Expression of xenogenous (human) immunoglobulins in cloned, transgenic ungulates

• US 20060117395A1
• Filed: 12/01/2005
• Published: 06/01/2006
• Est. Priority Date: 11/19/1999
• Status: Abandoned Application

• Current Assignee: Kyowa Hakko Kirin Company Limited (Kirin Holdings Company Limited)
• Original Assignee: Kirin Beer Kabushiki Kaisha, Hematech, LLC (MeritCare Health System)
• Inventors: Kuroiwa, Yoshimi, Goldsby, Richard A., Tomizuka, Kazuma, Ferguson, Stacy E., Osborne, Barbara A., Ishida, Isao, Robl, James M.
• Application Number:
  US11/292,164
• Publication Number:
  US 20060117395A1
• Time in Patent Office: Days
• US Class Current: 800/6
• CPC Class Codes: A01K 2207/15 Humanized animals
• CPC Class Codes: A01K 2217/00 Genetically modified animals
• CPC Class Codes: A01K 2217/05 Animals comprising random inserted nucleic acids (transgenic)
• CPC Class Codes: A01K 2217/075 inducing loss of function, i.e. knock out
• CPC Class Codes: A01K 2227/101 Bovine
• CPC Class Codes: A01K 2267/01 Animal expressing industrially exogenous proteins
• CPC Class Codes: A01K 67/0273 Cloned vertebrates
• CPC Class Codes: A01K 67/0276 Knock-out vertebrates
• CPC Class Codes: A01K 67/0278 Knock-in vertebrates, e.g. humanised vertebrates
• CPC Class Codes: C12N 15/8509 for producing genetically modified animals, e.g. transgenic
• CPC Class Codes: C12N 2800/30 Vector systems comprising sequences for excision in presence of a recombinase, e.g. loxP or FRT
• CPC Class Codes: C12N 9/1051 Hexosyltransferases (2.4.1)

Abstract

The present invention relates to the production of a transgenic bovine which comprises a genetic modification that results in inactivation and loss of expression of its endogenous antibodies, and the expression of xenogenous antibodies, preferably human antibodies. This is effected by inactivation of the IgM heavy chain expression and, optionally, by inactivation of the Ig light chain expression, and by the further introduction of an artificial chromosome which results in the expression of non-bovine antibodies, preferably human antibodies.

Claims

1. A transgenic ungulate whose cells comprise a mutation that reduces the expression of an endogenous antibody.

2. The transgenic ungulate of claim 1, wherein said mutation reduces the expression of a functional immunoglobulin heavy chain.

3. The transgenic ungulate of claim 2, wherein said mutation reduces the expression of a functional IgM heavy chain.

4. The transgenic ungulate of claim 1, wherein said mutation reduces the expression of a functional immunoglobulin light chain.

5. The transgenic ungulate of claim 1, wherein said mutation is a nonsense or deletion mutation.

6. The transgenic ungulate of claim 1, the cells of said ungulate comprising one or more nucleic acids, each nucleic acid comprising one or more xenogenous immunoglobulin heavy or light chain loci that undergo rearrangement and are expressed in B cells to produce xenogenous immunoglobulin in response to one or more antigens.

7. The transgenic ungulate of claim 6, wherein said xenogenous immunoglobulin is human immunoglobulin.

8. The transgenic ungulate of claim 6, wherein said xenogenous immunoglobulin loci comprise both a human immunoglobulin light chain locus and a human immunoglobulin heavy chain locus.

9. The transgenic ungulate of claim 6, wherein said one or more nucleic acids comprise one or more artificial chromosomes.

10. The transgenic ungulate of claim 9, wherein said one or more artificial chromosomes comprise a human artificial chromosome.

11. The transgenic ungulate of claim 10, wherein said human artificial chromosome is Δ

HAC or Δ

Δ

HAC.

12. The transgenic ungulate of claim 10, wherein said human artificial chromosome is derived from one or more of human chromosome 14, human chromosome 2, and human chromosome 22.

13. The transgenic ungulate of claim 6, wherein said one or more nucleic acids comprise a chromosome fragment.

14. The transgenic ungulate of claim 13, wherein said chromosome fragment is selected from the group consisting of a YAC, BAC, or cosmid vector.

15. The transgenic ungulate of claim 1, wherein said ungulate is a bovine, ovine, porcine, or caprine.

16. The transgenic ungulate of claim 15, wherein said ungulate is a bovine.

17. An ungulate somatic cell comprising a mutation that reduces the expression of an endogenous antibody.

18. The ungulate somatic cell of claim 17, wherein said mutation reduces the expression of a functional immunoglobulin heavy chain.

