Application of sodium new houttuyfonate in preparation of medicine for resisting respiratory syncytial virus

Abstract

The invention belongs to the technical field of medicines, and particularly relates to application of sodium new houttuyfonate in preparation of a medicine for resisting respiratory syncytial virus. The cytotoxicity of the sodium new houttuyfonate and the inhibition effect on the RSV virus are tested through a cell experiment, and the test finds that the sodium new houttuyfonate has an obvious inhibition effect on the respiratory syncytial virus B strain, is low in biotoxicity and provides a new drug choice for clinical treatment of the respiratory syncytial virus.

Description

Technical Field

[0001] The invention relates to the field of medical technology, and in particular to application of new houttuynia cordata sodium in the preparation of anti-respiratory syncytial virus drugs. Background Art

[0002] Houttuynia cordata, also known as Houttuynia cordata, Houttuynia cordata, and Dog Ear, is a perennial herb. Its active ingredient is sodium houttuyfonate, which is extremely unstable in chemical properties. SodiuM newhouttuyfonate (SNH) is a sodium bisulfite adduct of houttuyfonate, which is more stable than sodium houttuyfonate and has stronger pharmacological effects. SodiuM newhouttuyfonate has strong antibacterial and disinfection capabilities, and has obvious inhibitory effects on Candida albicans, Pseudomonas aeruginosa, Staphylococcus aureus, etc.

[0003] Respiratory syncytial virus (RSV) is the main cause of hospitalization for lower respiratory tract infection (LRTI) in children. RSV infection can cause repeated wheezing, asthma, decreased respiratory function, and increased sensitivity to allergic reactions in children. These can lead to a significant decline in the quality of life of children and increase the cost of utilizing healthcare resources. With age, the RSV infection rate in adults will gradually increase. Studies have shown that the number of elderly people infected with RSV in some developed countries exceeds five million cases per year, and the number of deaths caused by RSV infection in developing countries is much higher than that in developed countries. At present, the effective prevention and treatment drugs for RSV are ribavirin with relatively large side effects and expensive palivizumab, which are difficult to promote and apply. Compared with Western medicine, Chinese medicine is pure and natural, non-toxic and non-residual, non-resistant, and functionally diverse.

[0004] Therefore, it is of great social significance to develop low-toxic and highly effective anti-RSV drugs to meet the clinical needs of infected patients at home and abroad. Summary of the invention

[0005] In order to solve the problems of the prior art, the main purpose of the present invention is to use new houttuynia cordata sodium in the preparation of anti-respiratory syncytial virus drugs. The present invention has confirmed through cell experiments that new houttuynia cordata sodium can inhibit RSV virus infection in a dose-dependent manner in vitro, providing a theoretical basis for the development of new houttuynia cordata sodium as an anti-RSV virus drug.

[0006] To achieve the above purpose, the technical solution adopted by the present invention is: Application of new houttuynia cordata sodium in the preparation of anti-respiratory syncytial virus drugs.

[0007] Furthermore, the anti-respiratory syncytial virus drug is used to prevent or treat respiratory syncytial virus B strain infection.

[0008] Furthermore, the drug is a preparation prepared with new houttuynia cordata sodium as the only active ingredient and pharmaceutically acceptable excipients added.

[0009] Furthermore, the content of the active ingredient in the preparation is ≧0.1 μM, preferably 0.1-100 μM, and more preferably 20-100 μM.

[0010] Compared with the prior art, the present invention has the following beneficial effects: The new sodium houttuynia cordata of the invention comes from the medicinal plant houttuynia cordata, is easy to obtain, has low production cost, has strong inhibitory ability against respiratory syncytial virus (B strain), and can be developed into an anti-RSV specific drug with high economic benefits. BRIEF DESCRIPTION OF THE DRAWINGS

[0011] The present invention will be further described below in conjunction with the accompanying drawings: Figure 1 is the cytotoxic effect of different concentrations of SNH; Figure 2 is a graph showing the antiviral effect of different concentrations of SNH / RBV on RSV-B; Figure 3 The antiviral effect of different concentrations of SNH / RBV on RSV-A2. DETAILED DESCRIPTION

[0012] The technical scheme of the present invention is specifically described below in conjunction with the embodiments, but the protection scope of the present invention is not limited thereto. Unless otherwise specified, the raw materials used in the present invention are all known products, obtained by purchasing commercially available products.

[0013] The sources of some raw materials used in the present invention are as follows: Sodium houttuyfonate was purchased from Sichuan Weikeqi Biotechnology Co., Ltd.; Respiratory syncytial virus RSV-A2 strain was provided by Guangzhou National Laboratory; Respiratory syncytial virus RSV-B strain was provided by Wuhan Institute of Virology, Chinese Academy of Sciences.

[0014] Example 1: Cytotoxicity Assay 1.1 Cell culture and plating Hep-2 cells were seeded in 96-well plates (1×10 4 The cells were cultured overnight in a 37°C, 5% CO2 incubator.

[0015] 1.2 Preparation of different concentrations of new houttuynia cordata sodium The new houttuynia cordata sodium powder was dissolved in DMSO to prepare a 50mM stock solution. In the experiment, the drug stock solution was continuously diluted with DMEM solution containing 2% FBS into 8 gradient drug dilutions (0.01, 0.1, 1, 10, 20, 50, 100, 250μM), and added to a 96-well plate (100μl / well). At the same time, an equal volume of control wells was prepared, i.e., only DMSO group (final concentration 0.5%), and three replicate wells were set for each group.

