Monoclonal antibody specifically binding to human CD47 and application thereof

A monoclonal antibody and multispecific antibody technology, applied in the fields of antibody, application, anti-animal/human immunoglobulin, etc., can solve the problems of specific antibody to be studied, rapid antibody consumption, low blood drug concentration, etc., to achieve Good anti-tumor efficacy, good anti-tumor efficacy

Pending Publication Date: 2022-05-27
MABWELL (SHANGHAI) BIOSCIENCE CO LTD
View PDF23 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] Due to the large expression of CD47 on the surface of red blood cells, platelets and other cells, the clinical application of this target antibody faces two problems: 1. The toxicity of red blood cells and platelets can be restored by adjusting the dosage regimen and the body's own compensatory function. Candidate molecules with too high affinity still face greater risks; 2. Due to the excessive consumptio

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
  • Monoclonal antibody specifically binding to human CD47 and application thereof
  • Monoclonal antibody specifically binding to human CD47 and application thereof
  • Monoclonal antibody specifically binding to human CD47 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0075] Example 1: Sources of Antigen and Control Antibodies

[0076] Using the extracellular region (19-136aa) of human CD47 as the target antigen, we constructed the transient samples of unlabeled CD47 (crystallization experiment), CD47-his, CD47-mutC15G-his, CD47-mFc, CD47-hFc and SIRPα-mFc respectively. Expression vectors to obtain various forms of recombinant antigens and related identification proteins. A variety of human tumor cell lines that highly express CD47, such as CCRF-SB, U937, SKOV-3, A431, etc., are used for in vitro binding, blocking and in vivo pharmacodynamic evaluation.

[0077] Referring to the sequence disclosed in the patent, the following antibodies were prepared as reference antibodies, all in the form of IgG4. FortySeven's clinical research antibody hu5F9 (US2015 / 0183874A1, hu5F9V2, light and heavy chain variable region sequences: SEQ ID NO.1 and SEQ ID NO.2), Surface Oncology's SRF-231 (WO2017 / 053423A1, 2.3D11, light and heavy chain can be Variable...

Embodiment 2

[0091] Example 2: Anti-human CD47 antibody hybridoma antibody and chimeric antibody

[0092] Hybridoma production was performed by immunizing Balb / c mice with CD47-mFc. Serum antibody titers were detected 34 days or 57 days after the primary immunization, respectively. Select mice whose serum titer meets the requirements of the fusion experiment (serum antibody titer: 1:102400). Splenocytes and myeloma cells FO in an appropriate ratio (5:1) are fused under the action of a fusion agent to prepare hybridomas Monoclonal cells. Hybridoma cells were cultured in selective medium R1640-HAT, and 651 hybridoma supernatant samples were detected by ELISA on days 10-14, including cell binding activity (FACS), CD47 / SIPRα blocking, As well as the cross-reaction with recombinant cyno-CD47-mFc, clones 597 and 439 were screened out. The above cloned light and heavy chain variable region genes were obtained, and the light and heavy chain variable region genes were cloned into a recombinant e...

Embodiment 3

[0105] Example 3. Humanization and Transformation

[0106] 3.1 Affinity detection of humanized 439

[0107] The humanization design of 439 was carried out, and IGKV3-7|IGKJ4 and IGHV1-3*01|IGHJ1*01 were selected as germline templates for the light and heavy chains of 439, respectively. See SEQ ID NO.13 and SEQ ID NO.14 for the light and heavy chain sequences of Hz439 after humanization.

[0108] SEQ ID NO.13:Hz439 light chain amino acid sequence

[0109] ENVLTQSPPTLSLSPGERATLSCRASSSVSSSYFHWYQQKPGQAPRLLIYGTSTLASGIPARFSGSGSGTDYTLTISSLQPEDAAVYYCQQYSGYPYTFGGGTKVEIKR

[0110] SEQ ID NO.14:Hz439 heavy chain amino acid sequence

[0111] QVQLVQSGAEVKKPGASVKVSCKASGYTFSRYWIEWVRQAPGQRLEWMGEILPGDVQTSYNEKFKGRVTITADTSASTAYMELSSLRSEDTAVYYCARGGLRRFDYWGQGTLVTVSS

[0112] The affinity of 439 with recombinant CD47-His after humanization was detected, and the dissociation was carried out after binding for 300s. The results are as follows figure 2 , as shown in Table 2, the affinity of t...

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 provides a monoclonal antibody specifically binding to human CD47 or a fragment thereof, which can bind to CD47 on the surface of a cell and block binding of SIRP alpha to CD47 on the surface of the cell. The affinity KD value on the recombinant human CD47 is between 1 * 10 <-10 > and 8 * 10 <-8 >, and the tumor inhibition activity and the adverse reaction activity can be balanced. In the aspect of tumor inhibition activity, the antibody can inhibit tumor growth in a CD47/SIRP alpha double-transgenic mouse tumor-bearing hCD47-MC38 subcutaneous transplanted tumor model, and can enhance in-vivo cell phagocytosis and prolong the lifetime of a mouse in a mouse leukemia model of transplanted human acute B lymphocytic leukemia; in the aspect of adverse reaction activity, the antibody does not have or only has reduced erythrocyte agglutination activity, and does not have a significant effect or only has an over-sexual effect on erythrocytes, platelets and hemoglobin.

Description

[0001] priority information [0002] This application claims priority to and the benefit of the patent application No. 202011253602.X filed with the State Intellectual Property Office of China on November 11, 2020, which is hereby incorporated by reference in its entirety. technical field [0003] The invention belongs to the field of antibody engineering, and in particular relates to a monoclonal antibody that specifically binds to human CD47, its fragment, preparation method and use, in particular to a monoclonal antibody that can block the binding of SIRPα and CD47 and has low erythrocyte toxicity The anti-human CD47 monoclonal antibody and its application. Background technique [0004] CD47, also known as integrin-associated protein (IAP), ovarian cancer marker (OV-3), is a member of the immunoglobulin superfamily and is a 5-time transmembrane glycoprotein of about 50KD, containing three domains : The extracellular amino terminus is an IgV-like domain containing 5 glyco...

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
IPC IPC(8): C07K16/28C07K16/46C12N15/13A61K39/395A61P35/00A61P35/02
CPCC07K16/2803A61P35/00A61P35/02C07K2317/56C07K2317/24C07K2317/31C07K2317/73C07K2317/92A61K2039/505
Inventor 王双焦莎莎王荣娟张畅曾大地张姣
Owner MABWELL (SHANGHAI) BIOSCIENCE CO LTD
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