Hybrid light chain mice

• US 9,035,128 B2
• Filed: 06/22/2011
• Issued: 05/19/2015
• Est. Priority Date: 06/22/2010
• Status: Active Grant

• Current Assignee: Regeneron Pharmaceuticals Incorporated
• Original Assignee: Regeneron Pharmaceuticals Incorporated
• Inventors: MacDonald, Lynn, Stevens, Sean, Gurer, Cagan, Murphy, Andrew J., Hosiawa, Karolina A.
• Primary Examiner(s): Wehbe, Anne Marie S
• Application Number:
  US13/166,200
• Publication Number:
  US 20120073004A1
• Time in Patent Office: 1,427 Days
• US Class Current: 800/18
• CPC Class Codes: A01K 2207/15 Humanized animals
• CPC Class Codes: A01K 2217/05 Animals comprising random inserted nucleic acids (transgenic)
• CPC Class Codes: A01K 2217/052 inducing gain of function
• CPC Class Codes: A01K 2217/072 maintaining or altering function, i.e. knock in
• CPC Class Codes: A01K 2217/15 Animals comprising multiple alterations of the genome, by transgenesis or homologous recombination, e.g. obtained by cross-breeding
• CPC Class Codes: A01K 2227/105 Murine
• CPC Class Codes: A01K 2267/01 Animal expressing industrially exogenous proteins
• CPC Class Codes: A01K 2267/02 Animal zootechnically ameliorated
• CPC Class Codes: A01K 2267/0381 Animal model for diseases of the hematopoietic system
• CPC Class Codes: A01K 67/0275 Genetically modified vertebrates, e.g. transgenic
• CPC Class Codes: A01K 67/0278 Knock-in vertebrates, e.g. humanised vertebrates
• CPC Class Codes: C07K 16/00 Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies antibodies with enzymatic activity, e.g. abzymes C12N9/0002
• CPC Class Codes: C07K 16/18 against material from animals or humans
• CPC Class Codes: C07K 16/461 Igs containing Ig-regions, -domains or -residues form different species
• CPC Class Codes: C07K 16/462 Igs containing a variable region (Fv) from one specie and a constant region (Fc) from another
• CPC Class Codes: C07K 2317/10 characterized by their source of isolation or production
• CPC Class Codes: C07K 2317/14 Specific host cells or culture conditions, e.g. components, pH or temperature
• CPC Class Codes: C07K 2317/21 from primates, e.g. man
• CPC Class Codes: C07K 2317/24 containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
• CPC Class Codes: C07K 2317/50 characterized by immunoglobulin fragments
• CPC Class Codes: C07K 2317/515 : Complete light chain, i.e. VL + CL
• CPC Class Codes: C07K 2317/56 : variable (Fv) region, i.e. VH and/or VL
• CPC Class Codes: C07K 2317/64 : comprising a combination of variable region and constant region components
• CPC Class Codes: C12N 15/10 : Processes for the isolation, preparation or purification of DNA or RNA chemical preparation of DNA or RNA C07H21/00; preparation of non-structural polynucleotides from microorganisms or with enzymes C12P19/34
• CPC Class Codes: C12N 15/8509 : for producing genetically modified animals, e.g. transgenic
• CPC Class Codes: C12N 5/0606 : Pluripotent embryonic cells, e.g. embryonic stem cells [ES] embryonic germ cells C12N5/0611, induced pluripotent stem cells C12N5/0696

Abstract

Genetically modified mice are provided that express human λ variable (hVλ) sequences, including mice that express hVλ sequences from an endogenous mouse λ light chain locus, mice that express hVλ sequences from an endogenous mouse κ light chain locus, and mice that express hVλ sequences from a transgene or an episome wherein the hVλ sequence is linked to a mouse constant sequence. Mice are provided that are a source of somatically mutated human λ variable sequences useful for making antigen-binding proteins. Compositions and methods for making antigen-binding proteins that comprise human λ variable sequences, including human antibodies, are provided.

Claims

1. A mouse comprising at an endogenous κ

light chain locus in its germline;

(a) an unrearranged light chain variable region comprising a plurality of contiguous unrearranged functional human λ

light chain V (hVλ

) gene segments and a plurality of contiguous unrearranged functional human λ

light chain J (hJ λ

) gene segments, wherein the plurality of hV λ

gene segments and the plurality of hJ λ

gene segments are the only functional variable region gene segments in the unrearranged light chain variable region; and

(b) a mouse κ

light chain constant region gene,

wherein the plurality of contiguous unrearranged functional human λ

light chain V (hVλ

) gene segments and the plurality of contiguous unrearranged functional human λ

light chain J (hJ λ

) gene segments are operably linked to the mouse κ

light chain constant region gene such that the unrearranged light chain variable region is capable of rearranging to form a rearranged human λ

light chain variable region and the mouse expresses antibodies comprising a light chain comprising a variable region encoded by the rearranged human λ

light chain variable region and a constant region encoded by the mouse κ

light chain constant region gene.

