Screening method and application of neuraminidase inhibitor with drug-resistant mutation resistance potential and product
By constructing a three-dimensional structure and compound library screening of neuraminidase with mutation characteristics, the problem of lack of predicting small molecule structures of anti-influenza virus resistant strains in the prior art was solved, and the neural ammonia with anti-resistant mutation potential was screened out. Immunogenic enzyme inhibitors, effective in dealing with the threat of influenza virus drug resistance mutations.
Patent Information
- Application Number
- CN202411183133.7
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-08-27
- Publication Date
- 2024-12-10
AI Technical Summary
The existing technology lacks methods to predict the small molecule structure of potential NAIs influenza virus resistant strains, which leads to serious problems with anti-influenza drug resistance mutations, and it is urgent to develop new neuraminidase inhibitors with the potential to resist influenza virus resistance mutations.
By constructing a three-dimensional neuraminidase structure with mutation characteristics, using molecules in the compound library for docking screening, neuraminidase inhibitors with anti-drug mutation potential are screened. The specific steps include neuraminidase based on influenza virus Mutation analysis of sequences, Alpha Fold 2 modeling, MOE vertex mutation, AutoDock semi-flexible molecular docking screening and molecular docking verification of Schrodinger software.
Efficient screening of NAIs-resistant strains was achieved, and six compounds with new structures with anti-resistant potential were discovered, providing a convenient and efficient drug molecular screening method to effectively respond to the threat of drug-resistant mutations influenza viruses.
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Figure CN119101997A_ABST
Abstract
Description
Technical Field
[0001] The present invention belongs to the technical field of drug screening, and relates to a screening method, application and product of a neuraminidase inhibitor with the potential to resist drug-resistant mutation. Background Art
[0002] Influenza is an acute respiratory infection caused by influenza virus. It is still an unpredictable infectious disease that poses a serious threat to human life and health. Anti-influenza virus drugs are one of the main means of effectively preventing and treating influenza. Neuraminidase (NA), a glycoprotein on the surface of influenza virus, is a very effective anti-influenza virus target. Its representative drugs oseltamivir and zanamivir are the main anti-influenza virus drugs currently in clinical practice. However, with the increase in the duration and frequency of use, the problem of viral resistance mutation has become increasingly serious. Therefore, there is an urgent need to develop new neuraminidase inhibitors, especially innovative drugs with good potential to resist influenza virus resistance mutations.
[0003] Neuraminidase inhibitors (NAIs) are a class of antiviral drugs used to treat influenza A and B viruses. They compete with sialic acid for the active binding sites of neuraminidase by competitive inhibition, thereby inhibiting the hydrolysis of sialic acid by neuraminidase and reducing the spread of mature virus particles to uninfected cells. Oseltamivir and zanamivir are approved for prevention and treatment worldwide, and peramivir has also been approved for the treatment of influenza virus infection in many places. With the high frequency of NAIs used in influenza prevention and treatment and the easy mutation of viruses, there have been many reports on NAIs-resistant strains, and even a large number of strains resistant to oseltamivir and zanamivir have appeared. Most of these resistant strains directly or indirectly hinder NAIs binding by changing the neuraminidase sequence through gene mutations. Although seasonal influenza monitoring data in recent years show that the resistance rate of the four major epidemic strains to NAIs is about 2%, the problem of viral resistance mutation has become increasingly serious with the increase in the time and frequency of use. Therefore, it is clinically necessary to continue to develop new inhibitors, especially NA inhibitors with the potential to resist influenza virus resistance mutations, to cope with the threat of potential pandemics caused by existing NAIs-resistant strains.
[0004] However, the existing technology lacks a method to predict the structure of small molecules that can combat potential NAIs against drug-resistant strains of influenza virus. Summary of the invention
[0005] In view of this, in view of the bottleneck technical problem that the prior art lacks a method for predicting the structure of small molecules that can fight against potential NAIs influenza virus resistant strains, the purpose of the present invention is to provide a screening method, application and product of neuraminidase inhibitors with potential to resist drug-resistant mutations.
