Preparation method of glp-1 or its analogue and antibody fc fragment fusion protein

Abstract

The invention relates to the field of biotechnology, in particular to a method for preparing a fusion protein of GLP-1 or its analogue and an antibody Fc fragment. The present invention provides a preparation method of fusion protein of GLP-1 or its analogue and antibody Fc fragment, comprising the following steps: cloning the coding sequence of fusion protein of GLP-1 or its analogue and antibody Fc fragment into an expression vector; expressing The vector transfects CHO-DXB11 cells, cultures and screens out positive cell lines; the resulting cells are expressed and purified to obtain the fusion protein. The preparation method provided by the present invention expresses the GLP-1-Fc molecule in the CHO cell line DXB11 without degradation problems, the subsequent purification yield is greatly improved, and the biological activity is not weakened, which is very suitable for the needs of industrialized production.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a preparation method of fusion protein of GLP-1 or its analogue and antibody Fc fragment. Background technique [0002] Glucagon-like peptide-1 (GLP-1) promotes insulin release, reduces plasma glucagon levels, reduces the rate of gastric emptying, promotes satiety and stimulates islet biosynthesis and β-cell proliferation , can be used for the treatment of type 2 diabetes. Type 2 diabetes is common in our country. If it is not treated in time, it will easily transform into type 1 diabetes, which will cause a great burden to patients and the country. Therefore, the demand for developing such drugs is extremely urgent. At present, Exenatide (Byetta) of Eli Lilly and Company and Liraglutide (Novoli) of Danish Novo Nordisk Pharmaceutical Company have obtained import registration in my country. Both drugs have the effect of lowering blood sugar and reducing weight. effect, but requires ...

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Problems solved by technology

[0006] In the same article, it was mentioned that protease inhibitors (benzamidine hydrochloride) were added to control the degradation during the cell culture process, but the risk of drugging the product still increased, and it was also necessary to inactivate and eradicate the protease inhibitors in the subsequent process. It will make the preparation process of the whole product more complicated and uneconomical, and the addition of protease inhibitors can only reduce the degree of degradation of the N-terminal of the GLP-1-Fc molecule, but cannot effectively inhibit its occurrence, and cannot really solve the degradation problem

Another method to remove the N-terminal degradation products of GLP-1-Fc molecules is to use various chromatographic column chromatography for multi-step purification. Although very pure GLP-1-Fc fusion proteins can also be obtained, the whole process is Yield drops significantly below 20%, much lower than yield levels of greater than 70% for therapeutic antibody drugs

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