Human derived monoclonal antibody for identifying activated integrin alpha 4 beta 7

A monoclonal antibody and integrin technology, applied in the direction of antibodies, anti-receptors/cell surface antigens/cell surface determinant immunoglobulins, anti-inflammatory agents, etc., can solve the problem of non-specific antibody recognition

Inactive Publication Date: 2013-10-30
SHANGHAI INST OF BIOLOGICAL SCI CHINESE ACAD OF SCI
View PDF2 Cites 12 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this antibody has no specificity in recognition, it can recognize the activated form of α 4 Integrins also recognize the inactive form of α 4 Integrin, so in the process of clinical application, it will cause progressive multifocal leukoencephalopathy in some patients
Up to now, there is no integrin α with very good recognition specificity in this field 4 beta 7 Antibody drugs for clinical use

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Human derived monoclonal antibody for identifying activated integrin alpha 4 beta 7
  • Human derived monoclonal antibody for identifying activated integrin alpha 4 beta 7
  • Human derived monoclonal antibody for identifying activated integrin alpha 4 beta 7

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0072] Embodiment 1, integrin α 4 beta 7 Construction of wild-type and mutant soluble protein expression vectors and protein expression and purification

[0073] The following description is based on α 4 The amino acid sequence of the subunit (human) is shown in SEQ ID NO: 33 (999aa). The following description is based on β 7 The amino acid sequence of the subunit (human) is shown in SEQ ID NO: 9 (779aa).

[0074] It has been confirmed that an N-glycosylation site is introduced between the I and Hybrid domains of the integrin β subunit, which can activate the integrin molecule. Based on this, construct the full-length integrin α 4 beta 7 Wild-type and mutant soluble protein expression vectors with this mutation, α 4 The subunit has a splice body in the body, so in order to obtain the full-length α 4 Subunit, the inventors mutated the splice site R558A (ie the amino acid at position 558 was mutated from R to A). Soluble integrins do not have transmembrane and intracell...

Embodiment 2

[0077] Example 2. Using phage antibody library technology to screen and express antibodies that specifically recognize activated integrins

[0078] (1) Using purified wild-type non-activated and mutant-activated α 4 beta 7 Integrin protein, by panning (panning), screening scFv antibody library Tomlinson I+J (purchased from Geneservice), after three rounds of screening and removal of wild-type non-activated α 4 beta 7 Integrin protein combined with phage, and finally can be able to express specific binding activated α 4 beta 7 Integrin scFv phage. The specific screening process is to first label the soluble integrin protein with biotin, and incubate it with the human single-chain antibody phage library. The coated magnetic beads are mixed with it and incubated, and all the biotin-labeled integrins are immobilized on the magnetic beads, and the phages that bind the integrin are also immobilized on the magnetic beads, and finally the magnetic beads are separated by a magneti...

Embodiment 3、J19

[0101] Example 3, J19 specifically recognizes activated integrin α 4 beta 7

[0102] After the full-length J19 was purified, in order to further verify its recognition specificity, the inventors used flow cytometry detection method.

[0103] figure 2 The specific process of A experiment: Stable expression of integrin α 4 beta 7 K562 cells were treated with 1mM Ca 2+ +1mM Mg 2+ and 2mM Mn 2+ Wash twice with HBS buffer, then add J19 (10μg / ml), add the same amount of hIgG (purchased from Pierce) to the control, and incubate at 4°C for 0.5 hours; 2+ +1mM Mg 2+ and 2mM Mn 2+ Wash twice with HBS buffer respectively, then add Alexa Fluor 488-labeled goat anti-human IgG (1:500) (purchased from Invitrogen), and incubate at 4°C for 0.5 hours; 2+ +1mM Mg 2+ and 2mM Mn 2+ Wash twice with HBS buffer, and detect with a flow cytometer (purchased from BD).

[0104] figure 2 Experimental procedure of B: basic operation and figure 2 A is the same, except that before adding the...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention relates to a novel human derived monoclonal antibody for identifying activated integrin alpha 4 beta 7. The monoclonal antibody for specifically identifying activated integrin alpha 4 beta 7 is obtained by screening by an inventor. Based on the identification specificity, the monoclonal antibody can be used as a carrier for targeting transportation to take a medicament to positions of pathological tissues so as to fulfill an aim of treating diseases; and the monoclonal antibody also can be used as a tool for researching the activation status of the integrin alpha 4 beta 7.

Description

technical field [0001] The invention belongs to the field of biotechnology. Specifically, the present invention relates to a novel recognition of the activated form of integrin alpha 4 beta 7 humanized monoclonal antibodies. Background technique [0002] The migration of leukocytes is an important step and key link in the pathological process of inflammatory response, and it is a hot and frontier topic in the field of life science research. As an important class of cell surface adhesion molecules, integrins are important proteins that directly mediate the migration of leukocytes. Integrins are heterodimers composed of α and β subunits through non-covalent bonds. In vertebrates, 18 α subunits and 8 β subunits have been found to form 24 integrins. They are compatible with biological It is closely related to the immune response of the body, the movement and migration of cells, the tissue localization of immune cells, and blood coagulation. Integrin alpha 4 beta 7 It is m...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/28C12N15/13C12N15/63C12N1/21C12N1/19C12N5/10G01N33/68G01N33/577A61K39/395A61P29/00A61P1/00A61P9/00A61P7/02A61P35/00
Inventor 陈剑峰齐俊鹏
Owner SHANGHAI INST OF BIOLOGICAL SCI CHINESE ACAD OF SCI
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap