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Novel antibody molecule aiming at human CLDN18.2, antigen binding fragment and medical application thereof

A technology for binding fragments and antibody molecules, applied in the direction of anti-receptor/cell surface antigen/cell surface determinant immunoglobulin, medical preparations containing active ingredients, antibodies, etc., can solve the problems of immunogenicity risks and other problems

Active Publication Date: 2019-12-24
L&L BIOPHARMA CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

IMAB362 is an antibody against human CLDN18.2, which is a chimeric antibody of mouse origin, and there is a risk of immunogenicity

Method used

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  • Novel antibody molecule aiming at human CLDN18.2, antigen binding fragment and medical application thereof
  • Novel antibody molecule aiming at human CLDN18.2, antigen binding fragment and medical application thereof
  • Novel antibody molecule aiming at human CLDN18.2, antigen binding fragment and medical application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0171] Example 1: Construction of high-expression cell lines of claudin (Claudin, or CLDN) 18.1, 18.2 (English abbreviation: CLDN18.1, CLDN18.2)

[0172] The human CLDN18.1, human CLDN18.2, mouse CLDN18.1, and mouse CLDN18.2 high-expression cell lines used in the present invention are completed through the company's stable cell line construction platform. Specific steps are as follows:

[0173] On the first day of the experiment, 293T cells (Cat#GNHu17, a cell bank of the Type Culture Collection Committee of the Chinese Academy of Sciences) were inoculated in two 6 cm culture dishes, and the number of cells in each culture dish reached 7.5×10 5 . On the second day, wrap the plasmid (pGag-pol, pVSV-G, pBabe and other BioVector plasmid vector strain cell gene collection center) and the plasmid pBabe-CLDN18.2 or pBabe- Add 4µg each of CLDN18.1 to OPTI-MEM (Thermofisher Scientific Cat#31985070) to make the final volume 200µl, prepare 200µl OPTI-MEM and add 36µl transfection reag...

Embodiment 2

[0187] Example 2: Anti-CLDN18.2 antibody binding to CLDN18.2 + and CLDN18.1 + cell lines (ELISA) experiment

[0188] After expanding the human CLDN18.1, human CLDN18.2, mouse CLDN18.1, or mouse CLDN18.2 high-expression monoclonal cell line obtained in Example 1, press 5x10 4 Spread a 96-well plate per well, incubate overnight in a 37°C incubator, remove the supernatant, and fix with 100 μl / well of immunostaining fixative (Shanghai Beyond Biotechnology Co., Ltd. Cat#P0098) for half an hour at room temperature. After washing once with PBS (Yuan Pei Biology, Cat# B320), block with 5% milk at 37°C for 2 hours, and wash 3 times with PBST. Add sample to be tested (human or mouse antibody, Jackson Immuno Research). Incubate at 37°C for 1 hour, then wash 3 times with PBST. Add Anti-human or mouse HRP 1:2500 50μl / well, incubate at 37°C for 1 hour, wash 3 times with PBST, develop color with TMB (SurmodicCat#TTMB-1000-01), add 50μl / well 1MH 2 SO 4 Stop the reaction. Microplate reade...

Embodiment 3

[0189] Example 3: Cloning, expression and purification of recombinant proteins and antibodies

[0190] The cloning, expression and purification of the recombinant protein / antibody used in the present invention are carried out according to molecular cloning methods well known to those skilled in the art.

[0191] Specifically, the expression vector used in the present invention was purchased from Changsha Youbao Biotechnology Co., Ltd., and the EcoRI restriction site (GAATTC) was introduced by Shanghai Jianxin Biomedical Technology Co., Ltd. (Jianxin Biotechnology) to facilitate double enzyme digestion. Or homologous recombination method to clone foreign genes. Gene synthesis is completed by CRO (outsourcing) companies such as Sangon Bioengineering (Shanghai) Co., Ltd. (Sangon Biotechnology). 293 cells, CHO-K were purchased from the Cell Bank of the Type Culture Collection Committee of the Chinese Academy of Sciences.

[0192] The recombinant protein and antibody in the prese...

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Abstract

The invention discloses an anti-human CLDN18.2 antibody as well as an antigen binding fragment and a medical application thereof. Specifically, the present invention relates to a murine antibody comprising a CDR region of the anti-human CLDN18.2 antibody, a chimeric antibody and a humanized antibody, a fully humanized antibody, an antibody-dependent cytotoxicity (ADCC) comprising the antibody andthe antigen binding fragment thereof, complement dependent cytotoxicity (CDC), the killing effect on CLDN18.2 positive cells, the effect of inhibiting the growth of CLDN18.2 positive tumors, the in-vivo drug effect, a medicine containing the anti-human CLDN18.2 antibody and the antigen binding fragment thereof, a composition of the medicine thereof, and an application of the medicine in tumor treatment, in particular to treatment of CLDN18.2 positive tumor patients..

Description

technical field [0001] The present invention relates to a human CLDN18.2 antibody, a CLDN18.2 antigen-binding fragment, a murine antibody comprising the CDR region of the CLDN18.2 antibody, a chimeric antibody composed of a murine antibody variable region and a human antibody constant region, a human Antibody-dependent cytotoxicity (ADCC), and complement-dependent cytotoxicity (CDC) effects of humanized antibodies, fully humanized antibodies, and antigen-binding fragments thereof. The drug combination comprising the antibody and its antigen-binding fragment, its killing effect on tumor cells, especially CLDN18.2 high expression (CLDN18.2 positive) tumor cells, its pharmacodynamic activity in animals, and its single or drug combination The drug is used as an anticancer drug, especially for the treatment of patients with CLDN18.2 overexpression tumors. Background technique [0002] Cancer is a great threat to human health and one of the important causes of death in the field ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/28A61K39/395A61P35/00A61P35/02
CPCC07K16/28A61K35/00A61K35/02C07K2317/565C07K2317/24C07K2317/734A61K2039/505C07K2317/92C07K2317/732
Inventor 刘佳建
Owner L&L BIOPHARMA CO LTD
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