A chiral metal supramolecular compound, a pharmaceutical composition and its application in combating monkeypox virus.

Abstract

This invention relates to a chiral metal supramolecular compound, a pharmaceutical composition, and their application in treating monkeypox virus, belonging to the field of medicinal chemistry. To address the technical problems of neurotoxic side effects and poor drug-likeness in existing drugs used to treat or prevent poxviruses, this invention provides a chiral metal supramolecular compound and a pharmaceutical composition containing the above compound, wherein the chiral metal supramolecular compound is [Fe2L3]. 4+ The Λ enantiomer, abbreviated as MH3-Λ, has chiral selectivity and is more selective for monkeypox virus RNA than MH3-Δ. On the one hand, it can enhance the stability of A5L mRNA and the expression of the 39kDa core protein of monkeypox virus. On the other hand, it can also enhance the immune response induced by monkeypox virus, reduce the production of monkeypox virus, and prevent the further spread of monkeypox virus to the surrounding environment. It can be used to prepare anti-monkeypox drugs.

Description

Technical Field

[0001] This invention belongs to the field of medicinal chemistry technology, and relates to a chiral metal supramolecular compound, a pharmaceutical composition, and its application in the treatment of monkeypox virus. Background Technology

[0002] Monkeypox is a zoonotic disease caused by monkeypox virus (MPXV), a species of orthopoxvirus in the family Poxviridae. It is primarily transmitted from person to person through direct contact with infectious skin, oral cavity, or genital areas. It is unclear whether the virus can be transmitted in animal populations. Humans can also contract monkeypox through contaminated objects. Monkeypox virus is an enveloped linear double-stranded DNA virus with a genome size of approximately 190 kbp. The incubation period for monkeypox is typically 5-12 days. Clinical features include fever, body aches, rash, and swollen lymph nodes, and complications such as pneumonia, encephalitis, keratitis, and secondary bacterial infections may occur. Lesions begin in the oropharynx and can progress to systemic infection.

[0003] Currently, only reports suggest that tecovirimat (ST-246), a drug previously effective against smallpox, may be a potential treatment for monkeypox, but this has not yet been confirmed in clinical trials. Although the JYNNEOS vaccine received Emergency Use Authorization from the U.S. Food and Drug Administration in 2022, becoming the only monkeypox vaccine approved in Europe and the United States, no drugs have yet been approved clinically for treating monkeypox.

[0004] In view of the fact that existing drugs used to treat or prevent poxviruses have neurotoxic side effects and poor drug-likeness during use, those skilled in the art desire to develop a method that has high drug safety, good drug-likeness, and can effectively treat or prevent diseases caused by monkeypox virus infection. Summary of the Invention

[0005] To address the technical problems of neurotoxic side effects and poor drug-like properties in existing drugs used to treat or prevent poxviruses, this invention provides a chiral metal supramolecular compound, a pharmaceutical composition, and its application in the treatment of monkeypoxvirus.

[0006] One objective of this invention is to provide the application of a chiral metal supramolecular compound in the preparation of an anti-monkeypox drug, wherein the chiral metal supramolecular compound is [Fe2L3]. 4+ The Λ enantiomer.

[0007] In a preferred embodiment of the present invention, the chiral metal supramolecular compound is composed of 2 Fe atoms. 2+ It combines with three ligands through coordinate bonds to form a six-coordinated right-handed binucleated metallic triple-helix cylindrical structure.

[0008] In a preferred embodiment of the present invention, the structural formula of the ligand is:

[0009]

[0010] In a preferred embodiment of the present invention, the chiral metal supramolecular compound binds to the mRNA G4 structure of the A5L gene, which encodes a 39kDa core protein of monkeypox virus.

[0011] In a preferred embodiment of the present invention, the mRNA nucleotide sequence of the A5L gene is shown in SEQ ID No. 1.

[0012] In a preferred embodiment of the present invention, the effective safe concentration for addition of the chiral metal supramolecular compound in Vero cells is 10.27 μM.