19. The ungulate somatic cell of claim 18, wherein said mutation reduces the expression of a functional IgM heavy chain.

20. The ungulate somatic cell of claim 17, wherein said mutation reduces the expression of a functional immunoglobulin light chain.

21. The ungulate somatic cell of claim 17, wherein said mutation is a nonsense or deletion mutation.

22. The ungulate somatic cell of claim 17, comprising one or more nucleic acids, each nucleic acid comprising one or more xenogenous immunoglobulin heavy or light chain loci that undergo rearrangement and are expressed in B cells to produce xenogenous immunoglobulin in response to exposure to one or more antigens.

23. The ungulate somatic cell of claim 22, wherein said xenogenous immunoglobulin is human immunoglobulin.

24. The ungulate somatic cell of claim 22, wherein said xenogenous immunoglobulin loci comprise both a human immunoglobulin light chain locus and a human immunoglobulin heavy chain locus.

25. The ungulate somatic cell of claim 22, wherein said one or more nucleic acids comprise one or more artificial chromosomes.

26. The ungulate somatic cell of claim 25, wherein said one or more artificial chromosomes comprise a human artificial chromosome.

27. The ungulate somatic cell of claim 26, wherein said human artificial chromosome is Δ

HAC or Δ

Δ

HAC.

28. The ungulate somatic cell of claim 26, wherein said human artificial chromosome is derived from one or more of human chromosome 14, human chromosome 2, and human chromosome 22.

29. The ungulate somatic cell of claim 22, wherein said one or more nucleic acids comprise a chromosome fragment.

30. The ungulate somatic cell of claim 29, wherein said chromosome fragment is selected from the group consisting of a YAC, BAC, or cosmid vector.

31. The ungulate somatic cell of claim 17, wherein said cell is a fetal fibroblast or a B-cell.

32. The ungulate somatic cell of claim 17, wherein said ungulate is a bovine, ovine, porcine, or caprine.

33. The ungulate somatic cell of claim 32, wherein said ungulate is a bovine.

34. A method of producing xenogenous antibodies against one or more antigens, said method comprising the steps of:

(a) administering one or more antigens to a transgenic ungulate whose cells comprise a mutation that reduces the expression of an endogenous antibody and further comprise one or more nucleic acids, each nucleic acid comprising one or more xenogenous immunoglobulin heavy or light chain loci that undergo rearrangement and are expressed in B cells to produce xenogenous immunoglobulin, resulting in production of xenogenous antibodies against for said one or more antigens; and

(b) recovering said xenogenous antibodies from said ungulate.

36. The method of claim 34 or 35, wherein said mutation reduces the expression of a functional immunoglobulin heavy chain.

37. The method of claim 36, wherein said mutation reduces the expression of a functional IgM heavy chain.

38. The method of claim 34 or 35, wherein said mutation reduces the expression of a functional immunoglobulin light chain.

39. The method of claim 34 or 35, wherein said mutation is a nonsense or deletion mutation.

40. The method of claim 34 or 35, wherein said xenogenous antibodies are human antibodies.

41. The method of claim 34 or 35, wherein said one or more nucleic acids comprise one or more artificial chromosomes.

42. The method of claim 41, wherein said one or more artificial chromosomes comprise a human artificial chromosome.

43. The method of claim 42, wherein said human artificial chromosome is Δ

HAC or Δ

Δ

HAC.

44. The method of claim 42, wherein said human artificial chromosome is derived from one or more of human chromosome 14, human chromosome 2, and human chromosome 22.

45. The method of claim 34 or 35, wherein said one or more nucleic acids comprise a chromosome fragment.

46. The method of claim 45, wherein said chromosome fragment is selected from the group consisting of a YAC, BAC, or cosmid vector.

47. The method of claim 34 or 35, wherein said ungulate is a bovine, ovine, porcine, or caprine.

48. The method of claim 47, wherein said ungulate is a bovine.

35. A method of producing xenogenous antibodies against one or more antigens, said method comprising recovering xenogenous antibodies from a transgenic ungulate whose cells comprise a mutation that reduces the expression of an endogenous antibody and further comprise one or more nucleic acids, each nucleic acid comprising one or more xenogenous immunoglobulin heavy or light chain loci that undergo rearrangement and are expressed in B cells, resulting in production of xenogenous antibodies against said one or more antigens.