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EpCAM (epithelial cell adhesion molecule) Single-domain antibody E6

A single-domain antibody and anti-tumor drug technology, applied in the field of molecular biology, can solve the problems of unsatisfactory clinical trial results of immune-directed therapy, and achieve the effect of improving specific binding performance, small molecular weight, and weak immunogenicity

Active Publication Date: 2018-09-14
NANJING MATERNITY & CHILD HEALTH CARE HOSPITAL
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AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to disclose an EpCAM single-domain antibody for the problems of the unsatisfactory clinical trial results of EpCAM immune-directed therapy in the prior art.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] 1. Construction of EpCAM single-domain antibody library

[0057] (1) Purchasing commercially available EpCAM recombinant proteins (ORIGENE Company) Here, other commercially available EpCAM recombinant proteins can be used, and the way to purchase EpCAM recombinant proteins will not affect the realization of the purpose of subsequent experiments. According to the instructions, the purchased recombinant protein was taken out, mixed with an equal volume of Freund's incomplete adjuvant and fully emulsified, and used as a vaccine to immunize alpacas.

[0058] (2) Using the method of subcutaneous multi-point injection, immunize two adult male alpacas for 5 consecutive immunizations, with a one-week interval between each time, to promote the production of EpCAM antigen-specific single-domain antibodies;

[0059] (3) After 5 times of immunization, according to the detection titer (take 5ml of blood from the common carotid artery before each immunization, collect the upper serum...

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Abstract

The invention belongs to the field of molecular biology and particularly relates to EpCAM (epithelial cell adhesion molecule) single-domain antibody E6. A single-domain heavy-chain antibody library capable of specifically binding with target antigen EpCAM is screened from a camelid source immunized single-domain heavy-chain antibody library by means of a phage display technique; a gene sequence toencode the antibody is cloned to a prokaryotic expression vector so as to obtain the efficient-expression EpCAM single-domain antibody. The specific binding performance of the single-domain antibodycan be significantly improved, and basis is laid for immuno-guided clinical therapy of EpCAM.

Description

technical field [0001] The invention relates to an EpCAM single domain antibody and a preparation method thereof, belonging to the field of molecular biology. Background technique [0002] EpCAM refers to epithelial cell adhesion molecule (Epithelial Cell Adhesion Molecule,), also known as CD326, is a kind of glycoprotein widely expressed in human epithelial tissue, and has a very potent antigenic epitope. EpCAM not only has the function of epidermal cell adhesion molecule, but also participates in multiple life activities such as accelerating cell cycle, promoting cell proliferation, differentiation, migration and immune escape. [0003] Studies have shown that EpCAM activates the expression of proto-oncogenes such as c-myc and cyclinA / E by participating in the β-catenin-dependent Wnt cascade reaction, leading to tumorigenesis. [0004] Under physiological conditions, EpCAM is expressed to varying degrees in all normal epithelium except squamous epithelial cells, and most ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/30C12N15/13C12N15/70C12N1/21A61K39/395A61P35/00G01N33/574
CPCA61K39/00A61P35/00C07K16/30C07K2317/569G01N33/57484
Inventor 季晨博
Owner NANJING MATERNITY & CHILD HEALTH CARE HOSPITAL
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