[0013] A second objective of this invention is to provide a pharmaceutical composition comprising a chiral metal supramolecular compound and a pharmaceutically acceptable carrier, wherein the chiral metal supramolecular compound is [Fe2L3]. 4+ The Λ enantiomer.

[0014] A third objective of this invention is to provide the use of the above-mentioned pharmaceutical composition in the preparation of anti-monkeypox drugs.

[0015] The beneficial effects of this invention are as follows: This invention provides a chiral metal supramolecular compound and a pharmaceutical composition containing the above compound, wherein the chiral metal supramolecular compound is [Fe2L3]. 4+ The Λ enantiomer (MH3-Λ), which is composed of two Fe... 2+ It combines with three ligands through coordinate bonds to form a six-coordinated right-handed binucleate metallic triple-helix cylindrical structure.

[0016] Compared with existing technologies, the MH3-Λ provided by this invention exhibits chiral selectivity and strong selectivity for monkeypox virus RNA. Furthermore, the MH3-Λ provided by this invention can enhance the stability of A5L mRNA, with a binding constant of 1.22 × 10⁻⁴ to the MG G4 structure. 6 This indicates a strong interaction between MG and MH3-Λ. Furthermore, the MH3-Λ provided by this invention can enhance the expression of the 39kDa core protein of monkeypox virus, and also enhance the immune response induced by monkeypox virus, reduce monkeypox virus production, and prevent further spread of monkeypox virus. Compared with other antiviral drugs, the MH3-Λ provided by this invention has higher drug safety, better drug-likeness, and better monkeypox virus selectivity, and can be used to prepare drugs for treating or preventing diseases caused by monkeypox virus infection. Attached Figure Description

[0017] Figure 1 The chemical structural formula and characterization of the chiral metal supramolecular compound in Example 1 are shown in Figure A, which is the structural diagram of the MH3 ligand; Figure B is the structural diagram of MH3-Λ and MH3-Δ; and Figure C is the CD spectrum of the Λ and Δ enantiomers of MH3. The horizontal axis Wavelength represents the wavelength.

[0018] Figure 2 The following diagram shows the results of bioinformatics prediction of the monkeypox virus genome G-quadruplex in Example 2. A is a flowchart of the bioinformatics analysis of the potential G4 structure of monkeypox virus; B is a preliminary analysis result of GC% and QGRS G4 score; the horizontal axis represents the monkeypox virus genome; C is a graph of G4H score evaluation by PGQs; D is a graph of conservation analysis of the 39kDa virion core protein G1 in orthopoxvirus; E is a graph of conservation analysis of the 39kDa virion core protein G3 in orthopoxvirus; the vertical axis represents the frequency of occurrence of the corresponding bases.

[0019] Figure 3 The images shown are characteristic diagrams of MG G4 structure formation in vitro and in cells in Example 2. A is a migration rate measurement diagram, B is an NMM fluorescence result diagram, and C is... 1 H NMR spectrum, D is CD spectrum, the vertical axis Ellipicity is the ellipticity, E is the intracellular F-MG and BG4 antibody fluorescence colocalization map, the vertical axis FAM / BG4 foci per cell is the number of times FAM and BG4 colocalized fluorescent spots appear in each cell;

[0020] Figure 4 Figure A shows the stability results of MH3-Λ enhanced MPXV G4 in Example 3. Figure B is the fluorescence dissolution curve detection graph. The vertical axis is Normalized fluorescence, which is the normalized correlation fluorescence value. The horizontal axis is Temperature. Figure B is the UV titration detection graph.

[0021] Figure 5 Here is a schematic diagram of the EGFP reporter gene plasmid in Example 3; A is a schematic diagram of the MG-WT-EGFP reporter gene plasmid, and B is a schematic diagram of the MG-Mut-EGFP reporter gene plasmid.

[0022] Figure 6The diagram shows the effect of MH3 on the expression of the MG-WT-EGFP reporter gene in Example 3. A is the confocal fluorescence assay, B is the Image-J quantitative fluorescence assay, the vertical axis represents relative fluorescence intensity, and C is the Western blotting assay.

