Monoclonal antibody of human anti-Ebola virus envelope glycoprotein, and application thereof

An Ebola virus, monoclonal antibody technology, applied in the field of microbiology and immunology, can solve the problem of insufficient stimulation of Ebola GP membrane fusion process, and achieve the effect of excellent antigen binding activity and excellent neutralizing activity

Active Publication Date: 2017-08-11
INST OF BIOENG ACAD OF MILITARY MEDICAL SCI OF THE CHINESE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, simple furin cleavage is still insufficient to trigger membrane fusion of Ebola GP
[0005] After the Ebola virus enters the body, it binds to receptors on the surface of the cell membrane, but the adhesion factors on the surface of the host cell membrane are not the receptors for the membrane fusion of the Ebola virus into the host cell

Method used

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  • Monoclonal antibody of human anti-Ebola virus envelope glycoprotein, and application thereof
  • Monoclonal antibody of human anti-Ebola virus envelope glycoprotein, and application thereof
  • Monoclonal antibody of human anti-Ebola virus envelope glycoprotein, and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Example 1 Screening and preparation of human monoclonal antibodies against Ebola virus envelope glycoprotein

[0030] 1. Blood Sample Collection

[0031] After obtaining informed consent, 5 mL of blood samples were collected 14 days after the first immunization of recombinant Ebola vaccine clinical trial subjects for subsequent experiments.

[0032] 2. Sorting Single Cells by Flow Cytometry

[0033]The collected blood samples were separated from PBMC by Ficoll density gradient centrifugation, and the process was as follows:

[0034] 1) Take fresh anticoagulated whole blood and anticoagulate with EDTA. Whole blood was diluted with an equal volume of PBS or 0.09% NaCl.

[0035] 2) Add a certain volume of separation liquid into the centrifuge tube, spread the diluted blood sample above the liquid surface of the separation liquid, and keep the interface between the two liquid surfaces clear. The volume of separation medium, anticoagulated undiluted whole blood, and PBS ...

Embodiment 2

[0120] Example 2: Antigen epitope analysis of neutralizing monoclonal antibody 1B3 binding to GP

[0121] 1. Analysis of the binding effect between 1B3 and GP and its truncated antigen

[0122] Binding of mAbs to truncated antigens was detected by ELISA. Construct the expression plasmids of three GP truncated antigens GP1 (33-500aa), GPdmucin (33-310aa, 463-632aa), sGP (33-294aa), express the above-mentioned three truncated antigens with Expi293 mammalian cell system, and use the package The lectin on the ELISA plate captured the full-length and truncated antigens of GP respectively, and identified the binding activity of 17 specific antibodies to GP and its truncated antigens. Zmapp optimized strain MIL77-1 / 2 / 3 and anthrax-specific monoclonal antibody 8A7 were used as positive and negative controls, respectively.

[0123] Result: if Figure 5 , MIL77-3 can bind to all four forms of GP, and MIL77-1 / 2 can only bind to the full length of GP and GPdmucin, which is consistent w...

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Abstract

The invention discloses a monoclonal antibody of a human anti-Ebola virus envelope glycoprotein GP. The antibody has a unique CDR region and has a unique action site. Computer simulation analysis and biological experiments confirm that the antibody acts on the NPC1 receptor binding region of the GP, and also acts on the glycan cap region of the GP, so the antibody plays a role in the anti-Ebola virus infection through the unique neutralizing mechanism formed by the combined action of the antibody on the two structural domains of the GP. The antibody has high antigen binding activity and neutralizing activity, can protect 50% of virus attacked cells just at dose of 0.3 [mu]g / ml, and has a protection rate reaching 100% at an antibody concentration of 100 [mu]g / ml, so the antibody provided by the invention has a wide application prospect in the preparation of Ebola virus disease treatment medicines.

Description

technical field [0001] The invention discloses a monoclonal antibody against the envelope glycoprotein of Ebola virus and its application, belonging to the field of microbiology and immunology. Background technique [0002] Ebola virus (EBOV) can cause acute severe hemorrhagic fever in humans and non-human primates. It is one of the viruses with the highest fatality rate found so far, with a fatality rate as high as 90%. It can be transmitted directly through contact , highly contagious and lethal. The Ebola virus is listed as a Class A biological agent / biological agent (Biological agents) by the US NIH (National Institutes of Health) and CDC (Centers for Disease Control), and it is classified as a major threat to national security and public health. One of the most dangerous pathogens, there are currently no approved drugs to prevent or treat Ebola. After the Ebola virus was discovered in 1967, it experienced 12 large-scale transmissions. In 2014, the largest and most di...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/10C12N15/13C12N5/10A61K39/42A61P31/14
CPCA61K2039/505C07K16/10C07K2317/21C07K2317/565
Inventor 陈薇于长明迟象阳范鹏飞侯利华李建民徐俊杰吴诗坡刘威岑宋小红陈旖张晓鹏房婷付玲
Owner INST OF BIOENG ACAD OF MILITARY MEDICAL SCI OF THE CHINESE
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