Anti-human interleukin 5 (1L-5) monoclonal antibody and application thereof

An antibody, recombinant protein technology, applied in the field of biomedicine, can solve problems such as lack of

Pending Publication Date: 2020-06-19
SHANGHAI PHARMAEXPLORER
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] In order to overcome the current lack of fully human IL-5 antibody and the shortcomings in the activity of existing IL-5 antibodies, the present invention provides a high affinity and strong specificity IL-5 antibody and its preparation method and use

Method used

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  • Anti-human interleukin 5 (1L-5) monoclonal antibody and application thereof
  • Anti-human interleukin 5 (1L-5) monoclonal antibody and application thereof
  • Anti-human interleukin 5 (1L-5) monoclonal antibody and application thereof

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preparation example Construction

[0306] The preparation method of the nucleic acid is a conventional preparation method in the art. Preferably, it includes the following steps: obtaining the nucleic acid molecule encoding the above-mentioned protein by gene cloning technology, or obtaining the nucleic acid molecule encoding the above-mentioned protein by artificial full-sequence synthesis .

[0307] Those skilled in the art know that substitutions, deletions, alterations, insertions or additions can be appropriately introduced into the base sequence encoding the amino acid sequence of the above protein to provide a polynucleotide homologue. The homologue of the polynucleotide in the present invention can be prepared by replacing, deleting or adding one or more bases in the gene encoding the protein sequence within the scope of maintaining antibody activity.

[0308] carrier

[0309] The invention also provides a recombinant expression vector comprising the nucleic acid.

[0310] The recombinant expression v...

Embodiment 1

[0383] Example 1: Expression and purification of recombinant human IL-5

[0384] Using the PCR method, the 3' end of the DNA fragment encoding human IL-5 protein amino acid sequence (NP_000870.1) Met1-Ser134 was added to the DNA encoding six histidines, and the obtained encoding his tag recombinant human IL-5 protein The DNA fragments were cloned into expression vectors by molecular biology methods. Escherichia coli is used to amplify, the plasmid is extracted and purified by alkaline lysis, and the expression plasmid is transiently transfected to express the recombinant protein using insect cell SF21. After 5-7 days of culture, centrifuge and filter to collect the cell culture supernatant. The recombinant human IL-5 protein with His tag in the supernatant was purified by Ni-NTA affinity chromatography, and then further purified by using a molecular sieve column to remove impurities such as macromolecular polymers. The purified protein was stored in PBS buffer, filtered thro...

Embodiment 2

[0389] Example 2: Preparation of anti-human IL-5 antibody using hybridoma technology

[0390] 2.1 Mice were immunized with recombinant human IL-5 protein

[0391] For protein immunization, 6-8 week-old Balb / c, SJL / J mice (provided by Shanghai SLAC) were used. Mice were reared under SPF conditions after receipt. The initial immunization dose was 50 micrograms of recombinant human IL-5 protein per mouse. After the protein was emulsified with complete Freund's adjuvant, 0.25 ml was injected subcutaneously into the tail. Two weeks after the initial immunization, a booster immunization was given. Recombinant human IL-5 protein (25 μg protein per mouse) was emulsified with Freund's incomplete adjuvant and injected 0.25 ml intraperitoneally. After each booster immunization interval of 3 weeks. Serum samples were collected one week after each booster immunization, the antibody titer in the serum was detected by ELISA and the activity of the antibody in the serum was detected by r...

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Abstract

The invention discloses an IL-5-targeted antibody, and a preparation method and application thereof. Specifically, the invention discloses a novel IL-5-targeted murine or chimeric monoclonal antibody.The invention also discloses a method for preparing the monoclonal antibody. The monoclonal antibody can be combined with an IL-5 antigen with high specificity, has high affinity, and can well alleviate a series of asthma symptoms caused by IL-5, thereby playing a role in treating asthma.

Description

technical field [0001] The present invention relates to the field of biomedicine, in particular to an IL-5 antibody and its preparation method and application Background technique [0002] Bronchial asthma (referred to as asthma) is a common chronic airway inflammatory disease, usually accompanied by increased airway responsiveness, recurrent wheezing, shortness of breath, chest tightness and (or) cough and other symptoms. After the 1970s, asthma became widespread. By 2011, an estimated 235 million to 300 million people were affected worldwide, and approximately 250,000 people lost their lives as a result. Clinically, asthma is usually controlled with inhaled steroids and other glucocorticoids. For those with unsatisfactory disease control, adjuvant therapy such as inhaled long-acting β2 agonists, slow-release theophylline, or leukotriene regulators can be added. However, about 10% of patients still cannot control their disease well through conventional treatment methods....

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/24C12N15/13C12N15/70C12N1/21G01N33/577A61K39/395A61K47/68A61P11/00A61P11/06A61P17/00A61P29/00
CPCC07K16/244C12N15/70G01N33/577A61K47/6845A61P11/00A61P11/06A61P17/00A61P29/00C07K2317/51C07K2317/515C07K2317/56A61K2039/505G01N33/6869G01N2333/5409C07K2317/565C07K2317/76C07K2317/33C07K2317/92C07K2319/21A61K35/17C07K2317/24
Inventor 杨沂蓁杨松霖吴建杨欣秀邵小慧钟勤胡少平段清刘礼乐王冬旭戴朝晖
Owner SHANGHAI PHARMAEXPLORER
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