Genetic engineering of non-human animals for the production of chimeric antibodies

• US 10,626,188 B2
• Filed: 03/20/2017
• Issued: 04/21/2020
• Est. Priority Date: 03/31/2010
• Status: Active Grant

• Current Assignee: Ablexis LLC
• Original Assignee: Ablexis LLC
• Inventors: Green, Larry, Shizuya, Hiroaki
• Primary Examiner(s): Ton, Thaian N.
• Assistant Examiner(s): Montanari, David A.
• Application Number:
  US15/463,761
• Publication Number:
  US 20170188558A1
• Time in Patent Office: 1,128 Days
• CPC Class Codes: A01K 2207/15 Humanized animals
• 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 2227/105 Murine
• CPC Class Codes: A01K 2267/01 Animal expressing industrially exogenous proteins
• 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 2317/14 Specific host cells or culture conditions, e.g. components, pH or temperature
• CPC Class Codes: C07K 2317/24 containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
• CPC Class Codes: C07K 2317/51 Complete heavy chain or Fd fragment, i.e. VH + CH1
• CPC Class Codes: C07K 2317/515 Complete light chain, i.e. VL + CL
• CPC Class Codes: C07K 2317/52 Constant or Fc region; Isotype
• CPC Class Codes: C07K 2317/522 CH1 domain
• CPC Class Codes: C07K 2317/524 CH2 domain
• CPC Class Codes: C07K 2317/526 CH3 domain
• CPC Class Codes: C07K 2317/53 Hinge
• 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 15/8509 : for producing genetically modified animals, e.g. transgenic
• CPC Class Codes: C12N 2510/02 : Cells for production

Abstract

The invention provides non-human cells and mammals having a genome encoding chimeric antibodies and methods of producing transgenic cells and mammals. Certain aspects of the invention include chimeric antibodies, humanized antibodies, pharmaceutical compositions and kits. Certain aspects of the invention also relate to diagnostic and treatment methods using the antibodies of the invention.

Claims

1. A mouse whose genome comprises a transgene encoding a polypeptide comprising an immunoglobulin light chain variable region, wherein the transgene comprises (1) a plurality of immunoglobulin light chain variable (V) exons encoding human immunoglobulin light chain variable (V) polypeptides;

(2) non-coding sequences between the V exons;

(3) a plurality of immunoglobulin light chain joining (J) coding sequences encoding human immunoglobulin light chain joining (J) polypeptides; and

(4) non-coding sequences between the J coding sequences;

wherein the non-coding sequences between the V exons and the non-coding sequences between the J coding sequences are derived from mouse immunoglobulin light chain non-coding sequences and wherein the transgene is capable of undergoing gene arrangement and thereby upon expression to produce a polypeptide comprising the immunoglobulin light chain variable region.

2. The mouse according to claim 1, wherein the non-coding sequences between the V exons and the non-coding sequences between the J coding sequences are selected from the group consisting of intronic sequences and cis regulatory sequences.

3. The mouse according to claim 2, wherein the cis regulatory sequences are selected from promoters, enhancers, recombination signal sequences, splice acceptor sequences, and splice donor sequences.

4. The mouse according to claim 1, wherein the V exons encode human kappa light chain V (Vκ

) polypeptides or human lambda light chain V (Vλ

) polypeptides.

5. The mouse according to claim 1, wherein (1) the mouse non-coding sequences between the V exons and (2) the V are from non-orthologous immunoglobulin loci.

6. The mouse according to claim 1, wherein the transgene further comprises a coding sequence encoding a human immunoglobulin light chain constant (C) polypeptide.

7. The mouse according to claim 6, wherein the coding sequence encoding the human immunoglobulin light chain C polypeptide encodes a human Cλ

polypeptide or a human Cκ

polypeptide.

8. The mouse according to claim 1, wherein the transgene further comprises a coding sequence encoding a mouse immunoglobulin light chain constant (C) polypeptide.

9. The mouse according to claim 8, wherein the transgene comprises human Vλ

coding sequences and 2 to 7 Jλ

-Cλ

coding sequence pairs comprising a human Jλ and

a mouse Cλ

.

10. The mouse according to claim 1, wherein the transgene further comprises mouse non-coding sequences upstream of the V exons.

11. The mouse according to claim 1, wherein the transgene further comprises mouse non-coding sequences downstream of the J coding sequences.

12. A mouse whose genome comprises a transgene encoding a polypeptide comprising an immunoglobulin lambda light chain variable region, wherein the transgene comprises (1) a plurality of human immunoglobulin light chain variable Vλ

exons encoding human immunoglobulin light chain variable Vλ

polypeptides;

(2) mouse non-coding sequences between the Vλ

exons;

(3) 2-7 immunoglobulin light chain Jλ

-Cλ

coding sequence pairs, wherein the Jλ

sequence encodes a human immunoglobulin light chain Jλ

polypeptide and the Cλ

sequence encodes a mouse immunoglobulin light chain Cλ

polypeptide;

(4) mouse non-coding sequences between the Jλ

coding sequences and the Cλ

coding sequences in the Jλ

-Cλ

coding sequence pairs; and

(5) mouse non-coding sequences between the Jλ

-Cλ

coding sequence pairs, wherein the transgene is capable of undergoing gene arrangement and thereby upon expression to produce a polypeptide comprising the immunoglobulin lambda light chain variable region.

13. The mouse according to claim 12, wherein the transgene comprises 6 Jλ

-Cλ

coding sequence pairs.

14. The mouse according to claim 12, wherein the transgene further comprises mouse non-coding sequences upstream of the Vλ

coding sequences.

15. The mouse according to claim 12, wherein the transgene further comprises mouse non-coding sequences downstream of the Jλ

-Cλ

coding sequence pairs.

20. The mouse according to claim 12, wherein the transgene further comprises a human immunoglobulin light chain λ

3′

enhancer.

21. The mouse according to claim 12, wherein said transgene further comprises an Ig light chain 3′

LCR, or a functional fragment thereof.

22. The mouse according to claim 21, wherein said Ig light chain 3′

LCR, or a functional fragment thereof, is from a mammal selected from the group consisting of human, non-primate and rat.

16. A mouse whose genome comprises a transgene encoding a polypeptide comprising an immunoglobulin kappa light chain variable region, wherein the transgene comprises (1) a plurality of human immunoglobulin light chain variable Vκ

coding sequences encoding human immunoglobulin light chain variable Vκ

polypeptides;

(2) mouse non-coding sequences between the Vκ

coding sequences;

(3) a plurality of human immunoglobulin light chain variable Jκ

coding sequences encoding human immunoglobulin light chain variable Jκ

polypeptides;

(4) mouse non-coding sequences between the Jκ

coding sequences; and

(5) a Cκ

coding sequence encoding a mouse immunoglobulin light chain constant Cκ

polypeptide, wherein the transgene is capable of undergoing gene arrangement to produce upon expression a polypeptide comprising the immunoglobulin kappa light chain variable region.

17. The mouse according to claim 16, wherein the transgene further comprises mouse non-coding sequences upstream of the Vκ

coding sequences.

18. The mouse according to claim 16, wherein the transgene further comprises mouse non-coding sequences downstream of the Cκ

coding sequence.

19. The mouse according to claim 16, wherein the transgene further comprises 5 Jκ

exons.