[0006] To achieve the above-mentioned object of the invention, on the one hand, the present invention provides a method for screening neuraminidase inhibitors with potential to resist drug-resistant mutations, comprising the following steps:
[0007] S1. Construction of the three-dimensional structure of neuraminidase with mutation characteristics;
[0008] S2, using the three-dimensional structure of neuraminidase constructed in S1 to perform docking screening on the molecules in the compound library to obtain a compound group;
[0009] S3. Perform molecular docking verification on the compound group obtained in step S2 to obtain a neuraminidase inhibitor with potential to resist drug-resistant mutations.
[0010] Preferably, in step S1, the construction is: based on the mutation analysis and mutation pattern study of the influenza virus neuraminidase sequence, constructed through Alpha Fold 2 modeling and MOE vertex mutation.
[0011] More preferably, the construction is: based on mutation analysis and mutation pattern research of 49280 neuraminidase sequences of influenza A virus from NCBI influenza database, constructed by Alpha Fold 2 modeling and MOE vertex mutation.
[0012] Preferably, in step S2, the compound library is a compound library generated by performing skeleton transition from Chembl and Enamine databases using oseltamivir as the skeleton.
[0013] Preferably, in step S2, the docking screening is: using AutoDock to perform a semi-flexible molecular docking screening method to screen out from the compound library compounds having a higher affinity for the three-dimensional structure of the neuraminidase with mutation characteristics described in step S1 than for the three-dimensional structure of the wild-type neuraminidase.
[0014] Preferably, in step S3, the molecular docking verification is specifically performed using Schrodinger software and Hermite software.
[0015] In another aspect, the present invention provides a neuraminidase inhibitor with the potential to resist drug-resistant mutations obtained by the above screening method, which is selected from one or more of the following substances:
[0016]
[0017] In another aspect, the present invention provides the use of the above-mentioned screening method in screening neuraminidase inhibitors with the potential to resist drug-resistant mutations.
[0018] In another aspect, the present invention provides a device for screening neuraminidase inhibitors with potential to resist drug-resistant mutations, which is used to implement the above screening method, comprising:
[0019] Module 1: Construction of the three-dimensional structure of neuraminidase with mutation features;
[0020] Module 2: Use the three-dimensional structure of neuraminidase transmitted from module 1 to perform docking screening on the molecules in the compound library to obtain a compound group;
[0021] Module 3: Molecular docking verification of the compound group transferred from module 2 was performed to obtain neuraminidase inhibitors with potential to resist drug-resistant mutations.
[0022] In another aspect, the present invention provides an electronic device, comprising:
[0023] A memory storing executable instructions;
[0024] A processor runs the executable commands in the memory to implement the above screening method.
[0025] In another aspect, the present invention provides a computer-readable storage medium storing a computer program, wherein the computer program implements the above-mentioned screening method when executed by a processor.
[0026] Compared with the prior art, the present invention has the following beneficial effects:
[0027] The present invention provides a convenient and efficient method for screening drug molecules for possible NAIs resistant strains. Using the existing drug oseltamivir as the skeleton and combining with the analysis of influenza database sources, six compounds with new structures and potential for anti-resistance were screened. BRIEF DESCRIPTION OF THE DRAWINGS
[0028] Figure 1 This is the effect diagram of the MOE site-directed mutation modeling of the N1 group mutation model. A is the H274Y mutation site; B is the E119V mutation site.
[0029] Figure 2 This is the effect diagram of the MOE site-directed mutation modeling of the N2 group mutation model. A is the E119V mutation site; B is the R152K mutation site.
[0030] Figure 3 This is the 2D interaction diagram of 6 potential anti-drug resistance compounds docked with N1-WT molecule (source: Schrodinger).