[0023] Figure 7 The diagram shows the effect of MH3 on the expression of the MG-Mut-EGFP reporter gene in Example 3. A is the confocal fluorescence assay, and B is the Image-J quantitative fluorescence assay.

[0024] Figure 8 This is a schematic diagram of the A5L-EGFP reporter gene plasmid in Example 3;

[0025] Figure 9 The diagram shows the effect of MH3-Λ on the expression of the A5L-EGFP reporter gene in Example 3. A is the confocal fluorescence assay, B is the Image-J quantitative PCR assay, and C is the Western blotting assay.

[0026] Figure 10 This is a graph showing the results of detecting the effect of MH3-Λ on the stability of EGFP mRNA in A5L-EGFP cells in Example 3;

[0027] Figure 11 The graph shows the inhibition curve of MH3-Λ on Vero cell activity in Example 3. The vertical axis, Inhibition, represents the inhibition rate of the drug on cell activity, and the horizontal axis, Concentration, represents the drug concentration.

[0028] Figure 12 The graph shows the inhibitory effect of MH3-Λ on monkeypox virus in Example 3. A shows representative viral plaques after treatment with different concentrations of MH3-Λ (24-well plate). B shows the inhibition curve of MH3-Λ on monkeypox virus after quantitative analysis of plaque images by Image-J. The vertical axis Plaque Inhibitor represents the inhibition rate of the drug on viral plaques. C shows the anti-monkeypox virus effect of MH3-Λ evaluated by qPCR. The vertical axis qPCR Inhibition represents the virus inhibition rate detected by qPCR.

[0029] Figure 13 This is a graph showing the expression levels of inflammatory factors in MPXV-infected Vero cells by MH3-Λ in Example 3.

[0030] Figure 14 This is a schematic diagram illustrating how MH3-Λ regulates the antiviral activity of monkeypox virus core protein via G4 in Example 3. Detailed Implementation

[0031] Those skilled in the art can refer to the content of this document and appropriately improve the process parameters to achieve the desired results. It should be particularly noted that all similar substitutions and modifications are obvious to those skilled in the art and are considered to be included in this invention. The methods and applications of this invention have been described through preferred embodiments, and those skilled in the art can obviously make modifications or appropriate alterations and combinations to the methods and applications described herein without departing from the content and scope of this invention to implement and apply the technology of this invention.

[0032] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to specific embodiments and accompanying drawings. Unless otherwise specified, the experimental methods used in the following embodiments are conventional methods, and the materials, reagents, methods, and instruments used are all conventional materials, reagents, methods, and instruments in the art, and can be obtained commercially by those skilled in the art.

[0033] The chiral metal supramolecular compound [Fe2L3] is used in the following embodiments. 4+ The Λ enantiomer is abbreviated as MH3-Λ, and the chiral metal supramolecular compound [Fe2L3] is... 4+ The Δ enantiomer is abbreviated as MH3-Δ.

[0034] The experimental methods involved in the following embodiments are as follows:

[0035] (1) RNA extraction and cDNA transformation: Total RNA was extracted from cultured cells using a total RNA extraction kit (CWBIO, Beijing, China), and then transformed using PrimeScript. TM RT Master Mix (TaKaRa, Japan) converted it into cDNA.

[0036] (2) Quantitative RT-PCR detection: Quantitative RT-PCR detection was performed using SYBR Mix (Beyotime, China) on an ABI 7500 (Invitrogen, USA), and the relative expression level of RNA was calculated using the comparative Ct method.

[0037] The reaction system consisted of 2 μL of cDNA, 5 μL of 2×SYBR Green qPCR Mix, 0.5 μL each of 10 μM forward and reverse primers, and 2 μL of RNase-free ddH2O; the reaction procedure is shown in Table 1.