2. The mouse of claim 1, wherein the plurality of unrearranged hVλ

gene segments are at least 12 hVλ

gene segments.

3. The mouse of claim 1, wherein the plurality of hJλ

gene segments are selected from the group consisting of Jλ

1, Jλ

2, Jλ

3, Jλ

7, and a combination thereof.

4. The mouse of claim 1, wherein an endogenous mouse X light chain locus is deleted in whole or in part.

5. The mouse of claim 1, wherein about 10% to about 45% of the B cells of the mouse express the antibodies comprising a light chain comprising a variable region encoded by the rearranged human λ

light chain variable region and a constant region encoded by the mouse κ

light chain constant region gene.

6. The mouse of claim 5, wherein the light chain variable region of the antibody is derived from a rearranged hVλ

/hJλ

sequence selected from the group consisting of 3-1/1, 3-1/7, 4-3/1, 4-3/7, 2-8/1, 3-9/1, 3-10/1, 3-10/3, 3-10/7, 2-14/1, 3-19/1, 2-23/1, 3-25/1, 1-40/1, 1-40/2, 1-40/3, 1-40/7, 7-43/1, 7-43/3, 1-44/1, 1-44/7, 5-45/1, 5-45/2, 5-45/7, 7-46/1, 7-46/2, 7-46/7, 9-49/1, 9-49/2, 9-49/7 and 1-51/1.

7. The mouse of claim 1, wherein the unrearranged light chain variable region comprises a human Vκ

-Jκ

intergenic region from a human κ

light chain locus located downstream of the most 3′

hVλ

gene segment and upstream of the most 5′

hJλ

gene segment.

11. The mouse of claim 7, wherein the human Vκ

-Jκ

intergenic region comprises SEQ ID NO;

100.

12. An isolated cell of the mouse of claim 1.

13. The cell of claim 12, wherein the cell is a B cell.

16. The mouse of claim 1, wherein the plurality of hJλ

gene segments comprises 4 hJλ

gene segments.

17. The mouse of claim 16, wherein the 4 hJλ

gene segments are Jλ

1, Jλ

2, Jλ

3 and Jλ

7.

8. A mouse comprising at an endogenous κ

light chain locus in its germline;

(a) an unrearranged light chain variable region comprising;

(i) at least 12 contiguous unrearranged functional human λ

light chain variable region (hVλ

) gene segments and a plurality of contiguous unrearranged functional human λ

light chain J (hJλ

) gene segments, wherein the at least 12 functional hVλ

gene segments and the plurality of functional hJλ

gene segments are the only functional variable region gene segments in the unrearranged light chain variable region; and

(ii) a human Vκ

-Jκ

intergenic sequence located between the contiguous hVλ

gene segments and the plurality of contiguous hJλ

gene segments; and

(b) a mouse κ

light chain constant region gene;

wherein the at least 12 contiguous unrearranged functional human λ

light chain V (hVλ

) gene segments and the plurality of contiguous unrearranged functional human λ

light chain J (hJ λ

) gene segments are operably linked to the mouse κ

light chain constant region gene such that the unrearranged light chain variable region is capable of rearranging to form a rearranged human λ

light chain variable region and the mouse expresses antibodies comprising a light chain comprising a variable region encoded by the rearranged human λ

light chain variable region and a constant region encoded by the mouse κ

light chain constant region gene.

9. The mouse of claim 8, wherein the unrearranged light chain variable region comprises at least 28 unrearranged hVλ

gene segments.

10. The mouse of claim 8, wherein the unrearranged light chain variable region comprises at least 40 unrearranged hVλ

gene segments.

14. An isolated cell of the mouse of claim 8.

15. The cell of claim 14, wherein the cell is a B cell.

18. The mouse of claim 8, wherein the plurality of hJλ

gene segments comprises 4 hJλ

gene segments.

19. The mouse of claim 18, wherein the 4 hJλ

gene segments are Jλ

1, Jλ

2, Jλ

3 and Jλ

7.