Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

A fully human antibody or antibody fragment against CD19 and its method and application

A fully human antibody, antibody technology, applied in the field of biological treatment of malignant tumors, can solve the problem of poor targeting of cellular immunotherapy

Active Publication Date: 2021-03-02
CHANGZHOU VELOX PHARMA SCI & TECH CO LTD
View PDF4 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] The object of the present invention is to provide an anti-CD19 fully human antibody or antibody fragment and its method and application. Source CD19 protein binding, using chimeric antigen receptor T cell technology (CAR-T technology) to engineer the expression of human antibody fragments integrated in CAR and binding to CD19 in T cells, resulting in chimeric antigen receptor T cells Can be used to treat hematological cancers associated with CD19 expression

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
  • A fully human antibody or antibody fragment against CD19 and its method and application
  • A fully human antibody or antibody fragment against CD19 and its method and application
  • A fully human antibody or antibody fragment against CD19 and its method and application

Examples

Experimental program
Comparison scheme
Effect test

experiment example 1

[0092] Experimental example 1 Screening of anti-CD19 single chain antibody

[0093] 1. Establish a phage antibody library of fully human single-chain antibody

[0094] Design primers to amplify the heavy chain variable region and light chain variable region of the antibody, connect the heavy chain variable region and light chain variable region with Linker by overlapping extension PCR, and obtain the full-length PCR product, use SfiI Digest the PCR product and the pCGMT phagemid vector, electrotransform the ligated transformation product into XL1-blue competent cells, then add 3 mL of SOC medium to the competent cells, culture at 30°C for 1 hour, and add a final concentration of 50 μg / mL of ampicillin, 20 mL of 10 μg / mL tetracycline solution, and shake at 37°C for 2 hours. Then add a concentration of 10 13 / mL VCSM13 helper phage 50 μL, incubate at room temperature for 1 h, and shake once every 10 min. Shake culture at 37°C for 2 hours, then add kanamycin at a final concen...

experiment example 2

[0108] Preparation of Experimental Example 2 Anti-CD19 Single Chain Antibody (Expression and Purification)

[0109] The monoclonal plasmid obtained by screening in Experimental Example 1 was digested by restriction endonuclease SfiI, ligated, and then the fragment was inserted into the pFUSE expression vector by the method of recombination (the pFUSE expression vector was digested by restriction endonuclease SfiI site), thereby obtaining the pFUSE expression vector of the antibody of the present invention.

[0110] Mix the 293Fectin transfection reagent with the eukaryotic antibody expression vector obtained above at a volume-to-mass ratio of 30 μL: 30 μg, add 30 μL of 293Freestyle suspension cells, culture on a shaker at 125 rpm at 37°C for 48-72 hours, and collect after centrifugation Supernatant, using HiTrap Protein A HP columns in The antibody protein was purified on a protein purification instrument to obtain an anti-CD19 single-chain antibody (anti-CD19 fully human sc...

experiment example 3

[0111] Experimental example 3 Detection of binding specificity and binding kinetics characteristics of anti-CD19 fully human scFv antibody

[0112] The affinity and kinetic characteristics of the antibody and antigen were detected by the method of multi-cycle kinetics, and the antibody was immobilized by the capture method. The human scFv sample is diluted and flows over the surface of the chip, diluted to a concentration of 7.5 μg / mL and 15 μg / mL, the anti-CD19 fully human scFv antibody will be captured by the CD19 antigen, and the signal will be detected and recorded after the antigen is combined with the antibody. Finally, the antibody and antigen samples on the surface of the sensor were all eluted with a regeneration reagent (glycine solution at pH 1.5), and a new round of detection was performed.

[0113] Experimental results:

[0114] Such as figure 2 Shown is the binding kinetics curve of anti-CD19 fully human scFv antibody to CD19, and each line represents differen...

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 present invention discloses an anti-CD19 fully human antibody or antibody fragment as well as methods and applications thereof. The antibody or antibody fragment comprises: a heavy chain and a light chain; the heavy chain and the light chain comprise a variable region, and the variable region comprises a complementary decision area. The complementarity determining regions CDR1, CDR2 and CDR3 of the heavy chain are represented by HCDR1, HCDR2 and HCDR3 respectively; the complementarity determining regions CDR1, CDR2 and CDR3 of the light chain are represented by LCDR1, LCDR2 and LCDR3 respectively; the amino acid sequence of HCDR1 includes SEQ ID NO: 3 The amino acid sequence of HCDR2 includes SEQ ID NO: 4; the amino acid sequence of HCDR3 includes SEQ ID NO: 5; the amino acid sequence of LCDR1 includes SEQ ID NO: 6; the amino acid sequence of LCDR2 includes DVS; the amino acid sequence of LCDR3 includes SEQ ID NO: 7. The antibody or antibody fragment of the present invention can bind to human CD19 protein with high specificity, and the chimeric antigen receptor T cell technology is used to engineer expression and integration in T cells, and the obtained chimeric antigen receptor T cells can be used for treatment Hematologic cancers associated with CD19 expression.

Description

technical field [0001] The invention belongs to the technical field of biological treatment of malignant tumors, and relates to an anti-CD19 antibody, in particular to an anti-CD19 fully human antibody or antibody fragment as well as a method and application thereof. Background technique [0002] CD19 is expressed on B lineage cells (excluding mature plasma cells) and follicular dendritic cells. It is an important signal transduction molecule that regulates the growth activation and activation of B lymphocytes, and regulates B lymphocyte antigen receptors or It plays an important role in the signal threshold of other surface receptors. It is an important membrane antigen related to the differentiation, activation, proliferation and antibody production of B lymphocytes. It is the best marker for diagnosing B lymphocyte tumors and identifying B lymphocytes. [0003] Many patients with B-cell malignancies cannot be cured using standard therapies. In addition, traditional treat...

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 Patents(China)
IPC IPC(8): C07K16/28C07K19/00C12N15/13C12N5/10A61K35/17A61P35/02
CPCA61K35/17C07K14/7051C07K16/2803C07K2317/622C07K2317/92C07K2319/02C07K2319/33
Inventor 杨光闵晨雨
Owner CHANGZHOU VELOX PHARMA SCI & TECH CO LTD
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

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

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com