[0038] Table 1

[0039]

[0040] (3) Immunoblot analysis: For protein analysis of whole-cell lysates, cells were lysed with RIPA (CWBIO, Beijing, China) buffer, and the lysate was generated on ice for 20 min. The lysate was then centrifuged at 13,000 rpm for 15 min at 4°C. The protein-containing supernatant was transferred to fresh microcentrifuge tubes and stored at -20°C (short-term storage) for further use. Protein quantification was performed using the BCA protein assay (Beyotime, China). Total protein was electrophoresed on an SDS-polyacrylamide gel. The proteins were transferred to a polyvinylidene fluoride (PVDF) membrane and incubated overnight with β-actin primary antibody (Bioss, China) on a shaker at 4°C. The membrane was then washed three times (8 min each time) with TBST (Invitrogen, USA). The blot was then mixed with horseradish peroxidase-labeled goat anti-rabbit IgG. H&L (Bioss, China) was incubated on a plate shaker at RT for 60 min, and the membrane was washed with TBST (3 × 10 min). The bands of the immunoreaction were detected using a chemiluminescent substrate kit (Beyotime, China) and a Western blotting detection system (BioRad, USA), with β-actin used as a loading control for Western blotting detection.

[0041] Example 1: Chiral metal supramolecular compound [Fe2L3] 4+ Preparation of enantiomers

[0042] Step 1: Under a nitrogen atmosphere, (R)-2-phenylglycine (0.67 g, 6.0 mmol, 2.2 equivalents) was dissolved in 50 ml of anhydrous tetrahydrofuran, and 0.6 ml (0.67 g, 3.0 mmol, 1.3 equivalents) of 15-crown ether-5 was injected. Then, the above solution was added dropwise to pure sodium hydride (0.4 g, 10.0 mmol, 4.4 equivalents) to obtain a bubbly mixture. The bubbly mixture was placed under static vacuum and stirred at room temperature for 1 h. Then, 2,8-bis(bromomethyl)dibenzofuran (0.8 g, 2.3 mmol, 1 equivalent) was added dropwise to anhydrous tetrahydrofuran (40 ml of anhydrous tetrahydrofuran). l) The mixture was stirred in the solution under static vacuum to obtain a mixture. The mixture was first treated at room temperature for 1 h, and then at 65 °C for 5 h. The resulting dark green-gray reaction mixture was cooled and then quenched with a 2:1 saturated KCl aqueous solution / water (60 ml). The crude product was extracted with diethyl ether (3 × 100 ml), dried with sodium sulfate, filtered through diatomaceous earth, and the solvent was removed under low pressure, leaving a yellow oily substance as the crude product. The crude product was purified by distillation with a Krijl distillation head (150 °C, 45 min) to remove unreacted excess phenylglycine, and finally optically pure diamine was obtained.

[0043] Step 2: Dissolve 7.5 mg / mL optically pure diamine (3.0 equivalents) and 15 mg / mL 2-pyridinecarboxaldehyde (6.0 equivalents) obtained in Step 1 in methanol (50 mL). Stir at room temperature for 2 h to obtain a yellow solution containing the ligand. Then add 5 mg / mL anhydrous ferric chloride (II) (2.0 equivalents) to the above solution. Stop adding anhydrous ferric chloride (II) (2.0 equivalents) when the color immediately turns dark purple. Then reflux the above solution at 80 °C for 48 h. Filter through grooved filter paper and remove the solvent under low pressure to obtain a product that is a dark purple solid. Dry the product under vacuum at 50 °C overnight to obtain the chiral metal supramolecular compound [Fe2L3]. 4+ The Λ enantiomer, abbreviated as MH3-Λ; and the chiral metal supramolecular compound [Fe2L3]. 4+ The Δ enantiomer is abbreviated as MH3-Δ.

[0044] The chiral metal supramolecular compound [Fe2L3] prepared in this embodiment 4+ The structure of the ligand described in the text is as follows: Figure 1 As shown in Figure A, the enantiomers Λ and Δ are respectively; the metal helical compound structures of their enantiomers are as follows: Figure 1 As shown in B, the CD spectrum of its enantiomer is as follows: Figure 1 As shown in C.

