness in developed countries. In addition, HSV-1 can also cause oral and facial lesions, and the clinical manifestation is labial herpes. After HSV-1 infection, it will be latent in the trigeminal ganglion, when it is reactivated, the virus body moves to the epithelial cells and fibroblasts near the original site in a retrograde manner from the trigeminal ganglion (oral and facial infections) or dorsal root ganglion (genital-related infections) infected neurons, and re-infects new cells to cause lesions.

[0003] HSV-1 virus body is a spherical particle with an average diameter of 186 nm, which includes four parts: core, capsid, envelope and cortex. Acyclovir (ACV) is an effective clinical drug for treating HSV, and is widely used in the world. With the widespread use of ACV, it has also brought a difficult problem to human beings, that is, herpes virus drug-resistant strains gradually appear, and in recent years there is still an increase, this thorny problem prompts researchers to find new anti-HSV drugs, and these new drugs must be different from the mechanism of action of ACV, so it is necessary to explore new drugs through different ways.

[0004] Moracin M is a phenolic component extracted from sarsaparilla, which is reported to have anti-inflammatory activity. At present, there is no report that moracin M has anti-herpes simplex virus effect. SUMMARY

[0005] Based on the finding that moracin M can inhibit virus HSV-1, the present application provides a scheme for preparing anti-HSV-1 drugs using moracin M, which provides a new drug for treating HSV infection.

[0006] The technical scheme of the present application is as follows: In a first aspect, the present application provides the use of moracin M and / or a pharmaceutically acceptable salt thereof in the preparation of an anti-HSV-1 drug.

[0007] In the above scheme, the anti-HSV-1 medicine can further contain pharmaceutically acceptable adjuvants (such as disintegrants, fillers, stabilizers, etc.), and the dosage form of the anti-HSV-1 medicine includes, but is not limited to, tablets, sprays, granules, capsules, oral liquids, injections, etc.

[0008] In a second aspect, the present application provides use of morusin M and / or a pharmaceutically acceptable salt thereof in the preparation of a reagent for inhibiting the titer of HSV-1 virus.

[0009] In a third aspect, the present application provides use of morusin M and / or a pharmaceutically acceptable salt thereof in the preparation of a reagent for inhibiting the infection of HSV-1 to nerve cells. For example, in an embodiment of the present application, 12.5-50 μM of morusin M can significantly inhibit the infection of HSV-1 to sy5y cells.

[0010] In a fourth aspect, the present application provides a medicine for treating HSV-1 infection, and the medicine takes morusin M as one of the active ingredients. It can be understood that the medicine can further contain pharmaceutically acceptable adjuvants, and the dosage form thereof can be set according to actual needs.

[0011] Compared with the prior art, the present application has the following beneficial effects: Based on the finding that morusin M has anti-HSV-1 activity, the present application provides a new candidate drug for treating HSV-1 infection, which has a positive significance for the treatment of HSV-1. Morusin M has very low cytotoxicity and can significantly inhibit HSV-1 at a lower concentration, so it has excellent biological safety performance. In addition, morusin M can effectively inhibit the infection of HSV-1 to nerve cells, so morusin M has a good application prospect in the field of treating HSV-1 infection. BRIEF DESCRIPTION OF DRAWINGS

[0012] In order to more clearly illustrate the technical solutions of the present application, the drawings used in the present application will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present application, and other drawings can be obtained by those skilled in the art without creative labor on the basis of these drawings.

[0013] Figure 1 is a graph of the detection results of the survival rate of Vreo cells under different concentrations of morusin M in Example 1; Figure 2 is a graph of the calculation results of the CC 50 (half of the toxic concentration, Median cyctoxic concentration) of morusin M to Vreo cells in Example 1; Figure 3Figure of the results of the detection of the survival rate of sy5y cells in Example 1 under different concentrations of moracin M; Figure 4 Figure of the CC of moracin M on sy5y cells in Example 1; 50 Figure of the calculation results; Figure 5 Figure of the fluorescence microscopic observation results of Vreo cells after treatment with different concentrations of moracin M in Example 2; Figure 6 Figure of the flow cytometry detection results of Vreo cells after treatment with different concentrations of moracin M in Example 2; Figure 7 Figure of the fluorescence microscopic observation results of sy5y cells after treatment with different concentrations of moracin M in Example 2; Figure 8 Figure of the viral genome copy number results of Vreo cell samples after treatment with different concentrations of moracin M in Example 2; Figure 9 Figure of the viral genome copy number results of sy5y cell samples after treatment with different concentrations of moracin M in Example 2. DETAILED DESCRIPTION

[0014] The technical solutions of the present application will be described in detail below in combination with the drawings and examples.

[0015] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the present application belongs; the terms used herein are only for the purpose of describing specific embodiments and are not intended to limit the present application; the terms "include" and "have" and any variations thereof in the present document are intended to cover non-exclusive inclusion.

[0016] In order to solve the problem of drug resistance of existing herpes viruses to its clinical drug acyclovir, the present application discovers a new small molecule substance with anti-HSV-1 activity, i.e. moracin M, the structural formula of which is as follows: .

[0017] Moracin M has been reported to have anti-inflammatory activity, but there is no related report on its anti-herpes virus. The example data of the present application show that moracin M has excellent inhibitory activity on HSV-1 even at a lower administration concentration, so it is expected to be a candidate drug for treating HSV-1 infection.

[0018] Some specific examples are listed below. It should be noted that the examples described below are exemplary and are used to explain the present application, and are not to be construed as limiting the present application. The specific techniques or conditions described in the examples are not to be construed as limiting the present application unless otherwise specified. If a specific technique or condition is not described in the examples, it is performed according to the techniques or conditions described in the literature or according to the product manual. If the reagent or instrument used is not specified by the manufacturer, it is a conventional product that can be obtained on the market.

