Non-human transgenic mammal for the constant region of the class a human immunoglobulin heavy chain and applications thereof
The invention relates to a non-human transgenic mammal with an IgH locus modified by replacement of the switching sequence Sμ with all or part of a transgene comprising the gene Cα of a class A human immunoglobulin, including at least the exon, coding for the CH3 domain and the membrane exon and the applications of the above for the production of humanized class IgA antibodies.
1. A transgenic mouse, wherein an endogenous IgH locus comprises replacement of its switch sequence Sμ with a transgene consisting essentially of a human class A immunoglobulin heavy chain constant region gene Cα or a segment of said Cα gene comprising at least an exon encoding the CH3 domain and a membrane exon, wherein said transgenic mouse produces chimeric immunoglobulins A whose heavy chains comprise a mouse variable region and a human constant region or a segment thereof comprising at least the CH3 domain, and wherein said transgenic mouse produces no mouse immunoglobulins M.
2. The transgenic mouse of claim 1 , which is homozygous for said modified IgH locus.
3. The transgenic mouse of claim 1 , wherein said transgene consists of the entire Cα gene.
4. The transgenic mouse of claim 1 , wherein said transgene consists of the segment of the Cα gene comprising the exon encoding the CH3 domain and the membrane exon.
5. The transgenic mouse of claim 1 , wherein said Cα gene is the Cα1 gene.
6. The transgenic mouse of claim 1 , which further comprises another transgene encoding a human immunoglobulin light chain.
7. The transgenic mouse of claim 6 , wherein said light chain is a kappa light chain.
8. The transgenic mouse of claim 6 , wherein said transgene which encodes a human immunoglobulin kappa light chain, further comprises the intronic activator Eμ upstream of a DNA sequence encoding said human immunoglobulin kappa light chain and the palindrome hs3a/hs1,2/hs3b downstream of said DNA sequence.
9. The transgenic mouse of claim 8 , wherein said transgene is under the control of the promoter of the human immunoglobulin heavy chain.
10. The transgenic mouse of claim 6 , which is dizygous for said transgene.
11. The transgenic mouse of claim 6 , further comprising an inactivated endogenous immunoglobulin kappa light chain locus.
12. The transgenic mouse of claim 11 , which is homozygous for said inactivated endogenous immunoglobulin kappa light chain locus.
13. The transgenic mouse of claim 1 , further comprising an inactivated endogenous J chain gene.
14. The transgenic mouse of claim 13 , which is homozygous for said inactivated endogenous J chain gene.
15. The transgenic mouse of claim 13 , which further comprises another transgene encoding a human immunoglobulin J chain gene.
16. The transgenic mouse of claim 1 , wherein said:
a) endogenous mouse IgH locus comprises the replacement of its switch sequence Sμ with the entire human class A immunoglobulin heavy chain constant region gene Cα1, and
b) which transgenic mouse further comprises a human kappa light chain transgene comprising a VκI gene rearranged with a Jκ5 gene, a Jκ-Cκ intron and a Cκ gene, under the transcriptional control of the human heavy chain promoter (pVH), the intronic activator Eμ upstream of said promoter of and the palindrome hs3a/hs1,2/hs3b downstream of said Cκ gene.
17. A homologous recombination targeting vector, which comprises a human class A immunoglobulin heavy chain constant region gene Cα or a segment of said Cα gene comprising at least an exon encoding the CH3 domain and a membrane exon, flanked by the sequences SEQ ID NO: 7 and SEQ ID NO: 8.
18. The targeting vector of claim 17 , which comprises a cassette for expressing a selection marker, adjacent to said Cα gene or to a segment of said gene.
19. The targeting vector of claim 18 , wherein said expression cassette is flanked by site-specific recombination sequences.
20. The targeting vector of claim 19 wherein said sequences are LoxP sequences of Cre recombinase.
21. A mouse embryonic cell, which is modified with the targeting vector of claim 17 .
22. A method for preparing humanized class IgA antibodies or fragments thereof, which comprises at least the following steps:
a) immunizing the transgenic mouse of claim 1 , and
b) producing humanized class IgA antibodies or fragments of the antibodies from serum secretions or B lymphocytes of said transgenic mouse sacrificed beforehand.