[0045] Example 2: Determination of the G4 structure in the MPXV genome

[0046] This embodiment uses the National Center for Biotechnology Information (NCBI) to locate the genome sequence of monkeypox virus and uses the QGRS-mapper website and G4Hunter software to analyze potential G4 structures in the monkeypox genome. The analysis process is as follows: Figure 2 As shown in Figure A; preliminary analysis of potential G4 sequences was performed using GC% and QGRS G4 scores, and the results are as follows. Figure 2 As shown in B, two potential G4-forming sequences were identified. Then, sequence conservation in different monkeypox virus strains was analyzed using the NCBI database, and G4-forming ability was assessed using the G4H score of PGQs. The results are as follows: Figure 2 As shown in C, a highly conserved potential G4-forming sequence, 39kDa virion core protein G3 (as shown in SEQ ID No. 2), was identified and named MG. Conservation analysis was performed on the 39kDa virion core proteins G1 and G3 in the orthopoxvirus, and the results are as follows. Figure 2 As shown in D, it indicates that it has a high degree of conservatism.

[0047] In this embodiment, to demonstrate that only G-rich MG sequences can form G4 structures, the G base in the MG sequence was mutated to the U base, obtaining a mutant of MG, which was named MG-mut (as shown in SEQ ID No. 3). This embodiment tested the G4-forming ability of the above-mentioned MG and MG-mut by measuring gel migration rate, and the results are as follows... Figure 3 As shown in Figure A, the MG above has a faster migration rate compared to the corresponding mutant MG-mut, proving that MG folds into a compact secondary structure.

[0048] N-methyl-medium porphyrin IX (NMM) is a well-known fluorescent G4-specific ligand used to study the in vitro formation of G4. In this example, NMM was used to detect the fluorescence of the aforementioned MG, and the results are as follows: Figure 3 As shown in Figure B, MG can enhance the fluorescence effect of NMM.

[0049] This embodiment utilizes one-dimensional nuclear magnetic resonance (NMR) spectroscopy to analyze the above-mentioned MG and its mutant MG-Mut. 1 HNMR spectroscopy detection, results are as follows Figure 3 As shown in Figure C, the NMR results show that MG has a distinct peak in the characteristic chemical shift region of G4s (10.2-12.5 ppm), further proving that MG can fold into a G4 structure.

[0050] This embodiment uses 10mM Tris and 100mM K. + In a buffer solution with pH = 7.2, CD spectroscopy was performed on the above-mentioned MG and its mutant MG-Mut. The results are as follows: Figure 3 As shown in Figure D, the CD spectrum of MG shows a typical positive moiré elliptic peak at 262 nm and a negative moiré elliptic peak at 240 nm, while the mutant MG-Mut does not, further illustrating that MG can form a parallel G4 structure.

[0051] In this embodiment, an immunofluorescence co-localization assay was performed in 293t cells using BG4, a G4-specific antibody commonly used in the field. The results are as follows: Figure 3 As shown in E, the MG sequence labeled with FAM was found to co-localize with BG4, indicating that MG G4s can be formed in cells.

[0052] In summary, the highly conserved nucleotide sequence 39kDa virion core protein G3 (MG) of the G4 potential formation sequence, as shown in SEQ ID No. 2, discovered in this invention can fold into a stable G4 structure in vitro and intracellularly.

[0053] Example 3: Application of MH3-Λ in the treatment or prevention of diseases caused by monkeypox virus infection

[0054] 1. Verification of interaction forces

[0055] The gene encoding the MG core protein of the monkeypox virus described in Example 2 has a nucleotide sequence of A5L as shown in SEQ ID No. 1. The mRNA of the monkeypox virus encoding gene A5L has a parallel rG4 (RNA G-quadruplex) structure, and its structural strength differs from that of different enantiomers of various compounds. Therefore, this example uses the chiral metal supramolecular compound [Fe2L3] obtained in Example 1. 4+ The Λ enantiomer (MH3-Λ) and the chiral metal supramolecular compound [Fe2L3] 4+ The interaction / binding ability of the Δ enantiomer (MH3-Δ) with MG G4s in A5L mRNA rG4 was verified.

[0056] This embodiment examines the effects of MH3-Λ and MH3-Δ on the thermal stability of the MG G4 structure by plotting fluorescence dissolution curves. The results are as follows: Figure 4 As shown in Figure A, MH3-Λ significantly enhances the thermal stability of the MG G4 structure compared to MH3-Δ. The binding constant between MH3-Λ and the MG G4 structure was determined by UV titration, and the results are as follows... Figure 4 As shown in B, its binding constant is 1.22 × 10⁻⁶. 6 This indicates a strong interaction between MG and MH3-Λ.

