Method of making transgenic mice lacking endogenous heavy chains

• US 5,939,598 A
• Filed: 07/30/1992
• Issued: 08/17/1999
• Est. Priority Date: 01/12/1990
• Status: Expired due to Term

• Current Assignee: Abgenix Inc (Amgen Inc.)
• Original Assignee: Abgenix Inc (Amgen Inc.)
• Inventors: Jakobovits, Aya, Kucherlapati, Raju
• Primary Examiner(s): Crouch, Deborah
• Assistant Examiner(s): HAUDA, KAREN MARIE
• Application Number:
  US07/922,649
• Time in Patent Office: 2,574 Days
• Field of Search: 800/2, 800/21, 800/25, 435/172.3, 536/23.53
• US Class Current: 800/25
• 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 2217/072 maintaining or altering function, i.e. knock in
• CPC Class Codes: A01K 2217/075 inducing loss of function, i.e. knock out
• CPC Class Codes: A01K 2227/105 Murine
• CPC Class Codes: A01K 2267/01 Animal expressing industrially exogenous proteins
• 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 67/0276 Knock-out vertebrates
• CPC Class Codes: A01K 67/0278 Knock-in vertebrates, e.g. humanised vertebrates
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• 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 16/1282 from Clostridium (G)
• CPC Class Codes: C07K 16/241 Tumor Necrosis Factors
• CPC Class Codes: C07K 16/244 Interleukins [IL]
• CPC Class Codes: C07K 16/248 IL-6
• CPC Class Codes: C07K 16/2812 against CD4
• CPC Class Codes: C07K 16/2854 against selectins, e.g. CD62
• CPC Class Codes: C07K 16/2875 against the NGF/TNF superfamily, e.g. CD70, CD95L, CD153, CD154 against NGF C07K16/22, against TNF C07K16/241
• 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 2317/21 : 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: C12N 15/8509 : for producing genetically modified animals, e.g. transgenic

Abstract

The subject invention provides non-human mammalian hosts characterized by inactivated endogenous Ig loci. The hosts are produced by repetitive transformations of embryonic stem cells by homologous recombination, preferably in conjunction with breeding. Different strategies are employed for recombination of the human loci randomly or at analogous host loci.

Claims

1. A method for producing a transgenic mouse lacking expression of an endogenous immunoglobulin heavy chain, comprising:

(a) deleting the J segment genes from at least one endogenous heavy chain locus in a mouse embryonic stem cell to prevent rearrangement of the locus and to prevent formation of a transcript of a rearranged immunoglobulin heavy chain locus, the deletion being effected by a targeting vector comprising a gene encoding a selectable marker; and

(b) producing from the embryonic stem cell a transgenic mouse whose somatic and germ cells contain the gene encoding the selectable marker,

wherein the transgenic mouse lacks expression of an endogenous immunoglobulin heavy chain.

5. The method according to any one of claims 1-4, wherein the gene encoding a selectable marker is a neomycin resistance gene.

13. The method according to any one of claims 1, 2, 6 or 7, further comprising producing progeny of the transgenic mouse by breeding.

2. A method for producing a transgenic mouse lacking expression of an endogenous immunoglobulin heavy chain, comprising:

(a) deleting the J segment genes from at least one endogenous heavy chain locus in a mouse embryonic stem cell to prevent rearrangement of the locus and to prevent formation of a transcript of a rearranged immunoglobulin heavy chain locus, the deletion being effected by a targeting vector comprising a gene encoding a selectable marker; and

(b) producing from the embryonic stem cell a transgenic mouse whose somatic and germ cells contain an immunoglobulin heavy chain locus in which all of the J segment genes are deleted,

wherein the transgenic mouse lacks expression of an endogenous immunoglobulin heavy chain.

3. A method for producing a transgenic mouse and its progeny lacking expression of endogenous immunoglobulin heavy chains, comprising;

(a) deleting the J segment genes from at least one endogenous heavy chain locus in a mouse embryonic stem cell to prevent rearrangement of the locus and to prevent formation of a transcript of a rearranged immunoglobulin heavy chain locus, the deletion being effected by a targeting vector comprising a gene encoding a selectable marker;

(b) producing from the embryonic stem cell a transgenic mouse whose somatic and germ cells contain an immunolobulin heavy chain locus in which all of the J segment genes are deleted, and

(c) breeding the transgenic mouse as needed to produce a transgenic mouse and its progeny lacking expression of endogenous immunoglobulin heavy chains.

