A kind of nucleic acid molecule for preparing humanized antibody and its application

A nucleic acid molecule and carrier technology, which is applied to nucleic acid molecules and their application in the preparation of humanized antibodies to achieve high efficiency

Active Publication Date: 2020-09-29
CHONGQING JINMAIBO BIOTEC CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The disadvantage of this method is that secondary transformation is required to obtain fully humanized antibodies

Method used

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  • A kind of nucleic acid molecule for preparing humanized antibody and its application
  • A kind of nucleic acid molecule for preparing humanized antibody and its application
  • A kind of nucleic acid molecule for preparing humanized antibody and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Transfer the modified human immunoglobulin gene into mice, and then immunize mice containing human immunoglobulin genes to obtain fully humanized antibodies. The specific steps are as follows:

[0044] 1. Optimization and transformation of human immunoglobulin genes

[0045] 1) Transformation of human immunoglobulin heavy chain gene

[0046] The human CD79 gene is transferred to the YAC or BAC vector containing human Ig through homologous recombination, and the gene cluster of the human immunoglobulin heavy chain gene is constructed as follows: figure 2 Shown (the position in the black box is the sequence of the imported human CD79 gene). Including the V region, D region and J region of the human immunoglobulin heavy chain gene, hIgHC μ, hIgHC δ, hIgHC γ3, hIgHC γ1, hIgHC α1, hLCR, hCD79. The internal structure of hCD79 is as follows Picture 1-1 As shown, its gene sequence is shown in SED ID NO.1; including the coding sequence of CD79α protein peptide and CD79β prot...

Embodiment 2

[0143] The transformed human immunoglobulin gene was transferred into a pig as a host animal (immunoglobulin gene knockout pig) to obtain the following results:

[0144] Detection of human IgG expression in pig blood: Two transgenic pigs, 205 and 206, which were positive in PCR detection, were selected to collect blood, separate serum, and detect the expression of human IgG in blood. as comparison. The antibodies identified by ELISA were: sigma(I1886) and sigma(A8792).

[0145] The result is as follows:

[0146] 4115: 0.5mg / ml

[0147] 4116: 0.1mg / ml

[0148] 4117: 0.2mg / ml

[0149] Healthy adults: 10.2mg / ml

[0150] Ordinary pig: <0.1ng / ml

[0151] The results show that the transferred gene can express human antibody heavy chain protein in pigs.

[0152] The modification scheme of the human immunoglobulin gene of the present invention is also applicable to other mammals, for example, pigs, rabbits, sheep, horses, etc. as host animals can also achieve the beneficial eff...

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Abstract

Provided are nucleic acid molecules, comprising human immunoglobulin genes or fragments thereof, and a human CD79 gene sequence. Also provided is a method for preparing, by the nucleic acid molecules, a transgenic animal used for producing a humanized antibody. The nucleic acid molecules solve the problem of incompatibility of human immunoglobulin genes in different species due to interactions between BCR and Igα and between BCR and Igβ, and the expressed humanized antibody does not require secondary modification.

Description

technical field [0001] The invention belongs to the field of biotechnology, and mainly relates to a nucleic acid molecule and its application in the preparation of humanized antibodies. Background technique [0002] Antibodies are an important class of biomedical products, which play an important role in the prevention and treatment of human diseases. The development of therapeutic antibodies has gone through different development stages such as murine antibodies, chimeric antibodies, reshaped antibodies, resurfaced antibodies and fully humanized antibodies. Fully humanized antibody (Full Humanized Antibody) refers to an antibody that is completely identical to the protein sequence of a human antibody obtained through genetic modification or immunization of transgenic animals. Since fully humanized antibodies do not contain animal protein, they have lower side effects and better effects, and have become the main research and development direction of current and future antib...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/13C12N15/12C07K16/00C12N15/85A01K67/027
CPCA01K67/027C07K16/00C12N15/85
Inventor 刘作华葛良鹏刘雪芹邹贤刚丁玉春黄勇杨松全游小燕林保忠吴梦
Owner CHONGQING JINMAIBO BIOTEC CO LTD
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