Mice that make VL binding proteins

Patent metadata

Field	Value
Publication number	US-9516868-B2
Application number	US-201113195951-A
Country	US
Kind code	B2
Filing date	Aug 2, 2011
Priority date	Aug 2, 2010
Publication date	Dec 13, 2016
Grant date	Dec 13, 2016

Abstract

Genetically modified mice and methods for making an using them are provided, wherein the mice comprise a replacement of all or substantially all immunoglobulin heavy chain V gene segments, D gene segments, and J gene segments with at least one light chain V gene segment and at least one light chain J gene segment. Mice that make binding proteins that comprise a light chain variable domain operably linked to a heavy chain constant region are provided. Binding proteins that contain an immunoglobulin light chain variable domain, including a somatically hypermutated light chain variable domain, fused with a heavy chain constant region, are provided. Modified cells, embryos, and mice that encode sequences for making the binding proteins are provided.

First claim

We claim:

1. A mouse whose germline genome comprises a modified endogenous mouse immunoglobulin heavy chain locus comprising a replacement of all functional endogenous mouse immunoglobulin heavy chain variable (V H ) gene segments, all functional endogenous mouse immunoglobulin heavy chain diversity (D H ) gene segments and all functional endogenous mouse immunoglobulin heavy chain joining (J H ) gene segments at the endogenous mouse immunoglobulin heavy chain locus with a nucleotide sequence that comprises a plurality of contiguous unrearranged functional human immunoglobulin light chain variable V κ (hV κ ) gene segments and all five unrearranged functional human immunoglobulin light chain joining gene segments (hJ κ 1-hJ κ 5),
wherein the plurality of unrearranged functional hV κ gene segments and the five unrearranged functional human immunoglobulin light chain joining gene segments (hJ κ 1-hJ κ 5) are operably linked to an intact endogenous mouse immunoglobulin heavy chain constant region at the endogenous mouse immunoglobulin heavy chain locus,
wherein the plurality of unrearranged functional hV κ gene segments and the five unrearranged functional human immunoglobulin light chain joining gene segments (hJ κ 1-hJ κ 5) rearrange in a B cell during B cell development to form a rearranged human immunoglobulin light chain variable region V κ /J κ nucleotide sequence operably linked to the endogenous mouse immunoglobulin heavy chain constant region at the endogenous mouse immunoglobulin heavy chain locus, and
wherein the mouse comprises a CD19 + B cell comprising the rearranged human immunoglobulin light chain variable region V κ /J κ nucleotide sequence operably linked to the endogenous mouse immunoglobulin heavy chain constant region at the endogenous mouse immunoglobulin heavy chain locus.

2. The mouse of claim 1 , wherein all the functional endogenous mouse V H , D H , and J H gene segments are replaced with at least 6 human V κ gene segments and the five unrearranged functional human immunoglobulin light chain joining gene segments (hJ κ 1-hJ κ 5).

3. The mouse of claim 1 , wherein all the functional endogenous mouse V H , D H , and J H gene segments are replaced with at least 16 human V κ gene segments and the five contiguous unrearranged human immunoglobulin light chain joining gene segments (hJ κ 1-hJ κ 5).

4. The mouse of claim 1 , wherein all the functional endogenous mouse V H , D H , and J H gene segments are replaced with at least 30 human V κ gene segments and the five contiguous unrearranged human immunoglobulin light chain joining gene segments (hJ κ 1-hJ κ 5).

5. The mouse of claim 1 , wherein all the functional endogenous mouse V H , D H , and J H gene segments are replaced with at least 40 human V κ gene segments and the five contiguous unrearranged human immunoglobulin light chain joining gene segments (hJ κ 1-hJ κ 5).

6. An isolated cell from the mouse of claim 1 , wherein the cell comprises a modified endogenous mouse immunoglobulin heavy chain locus comprising a replacement of all functional endogenous mouse immunoglobulin heavy chain variable (V H ) gene segments, all functional endogenous mouse immunoglobulin heavy chain diversity (D H ) gene segments and all functional endogenous mouse immunoglobulin heavy chain joining (J H ) gene segments at the endogenous mouse immunoglobulin heavy chain locus with a nucleotide sequence that comprises a plurality of unrearranged functional human immunoglobulin light chain variable Vκ (hV κ ) gene segments and all five unrearranged functional human immunoglobulin light chain joining gene segments (hJ κ 1-hJ κ 5),
wherein the plurality of unrearranged functional hV κ gene segments and all five unrearranged functional human immunoglobulin light chain joining gene segments (hJ κ 1-hJ κ 5) are operably linked to an intact endogenous mouse immunoglobulin heavy chain constant region at the endogenous mouse immunoglobulin heavy chain locus,
wherein the plurality of unrearranged functional hV κ gene segments and the five unrearranged functional human immunoglobulin light chain joining gene segments (hJ κ 1-hJ κ 5) are capable of rearranging in a B cell during B cell development to form a rearranged human immunoglobulin light chain variable region V κ /J κ nucleotide sequence operably linked to the endogenous mouse immunoglobulin heavy chain constant region at the endogenous mouse immunoglobulin heavy chain locus.

7. The isolated cell of claim 6 , wherein the cell is an embryonic stem cell.

8. A cell isolated from the mouse of claim 1 , wherein the cell is the CD19 + B cell.

9. A hybridoma comprising a myeloma cell line fused with the CD19 + B cell of claim 8 , wherein the hybridoma produces a polypeptide encoded by the rearranged human immunoglobulin light chain variable region V κ /J κ nucleotide sequence operably linked to the endogenous mouse immunoglobulin heavy chain constant region.

10. The mouse of claim 1 , wherein the rearranged human immunoglobulin light chain variable region V κ /J κ gene sequence comprises at least one N addition.

11. The mouse of claim 1 , wherein the mouse is homozygous or heterozygous for the modified endogenous immunoglobulin heavy chain locus.

Assignees

• Macdonald Lynn
• Stevens Sean
• Gurer Cagan
• Hosiawa Karolina A
• Murphy Andrew J
• Regeneron Pharma

Inventors

• Macdonald Lynn
• Stevens Sean
• Gurer Cagan
• Hosiawa Karolina A
• Murphy Andrew J

Classifications

• A01K2217/072

  maintaining or altering function, i.e. knock in · CPC title

• C12N15/8509

  for producing genetically modified animals, e.g. transgenic · CPC title

• A01K67/0275 Primary

  Genetically modified vertebrates, e.g. transgenic · CPC title

• A01K2267/01

  Animal expressing industrially exogenous proteins · CPC title

• C07K16/00 Primary

  Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies · CPC title