[0019] Example 1 This example tests the cytotoxicity of morusin M, and the testing process is as follows: Vero cells were plated in a 96-well plate and incubated at 37°C in a 5% CO2 cell incubator for about 12 h. The cell culture medium was discarded, and cell maintenance solution containing different concentrations of morusin M (0 μM, 12.5 μM, 25 μM, 50 μM, 100 μM, 125 μM, 150 μM, 200 μM) was added, respectively, and incubated for 24 h. The cell survival rate was determined by CCK-8 method.

[0020] Vero cells were plated in a 96-well plate and incubated at 37°C in a 5% CO2 cell incubator for about 12 h. The cell culture medium was discarded, and cell maintenance solution containing different concentrations of morusin M (0 μM, 12.5 μM, 25 μM, 50 μM, 100 μM, 125 μM, 150 μM, 200 μM) was added, respectively, and incubated for 24 h. The cell survival rate was determined by CCK-8 method.

[0021] The cell survival rate was calculated according to the following formula: cell survival rate = (drug group average OD 450 - blank group average OD 450 / cell control group average OD 450 - blank group average OD 450 ) x 100%, wherein the control group is the single cell without drug group, and the blank group is the PBS group.

[0022] The cell survival rate results under different concentrations of morusin M are shown in Figures 1-4 According to the results of Figure 1 and Figure 3 , the CC 50 of morusin M for Vero cells and sy5y cells was calculated, respectively, and the results are shown in Figure 2 and Figure 4 : the CC 50 of morusin M for Vero cells and sy5y cells is 163.9 μM and 142.6 μM, respectively.

[0023] The above results show that morusin M has low toxicity to cells, and has good biological safety.

[0024] Example 2 This example detects the inhibitory effect of morusin M on HSV-1-GFP (laboratory- engineered virus, i.e., green fluorescent protein GFP is added to the virus HSV-1 to facilitate the reflection of infection rate according to fluorescence; the construction of HSV-1-GFP can refer to the prior art, and this example will not be described in detail) by in vitro cell test, and the test process is as follows: Vero cells were seeded in 12-well plates, and the cells were grown to 90% every other day, and then infected with HSV-1-GFP (MOI = 0.5); 2 hours later, the culture medium was discarded, and different concentrations of morusin M (0 μM, 12.5 μM, 25 μM, 50 μM) were added to treat the cells; 24 hours later, the green light expression of the virus was observed by fluorescence microscope and detected by flow cytometry, respectively.

[0025] Vero cells were seeded in 12-well plates, and the cells were grown to 90% every other day, and then infected with HSV-1-GFP (MOI = 0.5); 2 hours later, the culture medium was discarded, and different concentrations of morusin M (0 μM, 12.5 μM, 25 μM, 50 μM) were added to treat the cells; 24 hours later, the green light expression of the virus was observed by fluorescence microscope and detected by flow cytometry, respectively.

[0026] Figure 5 For the results of Vero cell infection observed by fluorescence microscope, Figure 6 For the results of Vero cell infection observed by fluorescence microscope, Figure 7 For the results of Vero cell infection observed by fluorescence microscope, Figures 5-7 As shown in the table, with the increase of the concentration of morusin M, the detected green fluorescent signal is weaker, indicating that the infection rate is lower.

[0027] In addition, after 24 h of morusin M treatment, sample DNA was extracted and the copy number of viral genomic DNA was detected by fluorescence quantitative PCR. The results are shown in Figure 8 and Figure 9 The copy number of viral genomic DNA is reduced.

[0028] The above results show that morusin M can significantly inhibit the viral titer of HSV-1, and is dose-dependent.

[0029] In summary, morusin M has good biological safety, and has a significant inhibitory effect on HSV-1, so it has application potential in the field of treating HSV-1 infection.

[0030] Note that the present application is not limited to the above-described embodiments. The above-described embodiments are merely examples, and embodiments having substantially the same configuration as the technical idea and achieving the same effects within the scope of the technical idea of the present application are included in the technical scope of the present application. Furthermore, other modes constructed by applying various modifications that can be thought of by those skilled in the art to the embodiments or by combining part of the constituent elements of the embodiments are also included in the scope of the present application without departing from the spirit of the present application.

Claims

1. The use of sancin M and / or its pharmaceutically acceptable salts in the preparation of anti-HSV-1 drugs.

2. The application according to claim 1, characterized in that, The anti-HSV-1 drug contains pharmaceutically acceptable excipients.

3. The application according to claim 1, characterized in that, The dosage form of the anti-HSV-1 drug is selected from any one of tablets, sprays, granules, capsules, oral liquids, and injections.

4. The use of sancin M and / or its pharmaceutically acceptable salts in the preparation of reagents for inhibiting HSV-1 viral titers.

5. Use of sancin M and / or its pharmaceutically acceptable salts in the preparation of reagents that inhibit HSV-1 infection of nerve cells.

6. The application according to claim 5, characterized in that, The nerve cells include sy5y cells.

7. The application according to claim 5, characterized in that, In the reagent, the concentration of sanguisorbin M is 12.5-50 μM.

8. A drug, characterized in that, The drug is used to treat HSV-1 infection, and one of the active ingredients of the drug is sanguisorbin M.

9. The medicament according to claim 8, characterized in that, The drug contains pharmaceutically acceptable excipients.

10. The medicament according to claim 8, characterized in that, The dosage form of the drug is selected from any one of tablets, sprays, granules, capsules, oral liquids, and injections.

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