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A kind of anti-human pdl1 monoclonal antibody and its application

A monoclonal antibody, antibody technology, applied in the direction of antibodies, antibody medical components, anti-animal/human immunoglobulins, etc., can solve the problems of inducing immune response, unproven cancer patients, etc., and achieve the effect of novel sequence

Active Publication Date: 2022-05-10
BIOSION INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] To date, no satisfactory method has been demonstrated to induce potent immune responses in cancer patients

Method used

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  • A kind of anti-human pdl1 monoclonal antibody and its application
  • A kind of anti-human pdl1 monoclonal antibody and its application
  • A kind of anti-human pdl1 monoclonal antibody and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1 Obtaining specific anti-PDL1 mouse monoclonal antibody by fusion hybridoma technology

[0043] 1.1 Animal immunity

[0044] Mice were immunized according to the general method in the literature (E Harlow, D. Lane, Antibody: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1998). 免疫原为重组人PDL1-Fc蛋白(公司自产,插入氨基酸序列为FTVTVPKDLYVVEYGSNMTIECKFPVEKQLDLAALIVYWEMEDKNIIQFVHGEEDLKVQHSSYRQRARLLKDQLSLGNAALQITDVKLQDAGVYRCMISYGGADYKRITVKVNAPYNKINQRILVVDPVTSEHELTCQAEGYPKAEVIWTSSDHQVLSGKTTTTNSKREEKLFNVTSTLRINTTTNEIFYCTFRRLDPEENHTAELVIPELPLAHPPNERTEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK)。 Recombinant human PDL1 with his-tagged protein (ACRO, Cat#PD1-H5229) was used as the detection antigen for determination of serum titer and hybridoma screening. Briefly, rem...

Embodiment 2

[0050] Example 2 In vitro analysis method for determining the functional activity of PDL1 monoclonal antibody

[0051] 2.1 Determination of antibody binding ability based on capture ELISA

[0052] Prepare goat anti-mouse IgG F(ab') in 1xPBS 2 The specific secondary antibody (Jackson ImmunoResearch Laboratories, Inc., Cat#115-005-072) was added to a 96-well microtiter plate at a final concentration of 2 μg / ml at 100 μl / well, and coated at 37° C. for 2 hours. After coating, the plate was washed once with PBS solution containing 0.05% Tween 20 (i.e. 1x PBST), then 200 μl / well of 1x PBST solution containing 5% skim milk powder was added and placed at 37°C for 2 hours to block. The plate was washed again, 100 μl / well of diluted antibody solution and PDL1-Benchmark (Tecentriq antibody, purchased from Roche) were added, incubated at 37° C. for 40 min, and then washed 4 times. Add biotin-labeled human PDL1 Fc protein solution (diluted 1:10000 in 1x PBST containing 2.5% skimmed milk ...

Embodiment 3

[0066] Example 3 DNA Cloning and Sequencing, Variable Region Sequencing of Anti-PDL1 Mouse Monoclonal Antibody

[0067] Total RNA was extracted from cultured mouse monoclonal cell lines using the FastPure Cell / Tissue Total RNA Isolation Kit (Vazyme, Cat#RC101). The process is briefly described as follows, centrifuged to collect 1.5x10 6Transfer the cells to a 1.5ml centrifuge tube and blot the supernatant dry. Add 500μl Buffer RL1 to the cell pellet and mix by pipetting. Transfer the treated cells to gDNA-Filter Columns (gDNA-Filter Columns have been placed in a collection tube), centrifuge at 13,000g for 2 minutes at room temperature, and save the supernatant in the collection tube. Add 1.6 times the volume of Buffer RL2 and mix gently. Transfer the mixture to RNAPureColumns, centrifuge at 13,000g for 1min at room temperature, and discard the waste. Add 500μl Buffer RW1 to RNAPure Columns, centrifuge at 13,000g for 1min at room temperature, and discard the waste. Add 700...

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PUM

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Abstract

The invention belongs to the field of biotechnology, relates to an anti-human PDL1 monoclonal antibody and its application, and provides a nucleic acid molecule encoding the antibody, an expression vector, a host cell, and a method for expressing the antibody. The invention can specifically recognize the monoclonal antibody of human PDL1, the binding affinity of the antibody and human PDL1 is stronger than that of the existing anti-human PDL1 monoclonal antibody, and the sequence is novel.

Description

technical field [0001] The invention belongs to the field of biotechnology and relates to a high-affinity and functional anti-human PDL1 monoclonal antibody or antibody fragment. The present invention also provides the encoding nucleic acid molecules of the antibody, expression vectors, host cells, and methods for expressing the antibody. Also provided are antibody immunoconjugates, bispecific molecules, chimeric antigen receptors, pharmaceutical compositions, and diagnostic and therapeutic methods comprising antibodies of the invention. Background technique [0002] Programmed death ligand 1 (PD-L1), also known as cluster of differentiation 274 (CD274) or B7 homolog 1 (B7-H1), is a 40 kDa type I transmembrane protein. PDL1 is the surface glycoprotein ligand of PD-1, a key immune checkpoint receptor expressed by activated T cells and B cells, and mediates immune suppression. PDL1 has been implicated in suppression of immune system responses during chronic infection, pregna...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/28C12N15/13A61K39/395A61P35/00A61K47/68A61K39/00
CPCC07K16/2827A61P35/00A61K47/68A61K39/001111C07K2317/565C07K2317/56C07K2317/92A61K2039/505
Inventor 陈明久马志清彭则羽
Owner BIOSION INC
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