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common light chain mouse

A mouse, light chain technology, applied in the field of genetically modified mice

Active Publication Date: 2017-03-08
REGENERON PHARM INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But preparing light chain components with suitable light chain components that can satisfactorily associate with each heavy chain of a bispecific antibody has proven problematic

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0192] Identification of the human heavy chain variable region combined with the selected human light chain variable region

[0193] An in vitro expression system was constructed to determine whether a single rearranged human germline light chain can be co-expressed with a human heavy chain derived from an antigen-specific human antibody.

[0194] Methods of producing human antibodies in genetically modified mice are known (see, for example, US 6,596,541, Regeneron Pharmaceuticals, ). The technology involves generating genetically modified mice having a genome that includes human heavy and light chain variable regions operably linked to endogenous mouse constant region loci, so that the mouse responds to antigen stimulation to produce human variable regions. And mouse constant region antibodies. Coded by The DNA of the variable regions of the heavy and light chains of antibodies produced in mice is fully human. Initially, high-affinity chimeric antibodies were isolated with hum...

Embodiment 2

[0206] Generation of rearranged human germline light chain loci

[0207] use Technology to prepare various rearranged human germline light chain targeting vectors (see, for example, US Pat. No. 6,586,251 and Valenzuela et al. (2003) High-throughput engineering of the mouse genome coupled with high-resolution expression analysis, Nature Biotech. 21(6):652-659) to modify the mouse genome bacterial artificial chromosome (Bacterial Artificial Chromosome, BAC) clones 302g12 and 254m04 (Invitrogen). Using these two BAC clones, the genomic construct was engineered to protect a single rearranged human germline light chain region and inserted into an endogenous kappa light chain locus, which was previously modified to delete the endogenous kappa Change and connect gene fragments.

[0208] Construction of a rearranged human germline light chain targeting vector. Three different rearranged human germline light chain regions were prepared using known standard molecular biology techniques. ...

Embodiment 3

[0224] Generation of mice expressing a single rearranged human light chain

[0225] The above target ES cells were used as donor ES cells and passed Method to introduce 8-cell stage mouse embryos (see, for example, US Pat. No. 7,294,754 and Poueymirou et al. (2007) F0 generationmice that are essentially fully derived from the donor gene-targeted ES cells allowing immediate phenotypic analyses Nature Biotech. 25(1) :91-99.). Carrying the engineered human germline Vκ1-39Jκ5 light chain region, Vκ3-20Jκ1 light chain region or VpreBJλ5 light chain region independently Allelic modification analysis was used to detect the presence of uniquely rearranged human germline light chain regions through genotyping (Valenzuela et al., supra).

[0226] The pups were genotyped and the pups heterozygous or homozygous for the rearranged human germline light chain region were selected to characterize the expression of the rearranged human germline light chain region.

[0227] Flow Cytometry. The exp...

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Abstract

The invention provides genetically modified mice, wherein the mice express an immunoglobulin light chain population characterized by a limited light chain variable region. The present invention provides mice expressing only one or a few immunoglobulin light chain variable regions from a limited population in their germline. The present invention provides methods for making light chain variable regions, including human light chain variable regions, in mice. The invention provides methods for preparing human variable regions suitable for use in multispecific binding proteins (eg, bispecific antibodies).

Description

Technical field [0001] The present invention provides a genetically modified mouse that expresses an antibody with a common human variable / mouse constant light chain associated with different human variable / mouse constant heavy chains. The present invention provides a method for preparing human bispecific antibodies from human variable region gene sequences of mouse B cells. Background technique [0002] Antibodies generally include homodimerized heavy chain components, where each heavy chain monomer is associated with the same light chain. Antibodies having heterodimerization components (e.g., bispecific antibodies) are desired as therapeutic antibodies. However, preparing suitable light chain components that can satisfactorily associate with each heavy chain of a bispecific antibody has proven to be problematic. [0003] In one method, the light chain can be selected by investigating the statistical data used by all light chain variable regions, identifying the most commonly us...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/85A01K67/027C07K16/00
CPCA01K67/0275A01K67/0278C07K16/00C07K16/22C12N15/8509C12N2800/107A01K2217/15A01K2217/072A01K2207/15A01K2227/105A01K2267/01C07K2317/567C07K2317/565C07K2317/56C07K2317/515C07K2317/24C07K2317/21C07K2319/30C07K2317/92C07K2317/76C12N2800/30C07K16/28C07K2317/14A01K67/02C12N15/85
Inventor R·巴布J·麦克沃特L·麦克唐纳S·史蒂文斯S·戴维斯D·R·巴克勒K·A·马尔A·J·莫菲
Owner REGENERON PHARM INC
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