Application of interferon kappa in preparation of anti-enveloped virus medicines

An interferon and envelope technology, applied in the field of biomedicine, can solve the problems of public health safety hazards, inability to determine IFN-κ enveloped virus, inconsistent replication mechanism, etc.

Active Publication Date: 2017-11-24
SHANDONG RUIYING PIONEER PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, since ECMV and HPV are non-enveloped viruses, their replication mechanism is not consistent with that of enveloped viruses. Non-enveloped viruses generally enter cells through receptor-mediated endocytosis, while enveloped viruses enter cells by binding to cell membranes or endosomal membranes. Fusion, the introduction of viral capsid and nucleic acid into cells for replication, therefore, it cannot be concluded whether IFN-κ can also inhibit the replication of enveloped viruses
[0003] Influenza virus in enveloped virus and Zika virus (ZIKV) caused by influenza and Zika outbreaks have caused major public health safety hazards and also imposed a heavy burden on social and economic development, and there is still a lack of effective vaccines to prevent infection
It has been reported in the literature that influenza virus can block the common IFN-I signaling pathway by binding to IFN upstream and downstream regulators, thereby escaping the antiviral effect of common IFN-I, while Zika virus can block STAT1 and STAT2 phosphorylation To escape the antiviral effect of common IFN-I, other antiviral small molecule drugs can effectively improve the prognosis of patients, but there are virus resistance mutations under the pressure of drug selection, and some patients with initial infection caused by drug resistance virus transmission Risks of Difficulty Medication Selection

Method used

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  • Application of interferon kappa in preparation of anti-enveloped virus medicines
  • Application of interferon kappa in preparation of anti-enveloped virus medicines
  • Application of interferon kappa in preparation of anti-enveloped virus medicines

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0014] Example 1: Construction of pSV1.0-IFN-κ overexpression plasmid

[0015] The present invention has cloned IFN-κ from the human genome, the nucleotide sequence of its coding gene is shown in SEQ ID No.1, the full-length amino acid sequence is shown in SEQ ID No.2, and the IFN-κ described in the present invention Belonging to the type I interferon family, it has only 30% homology with IFN-α and IFN-β.

[0016] In order to study the function of IFN-κ, we constructed the eukaryotic expression vector of IFN-κ, and eukaryotically expressed the mature secreted protein in cell lines in vitro, and then detected the effect of IFN-κ protein on the replication of influenza virus and Zika virus influences. First, we used the eukaryotic expression vector pSV1.0 to construct the IFN-κ eukaryotic expression plasmid. The construction method was as follows: the cDNA generated by reverse transcription of RNA extracted from A549 cells was used as a template, and the corresponding primers w...

Embodiment 2

[0020] Example 2: IFN-κ inhibits the replication of influenza viruses H7N9, PR8 and H9N2

[0021] The lung epithelial cell line A549 is derived from human non-small cell lung cancer epithelial cells, which is the main cell model for studying influenza virus infection. In order to verify the effect of IFN-κ on different subtypes of influenza virus infection, this example uses 12-well plates The pSV1.0-IFN-κ overexpression plasmid and the control plasmid pSV1.0-GFP were transfected in the A549 cell line. After 24 hours, 100 μL of three influenza virus PR8 ( figure 2 a), H9N2 ( figure 2 b), and H7N9 ( figure 2 c), the average number of virus-infected particles per cell (MOI) is 1. in CO 2 After continuing to incubate in the incubator for 2 hours, discard the virus solution, wash it twice with PBS, add DMEM complete medium and continue to culture for 48 hours, then collect the cells, and analyze the expression of IFN-κ and the expression of influenza virus nucleoprotein NP a...

Embodiment 3

[0023] Example 3: IFN-κ inhibits the replication of Zika virus

[0024] Astrocyte U-251 cells are one of the main target cells of Zika virus infection. In order to verify the effect of IFN-κ on Zika virus infection, U-251 cells were used to transfect control plasmids pSV1.0 and pSV1.0-IFN-κ plasmids. After 36 hours of transfection, cells in some wells were harvested to detect IFN in the cells -κ protein expression level ( image 3 a-left). The remaining cells were infected with Zika virus at a multiplicity of infection (MOI = 2), infected at 37 °C for 2 h, washed twice with PBS, and replaced with fresh DMEM medium containing 2% FBS to continue culturing. After 36 hours of Zika virus infection, the cells were harvested, and a part of the cells were used for western blot (WB) analysis of the expression levels of Zika virus non-structural proteins (NS2b, NS3, NS5) in U-251 cells ( image 3 b-left). Another part of cells was analyzed by immunofluorescence staining technique to...

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Abstract

The invention discloses application of an interferon kappa (IFN-kappa) in preparation of anti-enveloped virus medicines, belonging to the field of anti-virus medicines. A nucleotide sequence of a coding gene of the IFN-kappa is as shown in SEQ ID NO:1, and an amino acid sequence is as shown in SEQ ID NO:2; and an enveloped virus comprises but is not limited to flu and ZIKV. The condition that expression of antiviral protein IFITM3 is induced by IFN-kappa is found, so that infection and replication of flu virus and the ZIKV are prohibited. IFN-kappa can effectively prohibit infection and replication of the enveloped-virus in an in-vitro cell model, so as to relieve various diseases caused by massive replication of the flu virus and the ZIKV. The above finding shows that the IFN-kappa can be used for preparing the anti-enveloped virus medicines as well as medicines for treating and/or preventing various diseases caused by the enveloped-virus.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to the application of interferon-kappa (IFN-κ) in the preparation of anti-envelope virus drugs. Background technique [0002] Many members of the type I interferon (IFN-I) family, as antiviral drug candidates, have completed clinical drug trials, such as recombinant IFN-α2 being applied to anti-hepatitis B virus (HBV) and hepatitis C virus ( HCV) infection treatment drug, IFN-β is used in the treatment of multiple sclerosis (MS). Another relatively new member of the type I interferon family, IFN-κ reported in 2001 can activate the expression of antiviral factors by interacting with IFN receptor (IFNR) 1 / 2. The mechanism of action of IFN-κ may be that after IFN-κ binds to the receptor, it stimulates the phosphorylation of tyrosine kinase 2 (Tyk2) and Janus kinase 1 (Jak1), which further leads to the activation of signal transducer and activator of transcription 1 / 2 (...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K38/21A61K45/06A61P31/16A61P31/14
CPCA61K38/21A61K45/06Y02A50/30
Inventor 徐建青张晓燕傅卫辉陈健何涌泉
Owner SHANDONG RUIYING PIONEER PHARMA
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