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Structure and application of anti-ev71, Japanese encephalitis and influenza virus oligonucleotide targeting csnk2a2

An oligonucleotide and antisense oligonucleotide technology, applied in the field of bioengineering drugs, can solve the problems of no resistance to EV71 virus, Japanese encephalitis virus and influenza virus infection, affecting male germ cell function and anti-leukemia, etc. High specificity, less toxic and side effects

Inactive Publication Date: 2011-12-28
INST OF RADIATION MEDICINE ACAD OF MILITARY MEDICAL SCI OF THE PLA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, it has been reported that CSNK2A2 inhibitors can affect the function of male germ cells and have anti-leukemia effects, but there are no reports on anti-EV71 virus, Japanese encephalitis virus and influenza virus infection

Method used

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  • Structure and application of anti-ev71, Japanese encephalitis and influenza virus oligonucleotide targeting csnk2a2
  • Structure and application of anti-ev71, Japanese encephalitis and influenza virus oligonucleotide targeting csnk2a2
  • Structure and application of anti-ev71, Japanese encephalitis and influenza virus oligonucleotide targeting csnk2a2

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] This example mainly illustrates the design, synthesis and screening of antisense oligonucleotides targeting human CSNK2A2 gene against EV71 virus.

[0039] Materials and Methods

[0040] 1. Design and synthesis of antisense oligonucleotide CSNK2A2-3

[0041] Combined with the human gene expression profile of anti-EV71 (expression profile number GSE16358) and HPRD (human protein interaction network), a core subnetwork containing more than 100 nodes was designed by using the method of subnetwork identification, and then topologically important The node screened out the gene CSNK2A2. All oligonucleotides were synthesized with 8909 type automatic DNA synthesizer to synthesize antisense oligonucleotides modified with full sulfur. After the synthesis was completed, the concentrated ammonia solution was cleaved at 55°C and deprotected for 15 hours, then purified by a Micro Pure II reverse-phase purification column (Oligo Prep OP120, SAVANT), quantified by ultraviolet light,...

Embodiment 2

[0060] This example mainly illustrates the specificity of anti-EV71 virus of CSNK2A2-3 after designing the positive sense of CSNK2A2-3 and random verification.

[0061] 1. Design and synthesis of sense strand and random strand of CSNK2A2-3

[0062] The positive sense strand is directly obtained by base complementation with the antisense oligonucleotide antisense sequence of CSNK2A2-3, and its random strand is designed by using SMS2 (The Sequence Manipulation Suite 2), a design software such as DNA random protein, and Through online blast sequence comparison with GeneBank, the selected target sequences have good specificity and will not interfere with the expression of other normal human genes.

[0063] 2. Specific verification of CSNK2A2-3 anti-EV71 virus

[0064]Referring to Example 1, the RD cells were plated on a 96-well cell culture plate, and 75-80% of them were confluent the next day. The 96-well plate of DMEM medium (GIBCO) containing 10% fetal bovine serum, which was...

Embodiment 3

[0070] This example mainly illustrates that CSNK2A2-3 can dose-dependently inhibit the RNA of EV71 virus

[0071] Materials and methods

[0072] Dose-dependent effect of CSNK2A2-3 on inhibiting EV71 viral RNA

[0073] The RD cells were plated on a 96-well cell culture plate, and 75-80% of them were confluent the next day. The 75-80% confluent RD cells were replaced with maintenance solution containing 2% fetal bovine serum, and CSNK2A2-3 of different concentrations were added. Four concentration gradients of 0.25, 0.5, 1.0, and 2.0 μM were set up for CSNK2A2-3 respectively, and three auxiliary wells were set up for each sequence concentration, and a virus positive control group and a RD cell negative control group were set up. After acting for 60 minutes, add 3.3 μl of EV71 virus dilution solution of 100 TCID50 / 0.1 ml to each well, and incubate at 37° C. for 2 days. After 2 days, the supernatant of the culture medium was collected, and viral RNA was extracted using a viral...

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Abstract

The invention relates to the structure and application of an antisense oligonucleotide drug. Specifically, the structure of antisense oligonucleotides targeting casein kinase 2, alpha propeptide, and anti-enterovirus type 71, Japanese encephalitis virus and influenza virus was found, and these antisense oligonucleotides were found in Use in the preparation of medicines for treating EV71, Japanese encephalitis virus and influenza virus infection and related diseases.

Description

technical field [0001] The invention relates to the field of bioengineering drugs, in particular to a method targeting casein kinase 2, alpha prime peptide (CSNK2A2, casein kinase 2, alpha prime polypeptide) for treating EV71 (Human enterovirus 71) . The sequence, structure and therapeutic drug of antisense oligonucleotide (ASODN, antisense oligodexynucleotide) for infection. Background technique [0002] Viral disease infection is a serious hazard, especially EV71 virus, encephalitis virus, and influenza virus. EV71 is the latest virus discovered in the current enterovirus group. It has strong infectivity and high pathogenicity rate, especially the complications of the nervous system. Other viruses that belong to the enterovirus group include Polioviruses (3 types), Coxsackieviruses (23 types of type A, 6 types of type B), Iraqi virus (Echoviruses; with 31 types) and enteroviruses (Enteroviruses 68-72 types). Enterovirus 71 (English name: Human enterovirus 71). Referre...

Claims

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Application Information

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IPC IPC(8): C12N15/113A61K48/00A61P31/16A61P31/14
Inventor 王升启杨静刘娟伯晓晨张莉李康何丽娜
Owner INST OF RADIATION MEDICINE ACAD OF MILITARY MEDICAL SCI OF THE PLA
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