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Bispecific antibody

A bispecific antibody and bispecific antibody technology, applied in the direction of antibodies, specific peptides, anti-tumor drugs, etc., can solve problems such as tumor immune escape

Active Publication Date: 2019-12-20
BEIJING SHENOGEN PHARMA GRP
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Such a process is originally the normal immune regulation of the body to avoid damage or autoimmune diseases caused by excessive immunity, but in the process of tumor development, such a process will lead to immune escape or tolerance of the tumor

Method used

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Examples

Experimental program
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Effect test

Embodiment 1

[0059] Expression and purification of bispecific antibodies

[0060] (1) Synthesize the nucleotide sequence shown in SEQ ID NO: 10 and determine the correctness of the sequence. (2) Cloning the synthesized sequence into a mammalian expression vector in a manner well known to those skilled in the art. (3) Transfect the expression vector into the host cell CHO cell, and culture it for a suitable time. (4) After the set culture time, harvest the cell culture supernatant. (5) Purify the bispecific antibody by affinity chromatography with Ni strain. (6) Further purifying the bispecific antibody with anions and cations. (7) Determine the purity, concentration and charge heterogeneity of the bispecific antibody after purification by SDS-PAGE, SEC, iCE and other methods.

Embodiment 2

[0062] Using SPR to detect the affinity of bispecific antibodies to CD3 and PD-L1 proteins

[0063] (1) Prepare or purchase PD-L1 protein and CD3 protein. (2) Fix the PD-L1 or CD3 protein on the chip according to the instructions of the SPR manufacturer. (3) Prepare 4 to 5 different concentrations of the purified bispecific antibody solution. (4) Flow the bispecific antibody solution at a suitable speed through the chip immobilized with PD-L1 or CD3 protein and record the SPR reading. Calculate the affinity of the bispecific antibody to PD-L1 and CD3 proteins based on the reading and concentration.

Embodiment 3

[0065] Affinity determination of bispecific antibodies to cells expressing CD3 and PD-L1

[0066] (1) Preparation and culture of CD3-expressing cells, such as cytokine-induced human T cells or CHO cells expressing human PD-L1; (2) The purified bispecific antibodies of different concentrations are combined with a certain amount of CD3-positive cells or PD-L1 positive CHO cells are co-cultured; (3) Add fluorescently labeled anti-His-Tag antibody and culture; (4) Wash off antibodies and secondary antibodies that are not bound to the cells; (5) Use flow cytometry to determine The amount of the bispecific antibody bound to the cell surface; (5) Calculate the affinity of the bispecific antibody to CD3 positive cells and PD-L1 positive cells.

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Abstract

The invention provides a method. The core of the method is an anti-human PD-L1 antibody, and the anti-human PD-L1 antibody and an anti-human CD3 antibody constitute the bispecific antibody. The bispecific antibody can bind T cells and express PD-L1 or inductively express PD-L1 tumor cells and kill bond tumor cells. The method can bypass limitation of TCR and MHC-I antigens and also solve immunosuppression caused by binding of PD-L1 with PD-1 on surfaces of T cells after inducted expression. Therefore, tumor cells can be more effectively killed and growth of tumor can be inhibited.

Description

[0001] Related application [0002] This application claims the priority of the Chinese patent application 201811082753.6 whose application date is September 17, 2018 and the invention title is "a bispecific antibody". [0003] Invention field [0004] The present invention belongs to the field of biomedicine, and specifically relates to a bispecific antibody for treating tumors expressing PD-L1 or inducible expressing PD-L1. [0005] Background of the invention [0006] The immune system of higher animals is precisely controlled by the body itself to maintain a stable and balanced state. In this state, the immune system can not only quickly respond to and eliminate the abnormalities that occur in foreign invaders or the body, but also weaken the immune response and maintain after the abnormalities have been eliminated. Resting state. There are many factors involved in the regulation and maintenance of immune status, including some immune stimulating factors, such as CD40, CD137, OX40,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/46C12N15/63A61K39/395A61P35/00
CPCA61K2039/505A61P35/00C07K16/2809C07K16/2827C07K2317/31C07K2317/73C07K2317/76C07K2317/92
Inventor 孟坤周骏马
Owner BEIJING SHENOGEN PHARMA GRP
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