Anti-human SIRP alpha monoclonal antibody and application thereof

A monoclonal antibody, antibody technology, applied in the direction of antibodies, anti-animal/human immunoglobulins, applications, etc., can solve the problems of limited specificity of anti-SIRPα antibodies, negative effects on T cell proliferation and recruitment, etc., and achieve novel sequences. Effect

Active Publication Date: 2020-12-01
BIOSION INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Binding to SIRPγ expressed on T cells may negatively affect T cell prolifera

Method used

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  • Anti-human SIRP alpha monoclonal antibody and application thereof
  • Anti-human SIRP alpha monoclonal antibody and application thereof
  • Anti-human SIRP alpha monoclonal antibody and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

[0041] Example 1 Obtaining specific anti-SIRPα mouse monoclonal antibody by fusion hybridoma technology

[0042] 1.1 Animal Immunization

[0043] Mice were immunized according to general methods 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C端含有人IgG1 Fc标签的重组人SIRPα(氨基酸Met 1-Arg 370)蛋白(公司自产,所用质粒为CD172-ECD-Fc;载体为pCP;插入氨基酸序列为MEPAGPAPGRLGPLLCLLLAASCAWSGVAGEEELQVIQPDKSVLVAAGETATL RCTATSLIPVGPIQWFRGAGPGRELIYNQKEGHFPRVTTVSDLTKRNNMDFSIR IGNITPADAGTYYCVKFRKGSPDDVEFKSGAGTELSVRAKPSAPVVSGPAARA TPQHTVSFTCESHGFSPRDITLKWFKNGNELSDFQTNVDPVGESVSYSIHSTA KVVLTREDVHSQVICEVAHVTLQGDPLRGTANLSETIRVPPTLEVTQQPVRAE NQVNVTCQVRKFYPQRLQLTWLENGNVSRTETASTVTENKDGTYNWMSWLLVNVSAHRDDVKLTCQVEHDGQPAVSKSHDLKVSAHPKEQGSNTAAENTGSNER). Use recombinant human SIRPα with his-tagged protein (produced by the company, the plasmid used is SRPA-ECD-His; the vector is pCMV-C-His; the amino acid sequence inserted between EcoR I and Xba...

Example Embodiment

[0051] Example 2 In Vitro Assay for Determining the Functional Activity of SIRPα Monoclonal Antibodies

[0052] 2.1 Determination of antibody binding capacity based on capture ELISA

[0053] Goat anti-mouse IgG F(ab') in 1xPBS 2 A specific secondary antibody (Jackson ImmunoResearch Laboratories, Inc., Cat#115-005-072) was added to a final concentration of 2 μg / ml, 100 μl / well was added to a 96-well microtiter plate, and coated for 2 hours at 37 degrees Celsius. After the coating, the plate was washed once with 1×PBST, and then 200 μl / well of PBST solution containing 5% nonfat dry milk was added and placed at 37 degrees Celsius to block for 2 hours. Wash the plate again, add 100 μl / well of diluted antibody solution and Benchmark (ie KWAR23 antibody, produced by the company, its heavy chain amino acid is

[0054] EVQLQQSGAELVKPGASVKLSCTASGFNIKDYYIHWVQQRTEQGLEWIG RIDPEDGETKYAPKFQDKATITADTSSNTAYLHLSSLTSEDTAVYYCARWGAY WGQGTLVTVSAASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSW NSGALTSGV...

Example Embodiment

[0076] Example 3 Binding activity of anti-SIRPα mouse monoclonal antibody

[0077] According to the assay method described in Example 2, the binding activity of the SIRPα antibody was assessed and summarized in Table 3 below.

[0078] The SIRPαBenchmark was used as a control. It can be seen from Table 3 that the anti-human SIRPα monoclonal antibody of the present invention has slightly better binding activity to human SIRPα antigen than Benchmark, and can cross-react with SIRPβ antigen at the same time.

[0079] Table 3 Binding activity of SIRPα antibody

[0080]

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Abstract

The invention belongs to the technical field of biology, relates to an anti-human SIRP alpha monoclonal antibody and application thereof, and also provides coding nucleic acid molecules, an expressionvector and a host cell of the antibody, and a method for expressing the antibody. The invention also provides an antibody immunoconjugate containing the antibody, bispecific molecules, a chimeric antigen receptor, a pharmaceutical composition, and a diagnosis and treatment method. According to the monoclonal antibody capable of specifically recognizing human SIRP alpha, the affinity of combiningthe antibody with the human SIRP alpha is higher than that of existing anti-human SIRP alpha monoclonal antibodies, 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 SIRPα 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] Signal regulatory protein α (Signal regulatory protein α), also known as SHPS-1 or SIRPα, is a typical inhibitory immune receptor in the SIRP family, mainly expressed in myeloid cells such as macrophages and dendritic cells, and the surface of nerve cell membranes, while other cells in the body express less (Kharitonenkov, A. et al. (1997). "A family of proteins that inhibit signaling through tyrosine kinase r...

Claims

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Application Information

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IPC IPC(8): C07K16/28C12N15/13A61K39/395A61P35/00
CPCC07K16/2896A61P35/00C07K2317/51C07K2317/515C07K2317/56C07K2317/565C07K2317/33C07K2317/76C07K2317/92A61K2039/505
Inventor 陈明久
Owner BIOSION INC
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