EpCAM single domain antibody G7

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

Inactive Publication Date: 2018-09-04
NANJING MATERNITY & CHILD HEALTH CARE HOSPITAL
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  • Summary
  • Abstract
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AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to disclose a kind of EpCAM single-domain antibody G7 in order to solve the problem of the unsatisfactory clinical trial results of CD326 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 CD326 Nanobody Library

[0057] (1) Purchase commercially available CD326 recombinant protein (ORIGENE Company) Other commercially available EpCAM recombinant proteins can be used here, and the purchase route of EpCAM recombinant protein does not affect the purpose of subsequent experiments. The purchased recombinant protein was taken out according to the instructions, mixed with an equal volume of incomplete Freund's adjuvant and fully emulsified, and used as a vaccine to immunize alpaca.

[0058] (2) The method of subcutaneous multi-point injection was used to immunize two adult male alpacas for 5 consecutive immunizations with a one-week interval between each time to promote the production of CD326 antigen-specific nanobodies;

[0059] (3) After 5 times of immunization, according to the detection titer (before each immunization, 5ml of blood was collected from the common carotid artery, the upper serum was collected by centrifugation, and stored at...

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Abstract

The invention belongs to the field of molecular biology, and particularly relates to an EpCAM single domain antibody G7 and a nucleotide sequence for encoding the EpCAM single domain antibody G7. By utilizing a phage display technology, a single domain heavy chain antibody library specifically bound with a target antigen CD326 is screened from a camel-source immunizing single domain heavy chain antibody library, and a coding sequence of a positive clone is further guided into a prokaryotic expression vector, so that the high-efficiently expressed EpCAM single domain antibody G7 is obtained. Therefore, a basis is provided for clinic targeted immunological therapy of CD326.

Description

technical field [0001] The present invention relates to EPCAM single-domain antibody G7 and the VHH chain constituting the antibody, and belongs to the field of molecular biology. Background technique [0002] CD326 refers to Epithelial Cell Adhesion Molecule, also known as EpCAM, which is a kind of glycoprotein widely expressed in human epithelial tissue and has a very potent epitope. CD326 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 CD326 activates the expression of proto-oncogenes such as c-myc and cyclinA / E by participating in the β-catenin-dependent Wnt cascade, leading to tumorigenesis. [0004] Under physiological conditions, CD326 is expressed to varying degrees in all normal epithelia except squamous epithelial cells, and is mostly located in the tight junct...

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 季晨博郭锡熔崔县伟
Owner NANJING MATERNITY & CHILD HEALTH CARE HOSPITAL
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