Application of CHIR-090 in preparation of pharmaceutical composition or vaccine for treating and / or preventing vibrio parahaemolyticus infection

Abstract

The invention discloses an application of CHIR-090 in preparation of a pharmaceutical composition or a vaccine for treating and / or preventing vibrio parahaemolyticus infection. The MIC value of the small molecule drug CHIR-090 is 0.2 [mu] M, and compared with some antibiotics such as sertraline and conicillacin which are clinically used at present and used for treating vibrio parahaemolyticus infection, the MIC value of the small molecule drug CHIR-090 is extremely small. Results of an in-vitro killing and bacteriostatic activity experiment and a motility experiment show that the small molecule drug CHIR-090 has a certain effect on reduction of the number of bacteria and limitation of movement. The experiment result of zebra fish infection also shows that the small molecule drug CHIR-090 has better safety and bacteriostatic activity. Therefore, the small molecule drug CHIR-090 has a relatively good potential application value in inhibiting the activity of the vibrio parahaemolyticus.

Description

Technical Field

[0001] The present invention relates to the use of CHIR-090 in the preparation of a pharmaceutical composition or vaccine for treating and / or preventing Vibrio parahaemolyticus infection, belonging to a new application field of CHIR-090. Background Art

[0002] Vibrio parahaemolyticus is an important foodborne pathogen. Consuming food contaminated by it may cause symptoms such as intestinal spasm, diarrhea, vomiting, etc. Severe patients may also experience dehydration, shock, coma, and even death. At the same time, the abuse of antibiotics has led to an increasingly serious problem of bacterial drug resistance. To address this issue, screening for new antibacterial drugs has become crucial, and drug libraries have become an important resource for finding potential antibacterial agents.

[0003] The molecular drug CHIR-090 is a novel N-arylthreonine hydroxamic acid and has been identified as a potent, slow, and tight-binding inhibitor of LpxC deacetylase in the hyperthermophilic bacterium Aquifex aeolicus. Its main target is LpxC. Research has shown that CHIR-090 exhibits good antibiotic activity against both Pseudomonas aeruginosa and Escherichia coli. Notably, CHIR-090 can inhibit LpxC homologs of a variety of Gram-negative pathogens at low nanomolar levels, including Neisseria meningitidis and Helicobacter pylori, etc. Relatively speaking, CHIR-090 is a weaker competitive inhibitor of LpxC from Rhizobium leguminosarum, so this plant symbiont is resistant to CHIR-090. The results of the disc diffusion experiment show that CHIR-090 exhibits significant antibiotic activity comparable to ciprofloxacin against E. coli and P. aeruginosa. However, there is no report on the antibacterial activity of CHIR-090 against Vibrio parahaemolyticus, especially the antibacterial effects of Vibrio parahaemolyticus from different sources. Summary of the Invention

[0004] Object of the Invention: The technical problem to be solved by the present invention is to provide the use of CHIR-090 in the preparation of a pharmaceutical composition or vaccine for treating and / or preventing Vibrio parahaemolyticus infection.

[0005] Technical Solution: To solve the above technical problem, the present invention provides the use of a small molecule drug CHIR-090 in the preparation of a pharmaceutical composition or vaccine for treating and / or preventing Gram-negative bacterial infection, and the Gram-negative bacteria include Vibrio or Enterobacter.

[0006] Among them, the vibrios include Vibrio parahaemolyticus, Vibrio anguillarum or Vibrio alginolyticus; the enterobacteria include Edwardsiella tarda, Salmonella pullorum, Salmonella enteritidis or Escherichia coli.

[0007] Among them, the Gram-negative bacteria also include Pseudomonas plecoglossicida.

[0008] Among them, the concentration of the small molecule drug CHIR-090 is 0.16 - 2.5 μM.

[0009] Among them, when the Gram-negative bacteria is Vibrio parahaemolyticus, the concentration of the small molecule drug CHIR-090 is 0.1 - 1 μM.

[0010] The present invention also provides a pharmaceutical composition or a vaccine, which contains the small molecule drug CHIR-090; the Gram-negative bacteria include vibrios or enterobacteria. The pharmaceutical composition or the vaccine can be used for treating and / or inhibiting Gram-negative bacterial infections, treating and / or preventing aquatic products from being infected with Vibrio parahaemolyticus or increasing the survival rate of aquatic products infected with Vibrio parahaemolyticus.

