Mice expressing a limited immunoglobulin light chain repertoire
A genetically modified mouse is provided, wherein the mouse expresses an immunoglobulin light chain repertoire characterized by a limited number of light chain variable domains. Mice are provided that present a choice of two human light chain variable gene segments such that the immunoglobulin light chains expresses by the mouse comprise one of the two human light chain variable gene segments. Methods for making bispecific antibodies having universal light chains using mice as described herein, including human light chain variable regions, are provided. Methods for making human variable regions suitable for use in multispecific binding proteins, e.g., bispecific antibodies, and host cells are provided.
1. A method for generating a human immunoglobulin heavy or light chain variable domain sequence comprising the steps of:
(a) immunizing a genetically modified mouse with an antigen of interest, wherein the genetically modified mouse comprises in its germline genome:
(i) exactly two unrearranged human immunoglobulin Vκ gene segments and five unrearranged human immunoglobulin Jκ gene segments operably linked to a mouse immunoglobulin light chain constant region sequence at the endogenous kappa light chain loci of the mouse, wherein the two unrearranged human immunoglobulin Vκ gene segments are a human Vκ1-39 gene segment and a human Vκ3-20 gene segment; and
(ii) one or more unrearranged human immunoglobulin V H gene segments, one or more unrearranged human immunoglobulin D H gene segments, and one or more unrearranged human immunoglobulin J H gene segments operably linked to a mouse immunoglobulin heavy chain constant region sequence at the endogenous heavy chain loci of the mouse;
wherein the unrearranged human immunoglobulin heavy chain and kappa light chain gene segments of the genetically modified mouse are capable of rearranging and encoding human immunoglobulin variable domains of an antibody, wherein the genetically modified mouse does not comprise endogenous immunoglobulin Vκ or Jκ gene segments that are capable of rearranging to form an immunoglobulin light chain variable region sequence; and
(b) determining a human immunoglobulin heavy or light chain variable domain sequence of an antibody that specifically binds the antigen and that was generated by the genetically modified mouse.
2. The method of claim 1 , wherein determining the human immunoglobulin heavy or light chain variable domain sequence comprises determining a nucleotide sequence that encodes the human immunoglobulin heavy or light chain variable domain sequence.
3. The method of claim 1 , wherein the five unrearranged human immunoglobulin Jκ gene segments are a human Jκ1 gene segment, a human Jκ2 gene segment, a human Jκ3 gene segment, a human Jκ4 gene segment, and a human Jκ5 gene segment.
4. The method of claim 1 , wherein the human Vκ1-39 gene segment in the germline genome of the genetically modified mouse is a human germline Vκ1-39 gene segment and the human Vκ3-20 gene segment in the germline genome of the genetically modified mouse is a human germline Vκ3-20 gene segment.
5. The method of claim 1 , wherein the germline genome of the genetically modified mouse comprises in order: the human Vκ1-39 gene segment, the human Vκ3-20 gene segment, the human Jκ1 gene segment, the human Jκ2 gene segment, the human Jκ3 gene segment, the human Jκ4 gene segment, and the human Jκ5 gene segment.
6. The method of claim 1 , wherein the genetically modified mouse comprises a nonfunctional λ light chain locus.
7. The method of claim 1 , wherein the genetically modified mouse comprises a functional λ light chain locus.
8. A method of making a fully human immunoglobulin heavy chain or a fully human immunoglobulin light chain comprising the steps of:
(a) immunizing a genetically modified mouse with an antigen of interest, wherein the genetically modified mouse comprises in its germline genome:
(i) exactly two unrearranged human immunoglobulin Vκ gene segments and five unrearranged human immunoglobulin Jκ gene segments operably linked to a mouse immunoglobulin light chain constant region sequence at the endogenous kappa light chain loci of the mouse, wherein the two unrearranged human immunoglobulin Vκ gene segments are a human Vκ1-39 gene segment and a human Vκ3-20 gene segment; and
(ii) one or more unrearranged human immunoglobulin V H gene segments, one or more unrearranged human immunoglobulin D H gene segments, and one or more unrearranged human immunoglobulin J H gene segments operably linked to a mouse immunoglobulin heavy chain constant region sequence at the endogenous heavy chain loci of the mouse;
wherein the unrearranged human immunoglobulin heavy chain and kappa light chain gene segments of the genetically modified mouse are capable of rearranging and encoding human immunoglobulin variable domains of an antibody, wherein the genetically modified mouse does not comprise endogenous immunoglobulin Vκ or Jκ gene segments that are capable of rearranging to form an immunoglobulin light chain variable region sequence;
(b) determining a human immunoglobulin heavy or light chain variable domain sequence of an antibody that specifically binds the antigen and that was generated by the genetically modified mouse; and
(c) operably linking the human immunoglobulin heavy or light chain variable domain sequence to a human immunoglobulin heavy or light chain constant domain sequence, respectively, to form a fully human immunoglobulin heavy chain or a fully human immunoglobulin light chain.
9. The method of claim 8 , wherein determining the human immunoglobulin heavy or light chain variable domain sequence comprises determining a nucleotide sequence that encodes the human immunoglobulin heavy or light chain variable domain sequence.
10. The method of claim 8 , wherein operably linking the human immunoglobulin heavy or light chain variable domain sequence to the human immunoglobulin heavy or light chain constant domain sequence, respectively, comprises operably linking a nucleotide sequence encoding the human immunoglobulin heavy or light chain variable domain sequence to a nucleotide sequence encoding the human immunoglobulin heavy or light chain constant domain sequence.
11. The method of claim 8 , wherein the five unrearranged human immunoglobulin Jκ gene segments are a human Jκ1 gene segment, a human Jκ2 gene segment, a human Jκ3 gene segment, a human Jκ4 gene segment, and a human Jκ5 gene segment.
12. The method of claim 8 , wherein the human Vκ1-39 gene segment in the germline genome of the genetically modified mouse is a human germline Vκ1-39 gene segment and the human Vκ3-20 gene segment in the germline genome of the genetically modified mouse is a human germline Vκ3-20 gene segment.
13. The method of claim 8 , wherein the germline genome of the genetically modified mouse comprises in order: the human Vκ1-39 gene segment, the human Vκ3-20 gene segment, the human Jκ1 gene segment, the human Jκ2 gene segment, the human Jκ3 gene segment, the human Jκ4 gene segment, and the human Jκ5 gene segment.
14. The method of claim 8 , wherein the genetically modified mouse comprises a nonfunctional λ light chain locus.
15. The method of claim 8 , wherein the genetically modified mouse comprises a functional λ light chain locus.