Use of a small molecule inhibitor in the preparation of a product for combating ebola virus infection

Abstract

The application discloses application of a small molecule inhibitor in preparation of an anti-Ebola virus infection product. The application firstly confirms the anti-Ebola virus capability of TOFA, and the cell toxicity of TOFA is small (CC50 value is 7.717 muM), and TOFA can effectively inhibit replication of Ebola pseudovirus at a low concentration (IC50 value is 1.976 muM), so that a new candidate small molecule compound for developing a novel anti-Ebola virus infection drug is provided, a new technical selection for effectively preventing a severe infectious disease is provided, and the application has a wide application prospect.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, specifically to the application of a small molecule inhibitor in the preparation of products for treating Ebola virus infection. Background Technology

[0002] Ebola virus disease (EVD), also known as Ebola hemorrhagic fever (EHF), is a highly contagious and hemorrhagic disease caused by the Ebola virus (EOV) infecting humans and non-human primates. Ebola virus is an RNA virus, characterized by high infectivity, high mortality, and high pathogenicity. It has a biosafety level of 4, indicating extremely high risk and pathogenicity with a mortality rate as high as 90%. Currently, the U.S. Food and Drug Administration (FDA) has approved two monoclonal antibody (mAbs) drugs for the treatment of EVD: Inmazeb and Ebanga. However, in a PALM (Pamoja Tulinde Maisha) study, more than 30% of patients still died after receiving treatment with these two drugs. Therefore, there is an urgent need to develop new EBOV treatments to improve the treatment efficacy of EVD.

[0003] Compared to mAbs, small molecule compounds offer advantages in ease of production, transportation, and storage, and also have lower production costs. Furthermore, some EVD survivors persistently harbor Ebola virus. The large molecular weight of mAbs limits their penetration into the immune protection zone where EBOV persists, while the smaller molecular weight and greater permeability of small molecule compounds help to completely eliminate the virus, preventing potential transmission and long-term effects. Small molecule compounds capable of combating Ebola virus infection remain to be developed. Summary of the Invention

[0004] To address the problems in the prior art, this invention provides the application of a small molecule inhibitor in the preparation of products for treating Ebola virus infection.

[0005] The first objective of this invention is to provide the use of a small molecule inhibitor in the preparation of products for treating Ebola virus infection.

[0006] A second objective of this invention is to provide the use of a small molecule inhibitor in the preparation of a medicament for the prevention and / or treatment of diseases caused by Ebola virus.

[0007] A third objective of this invention is to provide the use of a small molecule inhibitor in the preparation of products for the prevention and / or treatment of Ebola hemorrhagic fever.

[0008] A fourth objective of this invention is to provide the application of a small molecule inhibitor in the preparation of products that inhibit Ebola virus replication.

[0009] A fifth objective of this invention is to provide the use of a small molecule inhibitor in the preparation of products that inhibit the expression of genes and / or proteins of Ebola virus.

[0010] The sixth object of this invention is to provide an anti-Ebola virus infection preparation.

[0011] The seventh object of the present invention is to provide a pharmaceutical composition.

[0012] To achieve the above objectives, the present invention is implemented through the following solution:

[0013] This invention utilizes the Ebola virus replication subsystem to clarify the anti-Ebola virus effect of acetyl-CoA carboxylase-α (ACCA) allosteric inhibitor (TOFA). TOFA can inhibit the expression of VP40 gene and protein and GP gene and protein, and can inhibit the replication and infection of Ebola pseudoviruses.

[0014] This invention seeks protection for the following:

[0015] The application of a small molecule inhibitor in the preparation of products for treating Ebola virus infection, wherein the structural formula of the small molecule inhibitor is as follows: , namely TOFA.

[0016] The use of a small molecule inhibitor in the preparation of a medicament for the prevention and / or treatment of diseases caused by Ebola virus, wherein the small molecule inhibitor has the following structural formula: .

[0017] The use of a small molecule inhibitor in the preparation of products for the prevention and / or treatment of Ebola hemorrhagic fever, wherein the structural formula of the small molecule inhibitor is as follows: .

[0018] The application of a small molecule inhibitor in the preparation of products that inhibit Ebola virus replication, wherein the structural formula of the small molecule inhibitor is as follows: .

[0019] The use of a small molecule inhibitor in the preparation of a product that inhibits the expression of genes and / or proteins of Ebola virus, wherein the structural formula of the small molecule inhibitor is as follows: .

[0020] Preferably, the genes of the Ebola virus include VP40 Genes and / or GP Gene.