4. A method for producing a transgenic mouse and its progeny lacking expression of endogenous immunoglobulin heavy chains, comprising:

(a) deleting the J segment genes from at least one endogenous heavy chain locus in a mouse embryonic stem cell to prevent rearrangement of the locus and to prevent formation of a transcript of a rearranged immunoglobulin heavy chain locus, the deletion being effected by a targeting vector comprising a gene encoding a selectable marker;

(b) producing from the embryonic stem cell a transgenic mouse whose somatic and germ cells contain the gene encoding the selectable marker, and

(c) breeding the transgenic mouse as needed to produce a transgenic mouse and its progeny lacking expression of endogenous immunoglobulin heavy chains.

6. A method for preventing the expression of an endogenous immunoglobulin heavy chain in a transgenic mouse, comprising:

(a) deleting the J segment genes from at least one endogenous heavy chain locus in a mouse embryonic stem cell to prevent rearrangement of the locus and to prevent formation of a transcript of a rearranged immunoglobulin heavy chain locus, the deletion being effected by a targeting vector comprising a gene encoding a selectable marker; and

(b) producing from the embryonic stem cell a transgenic mouse whose somatic and germ cells contain the gene encoding the selectable marker,

wherein the transgenic mouse lacks expression of an endogenous immunoglobulin heavy chain.

10. The method according to any one of claims 6-9, wherein the gene encoding a selectable marker is a neomycin resistance gene.

7. A method for preventing the expression of an endogenous immunoglobulin heavy chain in a transgenic mouse, comprising:

(a) deleting the J segment genes from at least one endogenous heavy chain locus in a mouse embryonic stem cell to prevent rearrangement of the locus and to prevent formation of a transcript of a rearranged immunoglobulin heavy chain locus, the deletion being effected by a targeting vector comprising a gene encoding a selectable marker; and

(b) producing from the embryonic stem cell a transgenic mouse whose somatic and germ cells contain an endogenous immunoglobulin heavy chain locus in which all of the J segment genes are deleted,

wherein the transgenic mouse lacks expression of an endogenous immunoglobulin heavy chain.

8. A method for preventing the expression of endogenous immunoglobulin heavy chains in a transgenic mouse and its progeny, comprising:

(a) deleting J segment genes from at least one endogenous heavy chain locus in a mouse embryonic stem cell to prevent rearrangement of the locus and to prevent formation of a transcript of a rearranged immunoglobulin heavy chain locus, the deletion being effected by a targeting vector comprising a gene encoding a selectable marker;

(b) producing from the embryonic stem cell a transgenic mouse whose somatic and germ cells contain an immunoglobulin heavy chain locus in which all of the J segment genes are deleted, and

(c) breeding the transgenic mouse as needed to produce a transgenic mouse and its progeny lacking expression of endogenous immunoglobulin heavy chains.

9. A method for preventing the expression of endogenous immunoglobulin heavy chains in a transgenic mouse and its progeny, comprising:

(a) deleting J segment genes from at least one endogenous heavy chain locus in a mouse embryonic stem cell to prevent rearrangement of the locus and to prevent formation of a transcript of a rearranged immunoglobulin heavy chain locus, the deletion being effected by a targeting vector comprising a gene encoding a selectable marker;

(b) producing from the embryonic stem cell a transgenic mouse whose somatic and germ cells contain the gene encoding the selectable marker, and

(c) breeding the transgenic mouse as needed to produce a transgenic mouse and its progeny lacking expression of endogenous immunoglobulin heavy chains.

11. A method for producing a mouse embryonic stem cell with at least one inactivated endogenous immunoglobulin heavy chain locus, comprising deleting the J segment genes of at least one endogenous immunoglobulin heavy chain locus in the mouse embryonic stem cell to prevent rearrangement of the locus and to prevent the formation of a transcript of a rearranged immunoglobulin heavy chain, the deletion being effected by a targeting vector comprising a gene encoding a selectable marker.

12. The method according to claim 11, wherein the gene encoding a selectable marker is a neomycin resistance gene.