[0011] Among them, the dosage form of the pharmaceutical composition includes tablets, capsules, granules, aerosols, sprays or injections.

[0012] The present invention also provides an application of the small molecule drug CHIR-090 in the preparation of a pharmaceutical composition or a vaccine for treating and / or preventing aquatic products from being infected with Vibrio parahaemolyticus.

[0013] The present invention also provides an application of the small molecule drug CHIR-090 in increasing the survival rate of aquatic products infected with Vibrio parahaemolyticus.

[0014] In the present invention, the small molecule drug CHIR-090 that can inhibit the activity and motility of Vibrio parahaemolyticus provides a safe and effective drug product for the prevention and control of Vibrio parahaemolyticus. The small molecule drug is mainly used for treating bacterial infections, especially against Gram-negative bacteria. CHIR-090 affects the synthesis of outer membrane lipid A of Gram-negative bacteria by inhibiting the activity of LpxC enzyme, thereby playing an antibacterial role. It shows great potential in the treatment of multi-drug resistant Gram-negative bacterial infections.

[0015] Among them, the small molecule drug CHIR-090 belongs to small molecule chemical drugs in terms of structure, and its molecular structural formula is C 24 H 27 N3O5, with a molecular weight of 437.49 g / mol, and its structural formula is:

[0016]

[0017] Among them, the small molecule drug CHIR-090 is safe on HeLa cells.

[0018] Among them, the small molecule drug CHIR-090 is safe for aquaculture applications.

[0019] Among them, the small molecule drug CHIR-090 can be used for sterilization in aquaculture.

[0020] Among them, the main feature of the small molecule drug CHIR-090 is that it is an inhibitor of LpxC (UDP-(3-O-(R-3-hydroxymyristoyl))-N-acetylglucosamine deacetylase). This drug is mainly used for the treatment of bacterial infections, especially showing great potential in the treatment of multi-drug resistant Gram-negative bacterial infections. CHIR-090 exerts its antibacterial effect by inhibiting the activity of the LpxC enzyme, thereby affecting the synthesis of outer membrane lipid A of Gram-negative bacteria.

[0021] Among them, the small molecule drug CHIR-090 can be used as a control agent for Vibrio parahaemolyticus.

[0022] Among them, the small molecule drug CHIR-090 has a certain antibacterial effect on Vibrio parahaemolyticus from different host sources.

[0023] Among them, the optimal growth temperature of Vibrio parahaemolyticus is 37 °C. Among them, the optimal pH of Vibrio parahaemolyticus is 7.4 - 8.0.

[0024] Among them, the optimal salt concentration of Vibrio parahaemolyticus is 3.5%.

[0025] Among them, the incubation period of Vibrio parahaemolyticus is 8 - 40 h, up to 96 h at most. The typical clinical symptoms are diarrhea (watery or bloody stools), upper abdominal cramps, nausea and vomiting, fever, etc.

[0026] Beneficial effects: Compared with the prior art, the present invention has the following remarkable advantages: The small molecule drug CHIR-090 of the present invention can effectively inhibit various sequence types of Vibrio parahaemolyticus in a targeted manner and can be applied to the prevention and control of most Vibrio parahaemolyticus. Description of the Drawings

[0027] Figure 1 is the growth curve of Vibrio parahaemolyticus after treatment with different concentrations of CHIR-090;

[0028] Figure 2 is the antibacterial activity of the small molecule drug CHIR-090 against Vibrio parahaemolyticus;

[0029] Figure 3 is the effect of the small molecule drug CHIR-090 on the motility of Vibrio parahaemolyticus: Figure 3A is the swimming ability of Vibrio parahaemolyticus on LB plates containing 0.3% agar supplemented with different concentrations of CHIR-090; Figure 3 B is a bar graph plotted based on Figure A, with the abscissa representing the concentration of the added drug and the ordinate representing the diameter of the bacterial circle; Figure 3 C is the effect of different concentrations of CHIR-090 on the biofilm formation ability of the standard strain of Vibrio parahaemolyticus RIMD2210633;

[0030] Figure 4 is the effect of different concentrations of small molecule drugs on the cytotoxicity of HeLa cells;

[0031] Figure 5 is the antibacterial activity of different concentrations of small molecule drugs against Vibrio parahaemolyticus on the HeLa cell model;

[0032] Figure 6 is the antibacterial activity of different concentrations of small molecule drugs against Vibrio parahaemolyticus on the zebrafish model. Specific Embodiments

[0033] The technical solutions of the present invention will be further described below with reference to the accompanying drawings.