[0016] 1.3 Effects of different concentrations of new houttuynia cordata sodium on Hep-2 cells Use a pipette to discard the culture medium in the 96-well plate containing Hep-2 cells, add the prepared drug dilutions respectively, and culture in a 37°C, 5% CO2 incubator for 48 hours.

[0017] 1.4 Drug cytotoxicity testing The cytotoxicity detection kit (CCK8 kit) was used to evaluate the toxicity of different concentrations of drugs on cells. The specific operation is as follows: Mix the CCK8 solution and sterile PBS in a biological safety cabinet at a ratio of 1:10 by pipetting and mixing. Use a pipette to discard the Chinese medicine dilution in the 96-well plate, add 100 μl of the prepared CCK8 detection solution to each well, place the culture plate in a 37°C, 5% CO2 incubator for 1.5 hours, place the culture plate in an ELISA reader and shake it slowly for 1 minute, and measure the optical density (OD value) at 450 nm.

[0018] The cell viability calculation formula is: .

[0019] Example 2: Detection of drug inhibition rate on virus 2.1 Cell plating Hep-2 cells were seeded in 96-well plates (1×10 4 The cells were cultured overnight in a 37°C, 5% CO2 incubator.

[0020] 2.2 Preparation of different concentrations of new houttuynia cordata sodium dilution and virus mixture SNH powder and ribavirin (RBV, positive control drug) were dissolved in DMSO to prepare a 50 mM stock solution. In the experiment, DMEM solution containing 2% FBS was used to continuously dilute the drug into 6 gradient dilutions (0.2, 2, 20, 40, 100, 200 μM) and added to a 96-well plate (50 μl / well). Then, 50 μl of RSV-B virus solution (Hep-2 cells were infected with an infection multiplicity of MOI = 0.1) was added and gently pipetted to mix, so that the final concentration of the drug was: 0.1, 1, 10, 20, 50, 100 μM; at the same time, 4 groups of control wells were prepared with equal volumes, including a virus-only group (final concentration of 1×10 3 PFU / well of RSV-B virus solution), DMSO only group (final concentration 0.5%), DMSO+virus group, and 2% DMEM blank group (cell control group), with three replicate wells in each group.

[0021] 2.3 Infection of Hep-2 cells with different concentrations of new houttuynia cordata sodium and RSV virus mixture Use a pipette to discard the culture medium in the cell plate, add the prepared mixture of drugs and viruses, and culture in a 35°C, 5% CO2 incubator for 6 days.

[0022] 2.4 Antiviral Detection A cell viability assay kit (CellTiter-Glo 2.0) was used to determine the neutralizing ability of different concentrations of traditional Chinese medicine against viruses. The more cells that survive, the stronger the neutralizing ability of the drug at that concentration against RSV. The specific operation is as follows: aspirate the drug and virus mixture in the 96-well plate, wash the plate twice with sterile PBS, and add 100μl 2% DMEM culture medium and an equal volume of CellTiter-Glo detection solution to each well in the plate. After incubating the culture plate at room temperature and away from light for 10 minutes, use a pistol to mix and transfer 50μl of the liquid to a transparent white well plate. Use an ELISA reader to slowly shake the white well plate for 1 minute and measure the chemiluminescence value.

[0023] The formula for calculating the inhibition rate of the drug on the virus is: .

[0024] The experimental results are as follows Figure 1-3 As shown in Figure 2, SNH has an obvious effect on the CC of Hep-2 cells. 50 >100μM, indicating that SNH has low cytotoxicity and good biosafety. The inhibitory effect of SNH on RSV-B is concentration-dependent, and the anti-RSV virus effect at a concentration of 20μM is better than that of RBV, indicating that SNH has a strong antiviral effect; according to Figure 1 and Figure 2 CC 50 and EC50 It can be concluded that the SI of SNH against RSV-B virus is > 7.58, indicating that SNH has high biosafety, meets the requirements of clinical candidate drugs, and has the potential to be developed as a specific drug against RSV virus. At a concentration of less than 20 μM, SNH has almost no inhibitory effect on RSV-A2, and the inhibition rate is greater than 20% at a concentration of 100 μM. In the concentration range of 0.01-100 μM, the antiviral ability of SNH against RSV-A2 is not as good as that of RBV, indicating that SNH's inhibitory effect on RSV-A2 strains is weaker than that of RBV. It can be seen that at the same concentration, SNH's inhibitory effect on RSV-B is better than RSV-A, indicating that SNH is selective for RSV virus.

[0025] The above embodiments are only preferred technical solutions of the present invention and should not be regarded as limiting the present invention. The protection scope of the present invention shall be the technical solutions recorded in the claims, including equivalent replacement solutions of the technical features in the technical solutions recorded in the claims. That is, equivalent replacement improvements within this scope are also within the protection scope of the present invention.

Claims

1. Application of new houttuynia cordata sodium in the preparation of anti-respiratory syncytial virus drugs.

2. The use according to claim 1, characterized in that: The anti-respiratory syncytial virus drug is used for treating respiratory syncytial virus B strain infection.

3. The use according to claim 2, characterized in that: The drug is a preparation prepared by using new houttuynia cordata sodium as the only active ingredient and adding pharmaceutically acceptable excipients.

4. The use according to claim 3, characterized in that: The content of the active ingredient in the preparation is ≧0.1 μM.

5. The use according to claim 4, characterized in that: The content of the active ingredient in the preparation is 0.1-100 μM.

6. The use according to claim 5, characterized in that: The content of the active ingredient in the preparation is 20-100 μM.

Images