[0021] Preferably, the proteins of the Ebola virus include VP40 Protein and / or GP protein.

[0022] An anti-Ebola virus infection preparation, comprising a structure as shown in the figure below. The small molecule inhibitor shown.

[0023] Preferably, the formulation is in the form of an injection, tablet, spray, granule, capsule, oral liquid, or patch.

[0024] Preferably, the content of the small molecule inhibitor in the formulation is 0.1 μM to 12.8 μM.

[0025] More preferably, the small molecule inhibitor is present in the formulation at a concentration of 0.1 μM to 2 μM.

[0026] More preferably, the small molecule inhibitor is present in the formulation at a concentration of 0.4 μM.

[0027] A pharmaceutical composition comprising a structure as shown in the figure below. The small molecule inhibitors shown are pharmaceutically acceptable carriers or excipients.

[0028] Preferably, the small molecule inhibitor is present in the pharmaceutical composition at a concentration of 0.1 μM to 12.8 μM.

[0029] More preferably, the small molecule inhibitor is present in the formulation at a concentration of 0.1 μM to 2 μM.

[0030] More preferably, the small molecule inhibitor is present in the pharmaceutical composition at a concentration of 0.4 μM.

[0031] Compared with the prior art, the present invention has the following beneficial effects:

[0032] This invention clarifies for the first time the anti-Ebola virus ability of TOFA, which exhibits low cytotoxicity (CC50 value of 7.717 μM). TOFA can effectively inhibit the replication of Ebola pseudovirus at low concentrations (IC50 value of 1.976 μM), providing a new candidate small molecule compound for the development of novel drugs against Ebola virus infection and offering a new technical option for the effective prevention of highly infectious diseases, with broad application prospects. Attached Figure Description

[0033] Figure 1 This is a graph showing the results of the cytotoxicity experiment of TOFA in Example 1.

[0034] Figure 2 This is an example of the effect of different concentrations of TOFA on Ebola virus replication in Example 2.

[0035] Figure 3This is the effect of TOFA on the expression levels of Ebola virus VP40 and GP proteins in Example 3.

[0036] Figure 4 This is the effect of TOFA on the expression level of the Ebola virus VP40 gene in Example 4.

[0037] Figure 5 This shows the effect of TOFA on the expression level of the Ebola virus GP gene in Example 4. Detailed Implementation

[0038] The present invention will be further described in detail below with reference to the accompanying drawings and specific embodiments. These embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Unless otherwise specified, the experimental methods used in the following embodiments are conventional methods; the materials and reagents used, unless otherwise specified, are commercially available.

[0039] The complete set of EBOV-trVLPs pseudovirus plasmids used in the following examples—p4cis-vRNA-RLuc (encoding proteins: Renilla luciferase, VP40, GP, and VP24), pCAGGS-NP, pCAGGS-VP35, pCAGGS-VP30, pCAGGS-L, pCAGGS-TIM1, pCAGGS-T7, and pCAGGS-firefly luciferase (i.e., pCAGGS-luc2)—was kindly provided by Professor Thomas Hoenen of the National Institute of Allergy and Infectious Diseases, National Institutes of Health. The structural formula of the allosteric inhibitor of acetyl-CoA carboxylase-α (TOFA) is as follows: Purchased from MCE.

[0040] Example 1: Cytotoxicity test of TOFA

[0041] Cytotoxicity was detected using Biosharp's CCK-8 assay kit (catalog number: BS350B). The specific steps are as follows:

[0042] In 96-well plates, 2000 cells were seeded with HEK293T cell suspension (100 μL / well) per well. After culturing for 12 h, TOFA was added at final concentrations of 100 nM, 200 nM, 400 nM, 800 nM, 1.6 μM, 3.2 μM, 6.4 μM, and 12.8 μM, respectively. The corresponding wells were designated as drug-treated wells. Three replicates were set up for each concentration. In addition, negative control wells without TOFA and blank wells without cells and without TOFA were also set up.

[0043] After TOFA treatment for 48 h, 10 μL of CCK-8 solution was added to each well, and the cells were incubated in a cell culture incubator for another 4 h. After the incubation, the absorbance of each well at 450 nm was measured using a microplate reader, and cell viability was calculated using the following formula: Cell viability = [OD(drug-treated wells) - OD(blank wells)] / [OD(negative control wells) - OD(blank wells)] × 100%.