[0034] Unless otherwise specified, the reagents involved in the embodiments of the present invention are all commercially available products and can be obtained through commercial channels.

[0035] Example 1 Evaluation of the Antibacterial Activity of CHIR-090 against Vibrio parahaemolyticus

[0036] Using the standard strain of Vibrio parahaemolyticus RIMD2210633 preserved in the laboratory as the experimental object, the antibacterial activity of CHIR-090 against Vibrio parahaemolyticus was evaluated. The results are shown in Table 1. The MIC value of the small molecule drug CHIR-090 is 0.2 μM, indicating that this small molecule drug CHIR-090 can achieve a good antibacterial effect at a relatively low concentration.

[0037] Table 1 Determination Results of the MIC Value of the Small Molecule Drug CHIR-090

[0038]

[0039] Example 2 Determination of the Growth Curve

[0040] (1) Pick a single colony of Vibrio parahaemolyticus RIMD2210633 and inoculate it into LB liquid medium. Incubate overnight at 37 °C and 180 r / min;

[0041] (2) Inoculate the overnight-cultured strain into LB liquid medium containing different concentrations of CHIR-090 (0, 0.2, 0.1, and 0.02 μM), and adjust the initial OD of the bacterial liquid 600To 0.05, continuously culture at 37°C and 180 r / min for 24 h;

[0042] (3) Take samples every 1 h to measure the OD 600 value, measure continuously for 24 times, and draw the bacterial growth curve according to the OD 600 value at different time points.

[0043] The results of the growth curve are as Figure 1 shown. Adding CHIR-090 at the MIC (0.2 μM) concentration can effectively inhibit the growth of Vibrio parahaemolyticus RIMD2210633. When adding CHIR-090 at the 1 / 2 MIC (0.1 μM) concentration, the bacterial growth rate can be slowed down, and the number of bacteria at the plateau phase is reduced. When adding CHIR-090 at the 1 / 2 MIC (0.02 μM) concentration, the growth curve of Vibrio parahaemolyticus is similar to that of the wild strain, with no significant difference. The above results indicate that the growth of Vibrio parahaemolyticus RIMD2210633 can be significantly inhibited under the condition of lower concentration of CHIR-090.

[0044] Example 3 Determination of in vitro killing activity

[0045] (1) Pick a single colony of Vibrio parahaemolyticus RIMD2210633 and inoculate it into LB liquid medium, culture overnight at 37°C and 180 r / min;

[0046] (2) Expand the bacterial solution in 5 mL of LB liquid medium and culture at 37°C and 180 r / min for 4 h;

[0047] (3) Adjust the initial OD of the bacterial solution 600 to 0.1;

[0048] (4) Dilute CHIR-090 with PBS buffer in a serial dilution to MIC, 1 / 2 MIC, 1 / 10 MIC, mix it with the bacterial solution in step (3), and immediately perform gradient dilution and plate counting after mixing, which is the 0 hour;

[0049] (5) At 1 h, 3 h, and 5 h after adding the bacteria, take samples for gradient dilution and plate, culture in a 37°C microbial constant temperature incubator for 24 h, and calculate the antibacterial activity.

[0050] The results are as Figure 2 shown. After adding small molecule drugs at different concentrations, the number of bacteria can be significantly reduced after 1 h, but increasing the culture time will not further reduce the number of bacteria. For the small molecule drug CHIR-090 at 1 h, the high concentration of the drug (MIC) can significantly reduce the number of bacteria compared with the low concentration of the drug (1 / 10 MIC), but there is no significant difference at subsequent time points.

