Construction method of CD73 humanized mouse model

A technology of mouse model and construction method, which is applied in the field of animal genetic engineering, can solve problems such as ethical restrictions, and achieve the effect of ensuring high efficiency

Active Publication Date: 2021-01-22
广东药康生物科技有限公司
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AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to provide a method for constructing a CD73 humanized mouse model, which has the advantages of efficient screening and application of CD73-targeted drugs in combination with other immune checkpoints or small molecule drugs, and solves the problem of human diseases The study of pathogenesis and the screening of effective therapeutic drugs require a large number of preclinical trials, and the direct use of human cells and tissues for preclinical research will be subject to ethical restrictions

Method used

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  • Construction method of CD73 humanized mouse model
  • Construction method of CD73 humanized mouse model
  • Construction method of CD73 humanized mouse model

Examples

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Embodiment 1

[0048] Example 1 Construction of CD73 humanized mouse model

[0049] The construction method of CD73 humanized mouse model is completed according to the following steps:

[0050] 1. Determine the replacement region of the human fragment and the inserted human sequence: use CRISPR Cas9 to replace the human CD73 coding gene with the mouse Cd73 coding gene to construct a mouse model that can express human CD73. The amino acid sequence (Aa: 1-574) encoded by the selected human CD73 gene is shown in the following sequence SEQ ID NO: 1; the amino acid sequence encoded by human CD73 is inserted behind the 5'UTR of the mouse CD73 gene, and a terminator sequence 3× is added to the end polyA prevents the expression of mouse CD73 protein, the strategy diagram is as follows. The human CD73 protein is regulated by the promoter and 5'UTR of the mouse CD73 gene, so that the expression of the human CD73 protein is regulated by the mouse sequence to maintain the similarity in the expression s...

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Abstract

The invention discloses a construction method of a CD73 humanized mouse model. The CD73 humanized mouse model is efficiently and successfully obtained by optimizing a targeting carrier and sgRNA usedin model preparation and verification steps. The model has remarkable advantages in screening application of CD73 targeted drugs and other immune checkpoints or small molecule drug combination.

Description

technical field [0001] The invention relates to the technical field of animal genetic engineering, in particular to a method for constructing a CD73 humanized mouse model. Background technique [0002] The first-generation immune checkpoint inhibitors, including PD1, PDL1, and CTLA4, have become the main force in tumor immunotherapy. Although these immune checkpoints have achieved considerable success in tumor immunotherapy, only a small number of patients show a sustained response to a single immune checkpoint, and some patients even do not respond to currently marketed checkpoint inhibitors. For how to overcome the drug resistance of solid tumor treatment, one of the most important aspects is the regulation of the tumor microenvironment. Regulating the immunosuppressive effect in the microenvironment can significantly improve the effect of tumor immunotherapy. [0003] CD73 is an extracellular-5'-nucleotidase, a glycoprotein anchored to the plasma membrane by glycosylphos...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/85A01K67/027C12N15/55
CPCC12N15/8509C12N9/16C12Y301/03005A01K67/0278A01K2207/15A01K2227/105A01K2267/0331C12N2800/107Y02A50/30
Inventor 琚存祥王韬杨笑柳于薇薇
Owner 广东药康生物科技有限公司
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