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anti-cd3 humanized antibody

A humanized antibody, VH-CDR3 technology, applied in the field of biomedicine, can solve problems such as limited success, uncertain efficacy, and limitations

Active Publication Date: 2022-02-01
NOVOPROTEIN SCI INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Existing CD3-binding bispecific antibodies currently in clinical trials for cancer therapy are limited by short half-lives and / or variable efficacy
Although many attempts have been made to obtain anti-CD3 antibodies with particularly favorable properties, these attempts have so far had limited success

Method used

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Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0200] Preparation of antibodies

[0201] Any method suitable for producing monoclonal antibodies can be used to produce the anti-CD3 antibodies of the invention. For example, animals can be immunized with a linked or naturally occurring CD3 homodimer or fragment thereof. Appropriate methods of immunization can be used, including adjuvants, immunostimulants, repeated booster immunizations, and one or more routes can be used.

[0202] Any suitable form of CD3 can be used as an immunogen (antigen) for the production of non-human antibodies specific for CD3, and the antibodies are screened for biological activity. The priming immunogen can be full-length mature human CD3, including native homodimers, or peptides containing single / multiple epitopes. The immunogens can be used alone or in combination with one or more immunogenicity enhancers known in the art. Immunogens can be purified from natural sources, or produced in genetically modified cells. The DNA encoding the immunog...

Embodiment 1

[0243] Example 1 Production of Humanized CD3 Monoclonal Antibody

[0244] Balb / C mice were immunized with recombinant human CD3E protein (C00E, Novoprotein). After the immunization, B cells were taken to construct a phage display library, and candidate antibodies were obtained through screening. A series of humanized antibodies were obtained by structural simulation and rational design. The VH and VL of the humanized sequence are connected by a (G4S)3 linker, and a 6HIS tag is added to the C-terminus to construct a single-chain antibody (scFv), which is expressed in E. coli and purified by a nickel column to obtain a single-chain antibody protein. The obtained humanized antibody heavy chain variable region (VH), light chain variable region (VL) sequence and CDR, such as the obtained humanized antibody heavy chain variable region (VH) sequence are as follows:

[0245] VH-CDR1 of humanized CD3 antibody (SEQ ID NO.1)

[0246] GFTFNKYA

[0247] VH-CDR2 of humanized CD3 antibody ...

Embodiment 2

[0261] Example 2 Affinity detection of CD3 antibody

[0262] This example mainly describes the affinity detection of CD3 antibody in the form of single-chain antibody and recombinant human and monkey CD3E protein.

[0263] Affinity with human CD3 protein

[0264] The affinity of CD3 antibody and recombinant hCD3 protein (CP19, Novoprotein) was determined by ELISA, and the recombinant hCD3 was coated on the plate, and the CD3 antibody was diluted 10-fold gradient (starting from 20 μg / ml), see figure 1 .

[0265] The EC50 was calculated to be 0.3642 μg / ml.

[0266] Affinity with monkey CD3 protein

[0267] The affinity of CD3 antibody and recombinant monkey CD3 protein (CW07, Novoprotein) was measured by ELISA, and the recombinant monkey CD3 was coated on the plate, and the CD3 antibody was diluted 10-fold gradient (starting from 20 μg / ml), see figure 2 .

[0268] The EC50 was calculated to be 0.9381 μg / ml.

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Abstract

The invention provides a novel anti-CD3 humanized antibody. The anti-CD3 humanized antibody can specifically bind human and monkey CD3 proteins, and the constructed bispecific antibody has higher tumor killing activity.

Description

technical field [0001] The present invention relates to the field of biomedicine, in particular, the present invention relates to an anti-CD3 humanized antibody. Background technique [0002] Activation of T cells is critical for stimulating immune responses. T cells exhibit immune specificity and direct most cellular immune responses. Although T cells do not secrete antibodies, they are required for the secretion of antibodies by B lymphocytes. [0003] CD3 is a homodimeric or heterodimeric antigen expressed on T cells in association with the T cell receptor complex (TCR) and is required for T cell activation. Functional CD3 is a dimer formed from two of four different chains (ε, ζ, δ and γ). The CD3 dimer arrangement includes γ / ε, δ / ε and ζ / ζ. Antibodies to CD3 have been shown to aggregate CD3 on T cells, causing T cell activation in a manner similar to the involvement of peptide-loaded MHC molecules in TCR. Therefore, anti-CD3 antibodies have therapeutic purposes inv...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/28C07K19/00C12N5/10C12N15/13A61K39/00A61K47/68A61P35/00
CPCC07K16/2809C07K14/7051A61K39/001111A61P35/00A61K47/6803C07K2317/56C07K2317/565C07K2317/567C07K2317/31C07K2319/00
Inventor 朱化星宋作伟王米李德彬于福涛秦照峰房继旋
Owner NOVOPROTEIN SCI INC
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