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Common light chain mouse

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

Active Publication Date: 2014-12-24
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

[0190] Identification of Human Heavy Chain Variable Regions Binding to Selected Human Light Chain Variable Regions

[0191] An in vitro expression system was constructed to determine whether a single rearranged human germline light chain could be co-expressed with a human heavy chain from an antigen-specific human antibody.

[0192] Methods for producing human antibodies in genetically modified mice are known (see, e.g., US 6,596,541, Regeneron Pharmaceuticals, ). The technique involves producing a genetically modified mouse having a genome comprising human heavy and light chain variable regions operably linked to endogenous mouse constant region loci such that the mouse responds to antigenic stimulation to produce and mouse constant region antibodies. encoded 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 human variable regions and mouse cons...

Embodiment 2

[0204] Generation of rearranged human germline light chain loci

[0205] use technology to prepare various rearranged human germline light chain targeting vectors (see, e.g., 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 (BAC) clones 302g12 and 254m04 (Invitrogen). Using these two BAC clones, genomic constructs were engineered to preserve a single rearranged human germline light chain region and inserted into the endogenous kappa light chain locus, which was previously modified to delete the endogenous kappa can Mutation and connection of gene segments.

[0206] Construction of a rearranged human germline light chain targeting vector. Three different rearranged human germline light chain regions were prepared using standard known molecular biology techniques. The human variable gene ...

Embodiment 3

[0222] Generation of mice expressing a single rearranged human light chain

[0223] The above-mentioned target ES cells were used as donor ES cells and passed Methods into mouse embryos at the 8-cell stage (see for example, US Pat. 1): 91-99.). Individuals carrying the engineered human germline Vκ1-39Jκ5 light chain region, Vκ3-20Jκ1 light chain region, or VpreBJλ5 light chain region The presence of uniquely rearranged human germline light chain regions was detected by genotyping identification (Valenzuela et al., supra) using allelic modification analysis.

[0224] Pups were genotyped and pups heterozygous or homozygous for the rearranged human germline light chain region were selected to characterize expression of the rearranged human germline light chain region.

[0225] Flow Cytometry. Expression of rearranged human light chain regions in common light chain mice of the normal antibody population was verified by expression of immunoglobulins kappa and lambda in spleno...

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Abstract

A genetically modified mouse is provided, wherein the mouse expresses an immunoglobulin light chain repertoire characterized by a limited number of light chain variable domains. Mice are provided that express just one or a few immunoglobulin light chain variable domains from a limited repertoire in their germline. Methods for making light chain variable regions in mice, including human light chain variable regions, are provided. Methods for making human variable regions suitable for use in multispecific binding proteins, e.g., bispecific antibodies, are provided.

Description

technical field [0001] The present invention provides a genetically modified mouse expressing antibodies with a common human variable / mouse constant light chain associated with a different human variable / mouse constant heavy chain. The invention provides a method for preparing a human bispecific antibody from the human variable region gene sequence of mouse B cells. Background technique [0002] Antibodies typically comprise homodimeric heavy chain components, wherein each heavy chain monomer is associated with the same light chain. Antibodies with heterodimerization components (eg, bispecific antibodies) are desired as therapeutic antibodies. However, preparation with suitable light chain components that can satisfactorily associate with each heavy chain of a bispecific antibody has proven problematic. [0003] In one approach, light chains can be selected by surveying the statistics of variable region usage of all light chains, identifying the most commonly used light ch...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01K67/027C12N15/85C07K16/00
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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