[0031] Figure 4This is the 2D interaction diagram of 6 potential anti-drug resistance compounds docking with R1-H274Y molecule (source: Schrodinger). DETAILED DESCRIPTION
[0032] The present invention is further described below in the form of specific embodiments. The following non-limiting examples can enable those of ordinary skill in the art to more fully understand the present invention, but do not limit the present invention in any way. The following content is merely an exemplary description of the scope of the present invention, and those skilled in the art can make various changes and modifications to the present invention based on the disclosed content, and they should also fall within the scope of the present invention.
[0033] Example 1
[0034] A method for screening neuraminidase inhibitors with potential to resist drug-resistant mutations.
[0035] A total of 49,280 NA sequences (FASTA format) of all subtypes of human influenza A viruses prevalent around the world were obtained from the influenza virus database of the National Center for Biotechnology Information (NCBI). There are 9 NA subtypes of influenza A viruses (N1 subtype-N9 subtype), which are divided into two groups according to the characteristics of the NA primary structure sequence. The first group (N1 group) includes N1, N4, N5 and N8 subtypes (17,891 sequences), and the second group (N2 group) includes N2, N3, N6, N7 and N9 subtypes (31,389 sequences). The active center of the NA enzyme is located in a deeper pocket in the center of each subunit, and is composed of 8 functional amino acids (R118, D151, R152, R224, E276, R292, R371, Y406) and 11 structural amino acids (E119, R156, W178, S179, D198, I222, E227, H274, E277, N294 and E425), which together maintain the physiological function of the NA enzyme to hydrolyze substrates and promote the lysis and release of viruses from infected cells. The method of the present invention uses the MAFFT sequence alignment program to respectively align two groups of NA sequences, and then uses Python to analyze and count the amino acid mutation frequencies of 19 active sites, and selects NA mutation sites with a mutation rate of ≥0.05% as the key to introducing the NA mutation model. The sites with actual mutation frequency ≥ 0.05% and their mutation frequencies in the N1 group are summarized in Table 1, and the sites with actual mutation frequency ≥ 0.05% and their mutation frequencies in the N2 group are summarized in Table 2.
[0036] Table 1
[0037] E191K D151N D151G D151E D198N I222T H274Y N294S 0.07％ 0.16％ 0.06％ 0.06％ 0.22％ 0.1％ 10.94％ 0.07％
[0038] Table 2
[0039] E119V D151N D151G D151E D198N I222T I222V R224T R292K 0.14％ 0.77％ 0.50％ 0.15％ 0.54％ 0.06％ 0.42％ 0.05％ 0.42％
[0040] A total of 3265 articles were searched through the SciFinder database in the limited time period from 1998 to 2022 using the keywords influenzavirus, neuraminidase, and drug resistance mutation. Then, 950 articles were reviewed in order according to the relevance of the search content, and the mutation frequency of NA resistance mutation sites was summarized. NA mutation sites with a mutation rate ≥ 3% were selected as the key to introduce the NA mutation model. The results of literature reports on NA resistance mutation frequencies ≥ 3% are summarized in Table 3.
[0041] Table 3
[0042] E119A E119G E119V E119D Q136K R152K H274Y R292K N294S 3.17％ 4.13％ 9.52％ 3.49％ 3.49％ 4.13％ 26.67％ 8.89％ 6.03％
[0043] According to the big data mutation analysis results of 19 active sites and the results of mutation resistance analysis reported in the literature, the basis for modeling and introducing mutation sites was formed by dividing the echelons according to the mutation rate, and the mutation sites were introduced by weight for homology modeling. The N1 group (PDB: 3B7E) and the N2 group (PDB: 7U4G) were respectively introduced with their own templates using MOE to introduce mutations, and the corresponding NA sequences of the R1-R7 real drug-resistant strains were downloaded from NCBI and homology modeled using AlphaFold2. Finally, 23 N1 models, 19 N2 models and 7 real drug-resistant NA models were constructed. Specifically, the establishment of the mutation pattern of the N1 group is shown in Table 4, the construction of the mutation model of the N1 group is shown in Table 5, the establishment of the mutation pattern of the N2 group is shown in Table 6, the construction of the mutation pattern of the N2 group is shown in Table 7, and the construction of the mutation model of the real drug-resistant group is shown in Table 8. In Tables 4-8, "\" represents no content here.