[0051] Example 4: Effect of CHIR-090 on the motility of Vibrio parahaemolyticus

[0052] LBA plate with 0.3% Agar (motility): 5 g NaCl, 5 g tryptone, 2.5 g yeast extract, 1.5 g agar powder and 500 mL distilled water, autoclaved at 121 °C for 20 min.

[0053] (1) Take out Vibrio parahaemolyticus RIMD2210633 from the -70 °C refrigerator and streak it onto an LB plate with 1.5% agar, and culture it at 37 °C for 12 h;

[0054] (2) Pick a single colony and inoculate it into LB liquid medium, culture it overnight at 37 °C with 180 rpm, wash the bacterial solution twice with PBS, centrifuge it at 8000 rpm for 2 min, and adjust OD 600 = 1;

[0055] (3) Take 2.5 μL of the bacterial solution and spot it onto LBA plates with 0.3% Agar containing CHIR-090 drug concentrations of 1 / 2 MIC, 1 / 5 MIC, 1 / 10 MIC and no drug added, and culture it in an incubator at 37 °C for 12 h;

[0056] (4) Observe the growth morphology of the strain and measure the diameter of the bacterial halo.

[0057] The results are as shown in Figure 3 Figures A and 3B. As the drug concentration increases, the diameter of the bacterial halo formed by Vibrio parahaemolyticus significantly decreases. The diameters of the bacterial halos in the Control group and the 1 / 10 MIC group both reach the maximum range of the culture medium. It can be considered that when the drug concentration is 1 / 10 MIC, there is basically no inhibitory effect on the motility of Vibrio parahaemolyticus, and the drug concentrations of 1 / 5 MIC and 1 / 2 MIC have an obvious inhibitory effect on the motility of Vibrio parahaemolyticus, indicating that the small molecule drug CHIR-090 can significantly inhibit the motility of Vibrio parahaemolyticus at the concentrations of 1 / 5 MIC and 1 / 2 MIC. The detection results of the biofilm show that CHIR-090 has no significant effect on the biofilm formation ability of Vibrio parahaemolyticus ( Figure 3 Figure C).

[0058] Example 5: Evaluation of the safety of small molecule drugs at the cellular level

[0059] Treat HeLa cells cultured overnight with different concentrations of antibacterial drugs to evaluate the cytotoxicity of CHIR-090.

[0060] (1) Seed HeLa cells at a density of 2×10 5 cells / well in a 24-well plate and culture them overnight in an incubator at 37 °C with 5% CO2;

[0061] (2) Take out the cell culture plates, add HeLa cells treated with 0, MIC, 5MIC, and 10MIC of CHIR-090 respectively, and set up maximum release wells at the same time. Place them in an incubator at 37°C and 5% CO2 and continue to culture for 2 h.

[0062] (3) In the maximum release wells, add 10% of the LDH release agent of the original culture volume. After mixing well, place them in an incubator at 37°C and 5% CO2 and continue to culture for 1 h.

[0063] (4) Centrifuge with a multi-well plate centrifuge at 400×g for 5 min, and transfer 120 μL of the supernatant from each well to a new 96-well plate respectively.

[0064] (5) Add 60 μL of the LDH detection working solution to each well, mix well, avoid light at room temperature for 30 min, measure the absorbance at 490 nm, and perform dual-wavelength measurement with 600 nm as the reference wavelength.

[0065] (6) Cytotoxicity (%) = (absorbance of the treated sample - absorbance of the sample control) / (absorbance of the maximum enzyme activity - absorbance of the sample control) × 100%.

[0066] The results are as Figure 4 shown. After treating with CHIR-090 at concentrations of MIC, 5MIC, and 10MIC for 3 h, there was no significant difference in the LDH release amount of HeLa cells compared with the untreated group (Control group), indicating that the small molecule drug CHIR-090 has good safety in the HeLa cell model.

[0067] Example 6 Evaluation of the antibacterial activity of small molecule drugs at the cellular level

[0068] Use Vibrio parahaemolyticus RIMD2210633 and antibacterial drug CHIR-090 at different concentrations to treat HeLa cells cultured overnight to evaluate the antibacterial activity of the antibacterial drug at the cellular level.