[0044] A nonlinear curve was fitted using GraphPad Prism to represent the logarithm of TOFA concentration versus cell viability. The inhibition rate was negatively correlated with cell viability. Based on the obtained curve, the TOFA concentration at which the inhibition rate was 50% was determined, which is the CC50 value.

[0045] like Figure 1 As shown, the CC50 value of TOFA is 7.717 μM, indicating that the small molecule drug TOFA has low cytotoxicity and high safety when the working concentration is below 7.717 μM.

[0046] Example 2: Effects of different concentrations of TOFA on Ebola virus replication

[0047] 1. Preparation of the first generation of Ebola pseudovirus

[0048] (1) Replicable pseudovirus system (EBOV virus-like particles)

[0049] The Ebola virus replication subsystem used in this invention is a mature system that has been reported in the prior art (DOI: 10.3389 / fcimb.2017.00479) and is recognized. Its working principle is to use reverse genetics technology and eukaryotic expression system to construct multi-life cycle Ebola virus-like particles, realize the simulation of multiple life cycle of Ebola virus adhesion, invasion, transcription, replication, virus particle packaging, virus budding, and continuous passage under BSL-2 conditions, thereby providing strong technical support for basic and applied research on Ebola virus.

[0050] The EBOV genome encodes seven structural proteins, including RNA polymerase (L), nucleocapsid protein (NP), polymerase cofactor (VP35), and transcription activator (VP30), which are involved in viral transcription and replication, as well as envelope glycoprotein (GP), matrix protein (VP40), and second matrix protein (VP24), which constitute the viral capsid structure and are involved in viral membrane binding, assembly, and budding.

[0051] The trVLPs-EBOV pseudovirus system mimics the entire life cycle of the Ebola virus. This system comprises a tetracistronic minigenome (MG), which undergoes initial transcription under the T7 polymerase promoter. The MG contains flanking non-coding regions and a central René luciferase. Renilla luciferase Gene, VP40 Gene, GP Genes and VP24 Gene.

[0052] The plasmid encoding MG (p4cis-vRNA-RLuc) and the plasmids encoding NP, VP35, VP30, L and T7 RNA polymerases (pCAGGS-NP, pCAGGS-VP35, pCAGGS-VP30, pCAGGS-L and pCAGGS-T7) were co-transfected into primary producer cells and packaged to obtain primary trVLPs-EBOV (i.e., P0 generation trVLPs).

[0053] P0 generation trVLPs were then used to infect target cells transfected with plasmids encoding TIM1, NP, VP35, VP30, and L, and packaged to obtain P1 generation trVLPs-EBOV. The replication level of trVLPs-EBOV was evaluated by detecting the activity of Renilla luciferase (Rluc) in the lysates of target cells.

[0054] (2) Preparation of P0 generation trVLPs-EBOV

[0055] To package multi-lifecycle Ebola virus-like particles, eukaryotic expression plasmids pCAGGS-NP, pCAGGS-VP35, pCAGGS-VP30, pCAGGS-L, pCAGGS-TIM1, and pCAGGS-T7, as well as the firefly luciferase Luc2 (pCAGGS-luc2), were prepared using pCAGGS as the backbone. In addition, the viral microgenomic plasmid p4cis-vRNA-RLuc was synthesized for subsequent virus-like particle packaging. The construction and sequencing of the above plasmids involved in this embodiment were jointly completed by the Academy of Military Medical Sciences, the National Institute of Allergy and Infectious Diseases of the National Institutes of Health (NIH), Shanghai Jikai Gene Chemical Technology Co., Ltd., and Jilin Kumei Biotechnology Co., Ltd.

[0056] The specific steps for virus packaging are as follows: revive HEK293T cells, passage them for 3 consecutive times, and then pack them at a cell density of 5×10⁶ cells / year. 5Cells were seeded at 0.5 mL / mL in 6-well plates and cultured at 37°C in a cell culture incubator containing 5% CO2. After 12 h of cell culture, plasmid transfection was initiated. The transfection reagent Lipofectamine™ 3000 was removed from the 4°C freezer and placed at room temperature (25°C) for later use. 100 µL of Opti-MEM was added to each well of the 6-well plate. Add 300 µL of Opti-MEM to a sterile centrifuge tube, add the corresponding plasmid according to the dosage in Table 1, and add 30 µL of P3000 and vortex to mix. Add another 300 µL of Opti-MEM to a sterile centrifuge tube, and add 7.5 µL of Lipofectamine™ 3000 transfection reagent per well, i.e., add 45 µL of Lipofectamine™ 3000 transfection reagent to the above sterile centrifuge tube, vortex gently to mix, and incubate each tube at room temperature for 5 min. Gently vortex the two tubes together and incubate at room temperature (25℃) for 5 min to obtain transfection system 1.