[0044] Table 4
[0045] First Tier H274Y Second Tier E119A / D / G / V, R152K, R292K, N294S Third Tier D151G / E, D198N, I222T Fourth Echelon I222V / M / K / R, E276D, R371K Fifth Echelon D151N、R224T Sixth Echelon Q136K、Y155H
[0046] Table 5
[0047]
[0048]
[0049] Table 6
[0050] First Tier E119A / D / G / V, R152K, R292K, N294S Second Tier D151G / E, D198N, I222T / V Third Tier I222M / K / R, E276D, R371K Fourth Echelon D151N、R224T Fifth Echelon Q136K
[0051] Table 7
[0052]
[0053]
[0054] Table 8
[0055]
[0056] The specific results of MOE site-directed mutation modeling in the N1 group mutation model are as follows: Figure 1 As shown. Among them, Figure 1 A in the figure shows that the mutation site at position H274Y mutates from HID (protonation state of histidine HIS) to TYR. Figure 1 B in the figure shows that the mutation site at position E119V mutates from GLU to VAL.
[0057] The specific results of MOE site-directed mutation modeling in the N2 group mutation model are as follows: Figure 2 As shown. Among them, Figure 2 A in the figure shows that the mutation site at position E119V changes from GLU to VAL. Figure 2 B in the figure shows that the mutation site at position R152K changes from ARG to LYS.
[0058] By analyzing the big data of influenza virus neuraminidase and the literature on drug-resistant mutations, the mutation pattern was determined and a NA 3D model with different mutation sites was constructed. A total of 231,183 compounds generated by oseltamivir skeleton transitions were screened from the Chembl and Enamine databases. A total of 3,528 compounds were obtained with the molecules whose maximum common substructure (MCS) of oseltamivir was greater than 0.6. Semi-flexible molecular docking screening was performed with the help of AutoDock, and the proportion of each compound with a higher affinity for mutants than for wild-type compounds in the N1, N2 and actual drug-resistant groups was calculated. Compounds with an affinity for different NA mutation pattern structures higher than that of the wild-type NA structure by more than 70% were found. After repeated verification with Schrodinger and Hermite molecular docking software, six inhibitor molecules with universal affinity for NA models carrying different mutations were found.
[0059]
[0060] Then, the binding energy performances of the above 6 anti-resistance potential compounds in N1 group, N2 group, and R1-R7 group were calculated respectively, and the results are shown in Tables 9 to 11 respectively.
[0061] Table 9
[0062]
[0063]
[0064] Table 10
[0065]
[0066]
[0067] Table 11
[0068]
[0069]
[0070] Furthermore, the Schrodinger software was used to perform molecular docking of the above six compounds OSV-SF-4, 8, 11, 12, 17, and 23 with N1-WT and R1-HY274, respectively. The results are as follows: Figure 3 , Figure 4 shown.
[0071] Example 2
[0072] A device for screening neuraminidase inhibitors with potential to resist drug-resistant mutations can implement the screening method in Example 1.
[0073] The device includes the following modules:
[0074] Module 1: Construction of the three-dimensional structure of neuraminidase with mutation features;
[0075] Module 2: Use the three-dimensional structure of neuraminidase transmitted from module 1 to perform docking screening on the molecules in the compound library to obtain a compound group;
[0076] Module 3: Molecular docking verification of the compound group transferred from module 2 was performed to obtain neuraminidase inhibitors with potential to resist drug-resistant mutations.
[0077] Example 3
[0078] An electronic device includes: a memory storing executable instructions; and a processor, wherein the processor runs the executable instructions in the memory to implement the screening method shown in Example 1.
[0079] In the electronic device described in this embodiment, the memory is used to store non-temporary computer-readable instructions. Specifically, the memory may include one or more computer program products, which may include various forms of computer-readable storage media, such as volatile memory and / or non-volatile memory. The volatile memory may, for example, include a random access memory (RAM) and / or a cache memory (cache), etc. The non-volatile memory may, for example, include a read-only memory (ROM), a hard disk, a flash memory, etc.