[0069] (1) Seed HeLa cells at a density of 2×10 5 cells / well in a 24-well plate and culture them overnight in an incubator at 37°C and 5% CO2.

[0070] (2) Pick a single colony of Vibrio parahaemolyticus RIMD2210633 and inoculate it into LB liquid medium, and continue to incubate overnight at 37°C and 180 r / min.

[0071] (3) Inoculate the cultured strain into fresh LB liquid medium at a ratio of 1:100 and culture at 37°C for 3 h.

[0072] (4) Centrifuge at 8000 r / min for 2 min to collect the bacterial cell precipitate. Discard the supernatant, wash twice with sterile PBS, resuspend in Opti medium, and adjust the OD 600 to 1.0;

[0073] (5) Aspirate the original cell culture medium and rinse once with DPBS. Add 1 mL of Opti medium containing the bacterial solution (MOI = 100), and simultaneously add CHIR-090 at different concentrations (0, MIC, 5MIC, and 10MIC). Also set up a control group. Centrifuge at 1000 r / min for 10 min at 23°C, and then place in a 37°C, 5% CO2 incubator for 2 h of conventional culture; measure the cytotoxicity by the same method as in Example 5.

[0074] The results are as Figure 5 shown. The control group is the cytotoxicity control group when neither bacteria nor drugs are added. After infection with Vibrio parahaemolyticus RIMD2210633, the cytotoxicity of HeLa cells was significantly enhanced, indicating that Vibrio parahaemolyticus infection can enhance cytotoxicity. Compared with the group infected with Vibrio parahaemolyticus alone, in the groups treated with different concentrations of the small molecule drug CHIR-090, the cytotoxicity of HeLa cells was significantly reduced and was similar to that of the control group. The above results indicate that the small molecule drug CHIR-090 has significant antibacterial activity in the HeLa cell model.

[0075] Example 7 Evaluation of the antibacterial activity of CHIR-090 in a zebrafish model

[0076] The experimental animals were 3-month-old wild-type zebrafish purchased from Shandong Yixiyue Biotechnology Co., Ltd. The zebrafish were set up with a PBS control group, MIC, 5MIC, and 10MIC treatment groups, a Vibrio parahaemolyticus RIMD221063 challenge group (WT group), and WT challenge groups after treatment with MIC, 5MIC, and 10MIC (WT+MIC group, WT+5MIC group, and WT+10MIC group, respectively). Stop feeding 24 h before challenge.

[0077] (1) Take an appropriate amount of the small molecule drug CHIR-090 and prepare working solutions at concentrations of MIC, 5MIC, and 10MIC respectively. Treat with CHIR-090 before WT challenge;

[0078] (2) Streak Vibrio parahaemolyticus RIMD221063 onto an LBA plate and culture at 37°C for 12 h;

[0079] (3) Pick a single colony and inoculate it into LB liquid medium and culture overnight at 37°C and 180 rpm;

[0080] (4) Collect 2 mL of each bacterial solution. After centrifugation to discard the supernatant, wash twice with sterile PBS solution and adjust the OD600 to 1.0, diluted to 10 8 CFU / mL for standby;

[0081] (5) Inject 5 μL of the diluted bacterial solution into zebrafish, place it in an aquarium at 26 - 28 °C and observe for 7 days, record the onset situation and death time.

[0082] In this invention, zebrafish was used as an animal model to evaluate the safety and antibacterial activity of the small molecule drug CHIR - 090 on the animal model. Zebrafish treatment groups with PBS, MIC, 5MIC, and 10MIC were respectively set up to evaluate the safety of CHIR - 090. The results are as Figure 6 shown. The survival rate of zebrafish after treatment with different concentrations of CHIR - 090 was consistent with that of PBS, both being 100%, indicating that CHIR - 090 has good safety on zebrafish. Subsequently, the antibacterial activity of CHIR - 090 against Vibrio parahaemolyticus was evaluated on zebrafish. The results are as Figure 6 shown. In the wild - type strain (WT) infection group of Vibrio parahaemolyticus, the survival rate of zebrafish was 26.3%. While in the WT - infected groups treated with MIC and 5MIC concentrations of CHIR - 090, the survival rates of zebrafish were 30.0% and 21.7% respectively, similar to the survival rate of the drug - untreated group. In the 10MIC pretreatment group, the survival rate of zebrafish was significantly increased, reaching 64.3%. The above results indicate that the small molecule drug CHIR - 090 can significantly inhibit the lethality of Vibrio parahaemolyticus to zebrafish at a concentration of 10MIC.