[0057] Table 1. Transfection dose of viral packaging plasmid (6-well plate)

[0058]

[0059] Add 100 µL of transfection system 1 to each well of a 6-well plate, shake the plate back and forth to ensure even distribution of the transfection system, and then place it in a 37°C cell culture incubator containing 5% CO2 for further cell culture. After 24 h, remove the cell culture supernatant, and add 2 mL of DMEM medium containing 5% v / v fetal bovine serum and 1% v / v penicillin antibiotics to each well of the 6-well plate. Place the 6-well plate with the replaced medium in a 37°C cell culture incubator containing 5% CO2 for further culture for 48 h. Collect the cell supernatant after culture and transfer it to a 15 mL centrifuge tube. Centrifuge at 800 g for 5 min at room temperature (25°C) to remove cell debris, thus obtaining P0 generation trVLPs-EBOV. Aliquot and store at -80°C.

[0060] 2. Helper plasmid transfection of target cells

[0061] Resuscitate additional HEK293T cells, passage them for 3 consecutive times, and then culture them in DMEM medium (containing 10% FBS + 1% penicillin and antibiotics) at a cell density of 4 × 10⁶ cells / year. 5 Cells were seeded at a density of 1 / mL in 24-well plates and cultured at 37°C in a cell culture incubator containing 5% CO2. After 12 h of culture, when the cell density reached 80%, plasmid transfection was initiated.

[0062] Take a sterile centrifuge tube and add 500 µL of Opti-MEM according to the dosage in Table 2, then add the corresponding plasmid and 48 µL of P3000 and vortex to mix. Take another sterile centrifuge tube and add 500 µL of Opti-MEM. Add 2 µL of Lipofectamine™ 3000 transfection reagent per well, that is, add 48 µL of Lipofectamine™ 3000 transfection reagent to the above sterile centrifuge tube, gently vortex to mix, and incubate at room temperature (25℃) for 5 min each. Gently vortex the two tubes together and incubate at room temperature (25℃) for 15 min to obtain transfection system 2.

[0063] Table 2. Target cell plasmid transfection dose (24-well plate)

[0064]

[0065] Add 44 µL of transfection system 2 to each well of a 24-well plate, shake the 24-well plate back and forth to disperse the transfection system evenly, and then place it in a cell culture incubator containing 5% CO2 at 37°C to continue cell culture. The resulting cells are the target cells.

[0066] 3. Infection of target cells by P0 generation trVLPs-EBOV and drug treatment

[0067] 24 h after transfection, discard the culture medium for the target cells and seed each well with 300 μL of P0 generation trVLPs-EBOV (1×10⁻⁶). 7 (Copies / mL) Simultaneously, TOFA was added to final concentrations of 100 nM, 200 nM, 400 nM, 800 nM, 1.6 μM, 3.2 μM, 6.4 μM, and 12.8 μM for drug treatment. Each well was designated as the drug-treated well, with three replicates for each concentration. A DMSO-only group was included as a negative control. After culturing for 24 h, the cell supernatant was discarded, and the medium was replaced with DMEM containing 4% v / v fetal bovine serum.

[0068] 4. Luciferase activity detection

[0069] After culturing for another 48 h, the cells were gently washed twice with pre-cooled PBS, and any residual liquid was aspirated. 200 μL of pre-prepared 1× Glo Lysis Buffer was added to each well, and the cells were incubated at room temperature (25°C) for 15 minutes to ensure complete lysis. The cell lysates were then collected.

[0070] Transfer 75 μL of cell lysis buffer to a 96-well opaque white microplate, add an equal volume (75 μL) of Dual-Glo® Luciferase Reagent, gently pipette to mix, and incubate at room temperature (25°C) in the dark for 10 minutes. Use a multi-functional microplate reader (Synergy H1, BioTek) to read the Firefly luciferase signal from each well.

[0071] After completing the Firefly signal detection, add an equal volume (75 μL) of Dual-Glo® Stop & Glo® Reagent to the same well, mix well with a pipette, and incubate at room temperature (25°C) in the dark for 10 min. Read the Renilla luciferase signal from each well.

[0072] The ratio of Firefly luciferase activity to Renilla luciferase activity was used as the standardization result, which is the dual luciferase activity of each well. A nonlinear curve of the logarithm of TOFA concentration versus viral infection rate was fitted using GraphPad Prism8, and the TOFA concentration at a viral infection rate of 50% was determined from the obtained curve, which is the IC50 value.