[0057] 2. Verification of Controlled Translation

[0058] Given that mRNA G4s are involved in regulating translation-related processes, it is hypothesized that G4 in the A5L mRNA promoter may be involved in regulating the translation process of A5L mRNA. To verify this hypothesis, this embodiment constructs an enhanced green fluorescent protein (EGFP) reporter gene system to investigate the effect of MG on translation described in Example 2.

[0059] Construction of EGFP reporter vectors: Using a reporter vector with pLV-EGFP-N as the backbone gene (Inovogen Tech.Co., Beijing, China), the gene sequences of MG (nucleotide sequence as shown in SEQ ID No. 2), MG-Mut (nucleotide sequence as shown in SEQ ID No. 3), and A5L (nucleotide sequence as shown in SEQ ID No. 1) were cloned and inserted into the pLV-EGFP vector via EcoRI and BamHI restriction sites, respectively, to obtain MG-WT-EGFP, MG-WT-EGFP, and A5L-EGFP reporter vectors (e.g., MG-WT-EGFP, MG-WT-EGFP, and A5L-EGFP). Figure 5 and Figure 8 (As shown).

[0060] In this embodiment, the MG-WT-EGFP and MG-WT-EGFP reporter vectors obtained above were transfected into HEK293T cells, and the effects of MH3-Λ and MH3-Δ on the expression of MG-WT-EGFP protein were analyzed by Western blotting and confocal fluorescence experiments.

[0061] like Figure 6 As shown, both MH3-Λ and MH3-Δ increased the expression of MG-WT-EGFP protein, as indicated. Figure 7 As shown, MH3-Λ and MH3-Δ did not significantly change the expression of MG-Mut-EGFP protein, indicating that the MH3-Λ and MH3-Δ metal helical compounds prepared in Example 1 enhanced the expression of MG protein by targeting the G4 structure. Furthermore, the above results all indicate that MH3-Λ has a higher ability to induce protein expression than MH3-Δ, further demonstrating a strong interaction between MG and MH3-Λ.

[0062] In this embodiment, the A5L-EGFP reporter vector obtained above was transfected into HEK293T cells, and the enhancer and promoter regions of A5L-EGFP mRNA and the reporter plasmid were analyzed. The results showed that no other G4 forming motifs were found. Western blotting analysis and confocal fluorescence experiments showed that, as Figure 9 As shown, both the MH3-Λ and MH3-Δ metal helical compounds prepared in Example 1 can upregulate the expression of the A5L-EGFP fusion protein, indicating that the formation of the MG G4 structure promotes the expression of the monkeypox core protein encoded by the A5L gene.

[0063] To further elucidate the regulatory mechanism by which MG G4 regulates the A5L gene encoding the monkeypox core protein, this embodiment tested the stability of EGFP mRNA in pMG-WT-EGFP transfected cells supplemented with MH3-Λ. The results are as follows: Figure 10As shown, the addition of MH3-Λ significantly enhanced the stability of EGFP mRNA in pMG-WT-EGFP transfected cells.

[0064] The gene encoding the monkeypox virus MG core protein described in Example 2 is the A5L gene, whose nucleotide sequence is shown in SEQ ID No. 1. The mRNA of the A5L gene has a parallel rG4 (RNA G-quadruplex) structure, and its interaction strength with different enantiomers of the compound varies. Based on the above experimental results, it can be demonstrated that the monkeypox virus encoding gene A5L mRNA rG4 interacts with [Fe2L3]. 4+ The Λ enantiomer (MH3-Λ) has a stronger binding affinity, while [Fe2L3] has a stronger affinity. 4+ The binding of the Δ enantiomer (MH3-Δ) is relatively weak. After MH3-Δ binds to A5L mRNA rG4, it effectively enhances the stability of A5L mRNA and the expression of the 39kDa core protein (MG). It can be seen that MH3-Δ can simultaneously enhance the immune response induced by monkeypox virus, reduce the production of monkeypox virus, and prevent the further spread of monkeypox virus.

