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Anti-respiratory syncytial virus fully human broad-spectrum neutralizing antibody 4F1 and application thereof

An antibody and antibody drug technology, applied in the field of medicine, can solve the problems of high price and high dosage

Active Publication Date: 2020-09-01
CENT FOR EXCELLENCE IN MOLECULAR CELL SCI CHINESE ACAD OF SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is no vaccine or commercially available treatment for RSV in China, and only ribavirin is approved for the treatment of RSV infection, but has serious side effects
Palivizumab targets the conserved region of the F protein and has a broad spectrum of action. However, the current dose of Palivizumab required for each administration is 15 mg / kg (body weight), and passive immunization is performed 5 times a year. High, need multiple administrations, expensive disadvantages, and Palivizumab is a humanized antibody, which still retains part of the mouse sequence, may have certain immunogenicity, and has a certain degree of safety concern

Method used

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  • Anti-respiratory syncytial virus fully human broad-spectrum neutralizing antibody 4F1 and application thereof
  • Anti-respiratory syncytial virus fully human broad-spectrum neutralizing antibody 4F1 and application thereof
  • Anti-respiratory syncytial virus fully human broad-spectrum neutralizing antibody 4F1 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0193] Example 1 Obtaining antibody gene and antibody expression by single cell RT-PCR method

[0194] 1. Obtaining peripheral blood mononuclear cells (PBMC)

[0195] Peripheral blood was drawn from healthy volunteers, and conventional Ficoll-Paque (manufacturer: -H (CEDARLANE) density gradient centrifugation to obtain 10 7 Above peripheral blood mononuclear cells (PBMC).

[0196] Ficoll separation method:

[0197] (1) Collect blood, collect 20ml of whole blood in a 50ml centrifuge tube (pre-containing 1ml of 4% sodium citrate), invert and mix 8-10 times. (even if the final concentration of sodium citrate is 0.4%);

[0198] (2) Add an equal volume of RPMI1640 (containing sodium citrate), and mix well;

[0199] (3) Use a 15ml transparent centrifuge tube to spread 3ml of lymphocyte separation medium, and carefully add 6ml of blood sample on it. Form a separation interface (or 4ml separation solution plus 8ml blood sample);

[0200] (4) centrifuge at room temperature 800g...

Embodiment 2

[0226] Example 2 Antibody Characteristic Analysis

[0227] 1. ELISA detection of antibody binding antigen activity

[0228] In order to study the ability of the 4F1 antibody to bind to the F protein of RSV A and B viruses, ELISA was used to detect whether the expressed antibody recognized RSV A2 and B9320 pre-fusion F protein and A2 post-fusion F protein. A2F protein sequence reference UniProtKB / Swiss-Prot:P03420.1, B9320 sequence reference UniProtKB / Swiss-Prot:Q6V2E7, RSV pre-fusion F protein design is based on the strategy adopted by Jason S.McLellan.Science 2013, RSV post-fusion F The protein was designed according to the strategy adopted by Davide Corti.Nature 2013, and the whole gene was synthesized in Shanghai Jierui Company, and constructed on the expression vector of invitrogen pcDNA3.1. Expressed by mammalian cell CHO expression system; refer to Invitrogen ExpiCHO-S TM Expression System Handbook. Palivizumab (Palivizumab, ) is a positive control antibody, purchas...

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Abstract

The invention discloses a fully-human neutralizing antibody of anti-respiratory syncytial virus fusion protein and application of the fully-human neutralizing antibody. Specifically, the invention discloses a fully human monoclonal antibody 4F1 aiming at respiratory syncytial virus fusion protein (F protein) and pre-fusion F protein (preF protein), a nucleic acid sequence for encoding an antibodyand an antibody fragment, and a preparation method of the nucleic acid sequence. In-vitro and in-vivo experiments prove that the 4F1 antibody can effectively prevent and control RSV infection, has lowimmunogenicity for a human body, can avoid antibody-mediated immunological rejection of human anti-mouse and other species sources, and can be clinically used for preventing and treating respiratorysyncytial virus infection.

Description

Technical field [0001] The invention relates to the field of medicine, and specifically to a fully human broad-spectrum neutralizing antibody 4F1 against respiratory syncytial virus and its application. Background technique [0002] RSV causes acute upper and lower respiratory tract infections and is one of the important pathogens of respiratory tract infections in infants and young children worldwide. RSV is also recognized as a high-risk disease for certain adults, such as elderly individuals and adults with chronic lung disease. Important pathogens in individuals as well as immunocompromised adults (e.g., bone marrow transplant patients). There are 30 million new cases of infection worldwide every year, and the number of deaths due to RSV infection is approximately 200,000. RSV is the most common viral pathogen (17.0%) among pneumonia syndromes in children under 2 years old in China. There is an urgent need around the world for safe, effective, and affordable preventive...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/10A61K39/42A61P31/14G01N33/577G01N33/569
CPCC07K16/1027A61P31/14C07K2317/565C07K2317/56C07K2317/76C07K2317/92A61K2039/505A61K39/42G01N33/569G01N33/577A61K39/395C07K19/00C07K16/1003
Inventor 孙兵王宾凌志洋赵干伊春艳
Owner CENT FOR EXCELLENCE IN MOLECULAR CELL SCI CHINESE ACAD OF SCI
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