[0073] like Figure 2 As shown, the higher the TOFA concentration, the lower the viral infection rate. The IC50 value of TOFA is 1.976 μM. Combined with the cytotoxicity experiment in Example 1, when the working concentration of TOFA is 400 nM, the cell viability is close to 100% and the viral infection rate is reduced by nearly 40%, indicating that the small molecule drug TOFA can effectively inhibit the replication of Ebola pseudovirus at a low concentration and has high safety.

[0074] Example 3: Effects of different concentrations of TOFA on the expression levels of Ebola virus VP40 and GP proteins

[0075] 1. Preparation of the first generation of Ebola pseudovirus

[0076] Same as Example 2.

[0077] 2. Helper plasmid transfection of target cells

[0078] Following the method described in Example 2, HEK293T cells were cultured in 12-well plates. When the cell density reached 80%, helper plasmids were transfected. Transfection system 3 was prepared by adding the corresponding plasmids according to the dosages in Table 3.

[0079] Table 3. Target cell plasmid transfection dose (12-well plate)

[0080]

[0081] 3. Infection of target cells by P0 generation trVLPs-EBOV and drug treatment

[0082] 24 h after transfection, the culture medium for the target cells was discarded, and 300 μL of P0 generation trVLPs-EBOV was seeded in each well. Wells treated with TOFA at a final concentration of 400 nM were designated as the experimental group, and wells treated with 0.05% DMSO were designated as the control group. Each group had three replicates. After culturing for another 24 h, the cell supernatant was discarded, and the medium was replaced with DMEM containing 4% v / v fetal bovine serum.

[0083] 5. Western Blot Detection

[0084] After culturing for another 48 h, the culture medium was discarded, the cells were washed twice with pre-cooled PBS, 0.1 mL of NP-40 lysis buffer was added to each well, the cells were lysed on ice for 15 min, centrifuged at 12000 g for 10 min at 4 °C, and the supernatant was collected.

[0085] Mix the supernatant and loading buffer at a 4:1 ratio and heat in a 95°C metal bath for 10 min to completely denature the proteins. Perform SDS-PAGE electrophoresis, adding 20 μL of protein sample to each well, with the protein marker used as a reference. The initial voltage is 80 V, adjusted to 120 V after bromophenol blue enters the separating gel, and electrophoresis continues until the bromophenol blue migrates to the bottom of the gel. For membrane transfer, immerse the NC membrane and filter paper in transfer buffer (containing methanol) and assemble the "sandwich" structure (filter paper-gel-PVDF membrane-filter paper), removing air bubbles. Transfer at a constant voltage of 150 V for 1 h at 4°C. Block with 5% skim milk powder (prepared with TBST) at room temperature (25°C) for 1 h to reduce non-specific binding. Dilute the VP40-specific antibody (Snio Biological, catalog number 40447-MM08), GP-specific antibody (Snio Biological, catalog number 40094-R106), and GAPDH-specific antibody (Proteintech, catalog number 60004-1-IG) and incubate overnight at 4°C. Wash the membrane three times for 10 min each time. Dilute the HRP-labeled secondary antibody (goat anti-rabbit IgG) (ThemoFisher, catalog number 31460) with TBST buffer and incubate at room temperature (25°C) for 1 h. Wash the membrane three times for 10 min each time with TBST buffer. Develop using chemiluminescence, cover with ECL reagent, and incubate at room temperature (25°C) for 1 min. Expose using a gel imaging system and adjust the time to obtain clear bands.

[0086] like Figure 3As shown, compared with the blank group, the expression levels of VP40 and GP proteins in the control group were significantly increased after Ebola pseudovirus infection; while after TOFA treatment, the expression of VP40 and GP proteins in the experimental group was significantly decreased compared with the control group. This indicates that TOFA can significantly inhibit the expression of VP40 and GP proteins and inhibit Ebola virus replication.

[0087] Example 4: TOFA against Ebola virus VP40 Genes and GP Effects of gene expression levels

[0088] 1. Preparation of the first generation of Ebola pseudovirus

[0089] Same as Example 2.

[0090] 2. Helper plasmid transfection of target cells

[0091] Same as Example 2.