[0065] Effect Experiment:

[0066] The viral core protein, with a molecular weight of 39 kDa, is encoded by the A5L gene and is a highly conserved immunogenic protein in poxviruses. It plays a crucial role in activating the host immune response and promoting the conversion of intracellular viral particles (IVs) into intracellular mature viral particles (IMVs).

[0067] This embodiment verifies whether MH3-Λ obtained in Example 1 can exert its anti-monkeypox virus effect by specifically targeting the G4 structure inside the virus.

[0068] This embodiment evaluated the cytotoxicity of MH3-Λ in Vero cells using CCK8 assay, and the results are as follows: Figure 11 As shown, the concentration of MH3-Λ that causes 50% cytotoxicity (CC) 50 The concentration was 10.27 μM. Antiviral experiments with different concentrations of MH3-Λ were conducted using monkeypox virus (MPXV). Quantitative analysis was performed using Image-J, and the efficacy of MH3-Λ against monkeypox virus was evaluated by qPCR. The primer sequences for the qPCR evaluation are shown in SEQ ID No. 4, SEQ ID No. 5, and SEQ ID No. 6. The results are as follows: Figure 12 As shown, the addition of 5 μM MH3-Λ to Vero cells infected with MPXV resulted in a dose-dependent decrease in the viral infectivity (PFU) of MPXV and the F3L-specific viral nucleic acid copy number, indicating that MH3-Λ has a potential antiviral effect on MPXV.

[0069] In this embodiment, GAPDH was used as an internal reference gene (the primer sequences of the internal reference gene are shown in SEQ ID No. 7 and SEQ ID No. 8). The relative mRNA expression levels of IL-6, IL-10, IL-α, IL-1β, and IL-12 were detected in Vero cells infected with monkeypox virus after the addition of a safe concentration of 5 μM MH3-Λ. The results are as follows: Figure 13 As shown, the primer sequences for detecting IL-6 expression level are shown in SEQ ID No. 9 and SEQ ID No. 10, the primer sequences for detecting IL-10 expression level are shown in SEQ ID No. 11 and SEQ ID No. 12, the primer sequences for detecting IL-α expression level are shown in SEQ ID No. 13 and SEQ ID No. 14, the primer sequences for detecting IL-1β expression level are shown in SEQ ID No. 15 and SEQ ID No. 16, and the primer sequences for detecting IL-12 expression level are shown in SEQ ID No. 17 and SEQ ID No. 18.

[0070] It is evident that the addition of MH3-Λ enhanced the expression of IL-6, IL-10, IL-α, IL-1β, and IL-12 in patients infected with monkeypox virus, indicating that it enhanced the immune response to MPXV and further strengthened the inhibition of the virus (e.g., Figure 14 (As shown).

[0071] The contents not described in detail in this specification are well-known to those skilled in the art. Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the invention. Anyone skilled in the art can make various modifications and alterations without departing from the spirit and scope of the invention. Therefore, the scope of protection of the present invention should be determined by the claims.

Claims

1. The application of a chiral metal supramolecular compound in the preparation of an anti-monkeypox drug, characterized in that, The chiral metal supramolecular compound mentioned is [Fe2L3]. 4+ The Λ-enantiomer; The chiral metal supramolecular compound is composed of 2 Fe atoms. 2+ It combines with three ligands through coordinate bonds to form a six-coordinated right-handed binucleate metallic triple-helix cylindrical structure; The structural formula of the ligand is: 。 2. The application according to claim 1, characterized in that, The chiral metal supramolecular compound and A5L The gene's mRNA G4 structure binds, A5L The gene encodes the 39 kDa core protein of monkeypox virus.

3. The application according to claim 2, characterized in that, The A5L The mRNA nucleotide sequence of the gene is shown in SEQ ID No.

1.

4. The application according to claim 1, characterized in that, The effective safe concentration for adding the chiral metal supramolecular compound in Vero cells is 10.27 μM.

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