Whole-humanized neutralizing antibody resisting H3N2 influenza virus and application of whole-human neutralizing antibody

A technology of antibodies and antibody drugs, which is applied in the field of medicine and can solve problems such as limited applications

Inactive Publication Date: 2020-05-08
INST PASTEUR OF SHANGHAI CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In 1986, the first murine monoclonal antibody for the treatment of rejection in organ transplantation—murmonab CD3 (murmonab CD3, hoclone OKT3) was approved by the US FDA for marketing, but because it can produce human anti-mouse antibodies in the human body ( HAMA) reaction, which limits the application

Method used

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  • Whole-humanized neutralizing antibody resisting H3N2 influenza virus and application of whole-human neutralizing antibody
  • Whole-humanized neutralizing antibody resisting H3N2 influenza virus and application of whole-human neutralizing antibody
  • Whole-humanized neutralizing antibody resisting H3N2 influenza virus and application of whole-human neutralizing antibody

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0188] Example 1 Obtaining antibody gene and antibody expression by single cell RT-PCR method

[0189] 1. Obtaining peripheral blood mononuclear cells (PBMC)

[0190] Peripheral blood was drawn from volunteers screened according to the hemagglutination inhibition test, and conventional Ficoll-Paque density gradient centrifugation was used to obtain 10 7 Above peripheral blood mononuclear cells (PBMC).

[0191] 2. HA-specific memory B cell sorting

[0192] Using FITC-CD19 / APC-IgG / Cy3-HA as a marker, specific B cells were obtained by flow cytometry to a 96-well RT-PCR plate, one cell per well, and several HA-specific memory B cells were obtained.

[0193] 3. Antibody gene

[0194] Use RT-PCR and Nested-PCR techniques to obtain the antibody gene (heavy and light chain variable region gene), connect the variable region of the antibody heavy and light chain gene from the same B cell to the T vector, and perform sequencing to verify the antibody gene. Then the heavy and light ch...

Embodiment 2

[0225] Example 2 Antibody Characteristic Analysis

[0226] 1. Antigen Specific Detection

[0227] Detect whether the expressed fully human antibody recognizes HA protein and H3N2 influenza virus vaccine lysate. For detection by ELISA, coat HA and H3N2 influenza vaccine lysates on ELISA plates respectively, 10 μg / ml, 100 μl per well, overnight at 4°C;

[0228] 1) Wash the plate with PBST, 3 times;

[0229] 2) Blocking: 1% BSA, 200 μl per well, 37°C, 2h;

[0230] 3) Wash the plate with PBST, 3 times;

[0231] 4) According to the measured concentration, adjust the whole human antibody cells to the same concentration, 100 μl per well, 37°C, 2h;

[0232] 5) Wash the plate with PBST, 3 times;

[0233] 6) Goat anti-human IgG (Fc specific)-Peroxidase antibody, diluted 1:10000, 100 μl per well was added to the ELISA plate, 37°C, 1h;

[0234] 7) Wash the plate with PBST, 3 times;

[0235] 8) Substrate A solution: B solution = 1:1, 100 μl per well, 37 ° C, 15 min, dark reaction; ...

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PUM

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Abstract

The invention discloses a whole-humanized neutralizing antibody resisting an H3N2 influenza virus and application of the hole-human neutralizing antibody. Particularly, the invention discloses a whole-humanized monoclonal antibody for the H3N2 influenza virus. The antibody can bind influenza virus hemagglutinin (HA) with a native conformation, can prevent the H3N2 subtype influenza virus from infecting susceptible cells, has lower immunogenicity for human bodies, can avoid medicated immune rejection response of antibodies derived from other specifies such as a human anti-mouse antibody, and has potential values of preventing and treating H3N2 subtype influenza virus infection clinically.

Description

technical field [0001] The invention relates to the field of medicine, in particular to a fully human neutralizing antibody against H3N2 influenza virus and its application. Background technique [0002] According to the survey of WHO, about 5-10% of adults and 20-30% of children suffer from seasonal influenza every year in the world, resulting in 3-5 million hospitalizations and 290-650 thousand deaths. In my country, up to 65 million to 190 million people suffer from influenza every year. The seasonally circulating strains are mainly H1N1, H3N2 and B influenza viruses. In recent years, H3N2 viruses have frequently circulated. In 2017, the H3N2 virus was prevalent in Hong Kong. As of August, there were about 400 deaths. In the winter of the same year, the "Australian H3N2 Influenza" spread rapidly in the UK. Except for a few areas, almost all the states in the UK fell. As of mid-January 2017, the number of influenza infection cases reached 2,000. Therefore, the prevalen...

Claims

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Application Information

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IPC IPC(8): C07K16/10C07K19/00C12N5/10A61K39/42A61P31/16G01N33/569
CPCA61K2039/505A61P31/16C07K14/7051C07K16/1018C07K2317/56C07K2317/565C07K2317/76C07K2319/30C12N5/0636C12N2510/00G01N33/56983
Inventor 孙兵孙晓玉卢晓凌志洋
Owner INST PASTEUR OF SHANGHAI CHINESE ACADEMY OF SCI
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