[0092] 3. Infection of target cells by P0 generation trVLPs-EBOV and drug treatment

[0093] 24 h after transfection, discard the culture medium for the target cells and seed each well with 300 μL of P0 generation trVLPs-EBOV (1×10⁻⁶). 7 The wells treated with TOFA at a final concentration of 400 nM were used as the experimental group, and the wells treated with 0.05% DMSO were used as the control group. Each concentration was tested in triplicate. After culturing for another 24 h, the medium was changed and the cell supernatant was discarded, and replaced with DMEM medium containing 4% v / v fetal bovine serum.

[0094] 5. q-PCR detection

[0095] After culturing for another 48 h, the culture medium was discarded, and the cells were washed twice with pre-chilled PBS. 1 ml of TRIZOL was added to each well to resuspend the cells, followed by 200 μL of chloroform. The cells were vigorously shaken for 15 s and incubated at room temperature (25°C) for 3 min. The cells were then centrifuged at 12000 g for 15 min at 4°C. After separation, the upper aqueous phase was carefully aspirated into a new tube. An equal volume of pre-chilled isopropanol was added to the aqueous phase, and the mixture was inverted and incubated at room temperature (25°C) for 10 min. The cells were then centrifuged at 12000 g for 10 min at 4°C, and the supernatant was discarded. A white RNA precipitate was visible. 1 ml of 75% pre-chilled ethanol was added to gently resuspend the precipitate, and the cells were centrifuged at 7500 g for 5 min at 4°C. The precipitate was dissolved in 30 μL of DEPC water, and the concentration was measured.

[0096] RNA was reverse transcribed into cDNA. The reverse transcription system was as follows: template RNA, 1 μg; 5× Evo M-MLV RT MasterMix, 4 μL; and RNase-free water was added to a final volume of 20 μL. The reverse transcription program was: 37℃, 15 min; 85℃, 5 sec.

[0097] Using the obtained cDNA as a template, q-PCR was performed with primers (VP40 upstream primer: 5'-AGTTGGACTGGCGGAAGAAC-3'; VP40 downstream primer: 5'-CAGAGTCAATCGGCTGGGTC-3'; GP upstream primer: 5'-CCGGAGACTTTGCCTTCCAT-3'; GP downstream primer: 5'-CTCTCAAGGGGTGTGAGCTG-3'; GAPDH upstream primer: 5'-AATGGGCAGCCGTTAGGAAA-3'; GAPDH downstream primer: 5'-GCCCAATACGACCAAATCAGAG-3').

[0098] The q-PCR reaction system (total volume 25 μL) consisted of: 12.5 μL of 2×SYBR Green Pro Taq HS Premix (ROXplus), 0.5 μL each of upstream and downstream primers, 3 μL of cDNA, and sterile deionized water to a total volume of 25 μL. The q-PCR reaction program was: 95℃, 30 s; 95℃, 15 s, 60℃, 60 s, 40 cycles.

[0099] After the reaction is complete, collect the amplification data, and then use 2... -ΔΔCt The method calculates the relative expression level of the target gene.

[0100] like Figure 4 and Figure 5 As shown, compared with the control group, the mRNA levels of VP40 and GP genes in the experimental group were significantly reduced. This indicates that TOFA inhibits the expression of VP40 and GP genes in EBOV.

[0101] In conclusion, TOFA can significantly inhibit the expression of Ebola virus genes and proteins, suppress viral replication, and has excellent anti-Ebola virus effects.

[0102] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the scope of protection of the present invention. For those skilled in the art, other variations or modifications can be made based on the above description and ideas, and it is neither necessary nor possible to exhaustively describe all implementation methods here. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention should be included within the scope of protection of the claims of the present invention.

Claims

1. The application of a small molecule inhibitor in the preparation of products for treating Ebola virus infection, characterized in that, The structural formula of the small molecule inhibitor is as follows: .

2. The use of a small molecule inhibitor in the preparation of a medicament for the prevention and / or treatment of diseases caused by Ebola virus, characterized in that, The structural formula of the small molecule inhibitor is as follows: .

3. The use of a small molecule inhibitor in the preparation of products for the prevention and / or treatment of Ebola hemorrhagic fever, characterized in that, The structural formula of the small molecule inhibitor is as follows: .

4. The application of a small molecule inhibitor in the preparation of products that inhibit Ebola virus replication, characterized in that, The structural formula of the small molecule inhibitor is as follows: .

5. The application according to any one of claims 1 to 4, characterized in that, The genes of the Ebola virus include the VP40 gene and / or the GP gene.

6. The application according to any one of claims 1 to 4, characterized in that, The proteins of the Ebola virus include VP40 protein and / or GP protein.

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