[0080] In the electronic device described in this embodiment, the processor may be a central processing unit (CPU) or other forms of processing units with data processing capabilities and / or instruction execution capabilities, and may control other components in the electronic device to perform desired functions. In one embodiment of the present disclosure, the processor is used to run the computer-readable instructions stored in the memory.
[0081] Example 4
[0082] A computer-readable storage medium stores a computer program, which, when executed by a processor, implements the screening method of Embodiment 1 of the present invention.
[0083] The computer-readable storage medium stores non-transitory computer-readable instructions. When the non-transitory computer-readable instructions are executed by a processor, all or part of the steps of the above-mentioned methods of various embodiments of the present disclosure are executed.
[0084] The computer-readable media include but are not limited to:
[0085] Optical storage media, such as CD-ROM and DVD; magneto-optical storage media, such as MO; magnetic storage media, such as magnetic tapes or mobile hard disks; media with built-in rewritable non-volatile memory, such as memory cards; media with built-in ROM, such as ROM boxes.
[0086] Finally, it should be noted that the above content is only used to illustrate the technical solution of the present invention, rather than to limit the scope of protection of the present invention. Simple modifications or equivalent substitutions of the technical solution of the present invention by ordinary technicians in this field do not deviate from the essence and scope of the technical solution of the present invention.
Claims
1. A method for screening neuraminidase inhibitors with potential to resist drug-resistant mutations, It is characterized in that The following steps are involved: S1. Construction of the three-dimensional structure of neuraminidase with mutation characteristics; S2, using the three-dimensional structure of neuraminidase constructed in S1 to perform docking screening on the molecules in the compound library to obtain a compound group; S3. Perform molecular docking verification on the compound group obtained in step S2 to obtain a neuraminidase inhibitor with potential to resist drug-resistant mutations.
2. The screening method according to claim 1, It is characterized in that In step S1, the construction is: Based on the mutation analysis and mutation pattern research of influenza virus neuraminidase sequence, it was constructed through Alpha Fold2 modeling and MOE vertex mutation.
3. The screening method according to claim 1, It is characterized in that In step S2, the compound library is a compound library generated by performing skeleton transition from Chembl and Enamine databases using oseltamivir as the skeleton.
4. The screening method according to claim 1, It is characterized in that In step S2, the docking screening is: using AutoDock to perform a semi-flexible molecular docking screening method to screen out from the compound library compounds having a higher affinity for the three-dimensional structure of the neuraminidase with mutation characteristics described in step S1 than for the three-dimensional structure of the wild-type neuraminidase.
5. The screening method according to claim 1, It is characterized in that In step S3, the molecular docking verification is specifically performed using Schrodinger software and Hermite software.
6. A neuraminidase inhibitor with the potential to resist drug-resistant mutations obtained by screening according to any one of claims 1 to 5, It is characterized in that One or more selected from the following substances:
7. Use of the screening method according to any one of claims 1 to 5 in screening neuraminidase inhibitors with the potential to resist drug-resistant mutations.
8. A device for screening neuraminidase inhibitors with potential to resist drug-resistant mutations, used for implementing the screening method according to any one of claims 1 to 5, It is characterized in that include: Module 1: Construction of the three-dimensional structure of neuraminidase with mutation features; Module 2: Use the three-dimensional structure of neuraminidase transmitted from module 1 to perform docking screening on the molecules in the compound library to obtain a compound group; Module 3: Molecular docking verification of the compound group transferred from module 2 was performed to obtain neuraminidase inhibitors with potential to resist drug-resistant mutations.
9. An electronic device, It is characterized in that The electronic device comprises: A memory storing executable instructions; A processor, wherein the processor runs the executable instructions in the memory to implement the screening method according to any one of claims 1 to 5.
10. A computer-readable storage medium, It is characterized in that The computer-readable storage medium stores a computer program, and when the computer program is executed by a processor, the screening method according to any one of claims 1 to 5 is implemented.