[0083] Example 8 Antibacterial effect of CHIR - 090 against 60 Vibrio parahaemolyticus strains from different sources

[0084] In this example, 60 Vibrio parahaemolyticus isolates purified from seawater farms, freshwater farms, seafood, freshwater aquatic products, and human clinical diarrhea samples in areas such as Yangzhou, Nantong, and Taizhou were selected (published in two papers respectively: DOI: 10.1016 / j.fm.2023.104277; DOI: 10.27441 / d.cnki.gyzdu.2024.001069) to determine the antibacterial effect.

[0085] (1) Take out 60 Vibrio parahaemolyticus strains from different sources from the - 70 °C refrigerator and streak them onto LBA plates, and culture at 37 °C for 12 h;

[0086] (2) Respectively pick single colonies and inoculate them into MH liquid medium, and culture overnight at 37 °C with 180 rpm;

[0087] (3) Respectively take appropriate amounts of the antibacterial drug CHIR - 090 and dilute them to concentrations of 5MIC, 2MIC, MIC, and 1 / 2MIC to prepare working solutions;

[0088] (4) Take a sterile 96-well plate and dilute the drug in a biosafety cabinet. The specific operation is as follows: Add 100 μL of the drug to the first to fourth columns and the seventh to tenth columns in the order of decreasing concentration, where the fifth and eleventh columns are the control groups without adding the drug;

[0089] (5) Take an appropriate amount of bacteria cultured overnight, adjust OD 600 = 1.0, dilute 5 times for use, and take 100 μL of the bacterial solution and add it to the diluted drug above. The sixth and twelfth columns are the control groups without adding bacteria;

[0090] (6) Place the 96-well plate in an incubator and incubate for 12 hours to observe the growth of the bacteria.

[0091] The results are shown in Table 2. Except for 2 clinical isolates (YZU2256 and YZU2302), 1 freshwater farm isolate (YZU5311), and 3 seawater farm isolates (YZU5177, YZU5360, and YZU5362) that can still grow normally under conditions greater than 5 times the MIC, the small molecule drug CHIR-090 has good inhibitory effects on other Vibrio parahaemolyticus isolates, indicating that CHIR-090 has potential application value for the prevention and control of Vibrio parahaemolyticus in different scenarios.

[0092] Table 2 Inhibitory effects of the small molecule drug CHIR-090 on different species of Vibrio parahaemolyticus

[0093]

[0094]

[0095]

[0096] Example 9 Antibacterial effects of CHIR-090 against different Vibrio and Salmonella

[0097] In this example, 9 Gram-negative bacteria were selected for the test of antibacterial effect determination. The highest drug concentration was set at 10 μM and diluted in gradient successively. Drug concentrations above 10 μM were considered to have no obvious inhibitory effect.

[0098] (1) Take Salmonella pullorum S06004, Salmonella enteritidis Z11, Escherichia coli ATCC25922, Vibrio anguillarum MVM425, Edwardsiella tarda EIB202, Staphylococcus aureus USA300, Campylobacter jejuni 81-176, Vibrio alginolyticus EPGS, Pseudomonas plecoglossicida XSDHY-P from a -70°C refrigerator, streak them onto LB plates respectively, and culture them overnight in an incubator at 37°C and 180 rpm or 30°C and 150 rpm;

[0099] (2) Pick a single colony and inoculate it into MH liquid medium, and incubate it overnight in an incubator at 37°C with 180 rpm or 30°C with 150 rpm;

[0100] (3) Dilute the initial concentration of the small molecule drug CHIR-090 to 10 μM and place it in a 4°C refrigerator for later use;

