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Bispecific antibodies and antibody conjugates for tumor therapy and applications thereof

A bispecific antibody and antibody conjugate technology, which is applied in the field of tumor treatment, can solve the problems of immunogenicity manufacturing difficulties, poor pharmacokinetics and physical properties of bispecific antibodies, and achieve high solubility and antigenicity Weak, good thermal stability effect

Active Publication Date: 2021-11-12
BENHEALTH BIOPHARMACEUTIC SHENZHEN CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] In addition, bispecific antibodies may have poor pharmacokinetic and physical properties (such as immunogenicity) and manufacturing difficulties

Method used

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  • Bispecific antibodies and antibody conjugates for tumor therapy and applications thereof
  • Bispecific antibodies and antibody conjugates for tumor therapy and applications thereof
  • Bispecific antibodies and antibody conjugates for tumor therapy and applications thereof

Examples

Experimental program
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Effect test

preparation example Construction

[0077] Preparation of conjugates of the present invention

[0078] The preparation method of the bispecific antibody conjugate of the present invention is described below. The preparation method comprises the following steps:

[0079] (1) Preparation, collection and activation of nanomaterials;

[0080] (2) Linking the nanomaterial obtained in step (1) with the bispecific antibody.

[0081] In step (1), the preparation of the nanomaterials includes: using a solvent to completely dissolve the nanomaterials, stirring, and adding water to form a uniform emulsion. Wherein said stirring can be carried out under the rotating speed of 500-20000rpm / min, for example rotating speed can be 500rpm / min, 700rpm / min, 800rpm / min, 1000rpm / min, 1100rpm / min, 1200rpm / min, 1300rpm / min, 1400rpm / min min, 1480rpm / min, 1500rpm / min, 2000rpm / min, 2200rpm / min, 2500rpm / min, 3000rpm / min, 3500rpm / min, 4000rpm / min, 4200rpm / min, 4500rpm / min, 5000rpm / min, 5500rpm / min, 6000rpm / min, 6500rpm / min, 7000rpm / min,...

Embodiment 1

[0117] Embodiment 1-design and synthesis of anti-CD16-MUC1 bispecific nanobody nucleotide sequence

[0118] According to the sequence and connection form of the bispecific antibody, the nucleotide sequence was redesigned and optimized, and an NcoI restriction site was added at the 5' end of the sequence, and a HindIII restriction site was added at its 3' end. DsbA-anti-MUClVHH-GS-anti-CD16 VHH-6His was directly synthesized and ligated into the vector pETDuet by double enzyme digestion to form an expression plasmid. The schematic diagram of the structure of the anti-CD16-MUC1 bispecific nanobody is shown infigure 1 shown.

Embodiment 2

[0119] Embodiment 2-vector transformation BL21 (DE3)

[0120] Transform the plasmid into Escherichia coli DH5α competent cell strain, select positive clones, culture overnight at 37°C in LB medium (3mL) containing 100 μg / ml ampicillin, centrifuge the bacterial solution at 5000rpm at room temperature and discard the supernatant, and the obtained Escherichia coli was purified by Qiagen The company's plasmid extraction kit lyses and extracts the plasmid to obtain an expression vector.

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Abstract

The present invention provides a bispecific antibody, which comprises an antibody fragment against MUCl VHH and an antibody fragment against CD16 VHH. The antibody fragments used in the present invention are variable region sequences from camelid heavy chain antibodies, and have high binding affinity with antigens. In the present invention, antibody fragments that recognize MUC1 and CD16 are constructed in the same antibody molecule, which can specifically bind to MUC1 and CD16 molecules, promote the killing effect of NK cells on MUC1-positive cells, and inhibit the growth of MUC1-positive tumors . The present invention also provides conjugates of the bispecific antibody, related pharmaceutical compositions and applications.

Description

technical field [0001] The present invention relates to a bispecific antibody and its preparation, more specifically, to an anti-MUC1 and anti-CD16 bispecific antibody for use in the field of tumor treatment. Background technique [0002] In tumor immunotherapy, bispecific antibody (bispecific antibody, bsAb) plays a very important role. Bispecific antibodies are artificial antibodies containing two specific antigen-binding sites. Through the binding sites, they can simultaneously bind to specific antigens on the surface of effector cells and target cells, activate quiescent effector cells and recruit them to the surrounding target cells. induce apoptosis or lysis of target cells. A variety of bsAbs targeting different immune effector cells and tumor cells have been developed, among which the main immune effector cells in tumor immunotherapy are T cells, NK cells and macrophages. NK cells and T cells are the most studied effector cells. [0003] Molecules that have trigge...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/46C07K16/28C12N15/13C12N15/85A61K39/395A61P35/00C07K17/08
CPCC07K17/08A61P35/00C07K16/3092C07K16/283C07K2317/31C07K2317/569C07K2317/73A61K47/6937A61K47/593A61K47/65A61K47/6883A61K35/17C07K16/46C12N15/85
Inventor 于浩洋王忠李正成
Owner BENHEALTH BIOPHARMACEUTIC SHENZHEN CO LTD
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