[0101] (4) Take a sterile 96-well plate and perform drug dilution in a biosafety cabinet. The specific operation is as follows: Add 200 μL of the diluted drug to the first well (A1), add 100 μL of the diluted drug to A12, add 100 μL of MH liquid medium to wells A2 - A12. Then, aspirate 100 μL from well A1 and add it to well A2, mix well and then aspirate from well A2, and perform serial dilution to well A10. After diluting the drug in wells A1 - A10, discard the excess liquid to ensure that each well contains 100 μL of liquid. Well A11 is the control group without drug addition, and well A12 is the control group without bacteria addition;

[0102] (5) Adjust the OD 600 of the bacteria to 1, dilute it 5 times, and add 100 μL of the bacterial solution to each of wells A1 - A11 containing the above-diluted drug;

[0103] (6) Place the 96-well plate in an incubator and incubate for 10 to 12 hours. Take out the 96-well plate and observe the growth of the bacteria;

[0104] (7) Repeat the experiment to clarify the antibacterial effect of the CHIR-090 drug against different Vibrio and Salmonella, etc.

[0105] Table 3 Antibacterial effects of the small molecule drug CHIR-090 against different Vibrio and Salmonella, etc.

[0106]

[0107] Among them, 9 Gram-negative bacteria have been disclosed in the paper: Salmonella pullorum S06004: DOI: 10.3382 / ps / pey482; Salmonella enteritidis Z11: DOI: 10.1016 / j.ijmm.2019.151337; Escherichia coli ATCC25922: DOI: 10.3390 / pathogens10050536; Vibrio anguillarum MVM425: DOI: 10.1016 / j.micres.2018.08.011; Edwardsiella tarda EIB202: DOI: 10.1371 / journal.pone.0007646; Staphylococcus aureus USA300: DOI: 10.1016 / S0140-6736(06)68231-7; Campylobacter jejuni 81-176: DOI: 10.3390 / antibiotics10030246; Vibrio alginolyticus EPGS: DOI: 10.1371 / journal.ppat.1005645; Pseudomonas plecoglossicida XSDHY-P: DOI: 10.1016 / j.micres.2022.127220.

[0108] As shown in Table 3, the small molecule drug CHIR-090 showed good inhibitory effects on some Gram-negative bacteria, such as Salmonella pullorum S06004, Salmonella enteritidis Z11, Escherichia coli ATCC25922, Vibrio alginolyticus EPGS, Vibrio anguillarum MVM425 and Pseudomonas plecoglossicida XSDHY-P, etc., but no antibacterial effect was shown on Staphylococcus aureus USA300 and Campylobacter jejuni 81-176.

Claims

1. Use of a small molecule drug CHIR-090 in the preparation of a pharmaceutical composition or vaccine for treating and / or preventing Gram-negative bacterial infection, characterized in that: The Gram-negative bacteria include Vibrio or Enterobacter.

2. The application according to claim 1, characterized in that: The Vibrio includes Vibrio parahaemolyticus, Vibrio anguillarum or Vibrio alginolyticus; the Enterobacterium includes Edwardsiella tarda, Salmonella pullorum, Salmonella enteritidis or Escherichia coli.

3. The application according to claim 1, characterized in that: The Gram-negative bacteria also include Pseudomonas auriculatida.

4. The use according to claim 1, characterized in that: The concentration of the small molecule drug CHIR-090 is 0.16-2.5 μM.

5. The use according to claim 1, characterized in that: When the Gram-negative bacteria is Vibrio parahaemolyticus, the concentration of the small molecule drug CHIR-090 is 0.1-1 μM.

6. A pharmaceutical composition or vaccine for treating and / or inhibiting Gram-negative bacterial infection, characterized in that: The invention contains the small molecule drug CHIR-090; the Gram-negative bacteria include Vibrio or Enterobacter.

7. The use according to claim 6, characterized in that: The Gram-negative bacteria also include Pseudomonas auriculatida.

8. The pharmaceutical composition or vaccine according to claim 6 or 7, characterized in that: The dosage form of the pharmaceutical composition includes tablets, capsules, granules, aerosols, sprays or injections.

9. Use of a small molecule drug CHIR-090 in the preparation of a pharmaceutical composition or vaccine for treating and / or preventing Vibrio parahaemolyticus infection in aquatic products.

10. Application of a small molecule drug CHIR-090 in improving the survival rate of aquatic products infected with Vibrio parahaemolyticus.

Images