Animal models and therapeutic molecules

Abstract

The invention discloses methods for the generation of chimaeric human—non-human antibodies and chimaeric antibody chains, antibodies and antibody chains so produced, and derivatives thereof including fully humanised antibodies; compositions comprising said antibodies, antibody chains and derivatives, as well as cells, non-human mammals and vectors, suitable for use in said methods.

1 . A method of obtaining an antigen specific antibody or antigen binding fragment thereof, said antigen-specific antibody comprising a human immunoglobulin light (IgL) chain comprising a human IgL chain lambda (λ) variable region and a human IgL chain constant region, said antigen-binding fragment thereof comprising said human IgL chain λ variable region, the method comprising:
 expressing said antibody from a cell comprising nucleic acid encoding said human IgL chain λ variable region and said human IgL chain constant region of said antibody, or expressing said antigen binding fragment thereof, from a cell comprising nucleic acid encoding said human IgL chain λ variable region, 
 wherein said nucleic acid encoding said human IgL chain λ variable region is of a transgenic mouse contacted with said antigen, 
 wherein the germline of said transgenic mouse comprises an immunoglobulin light chain (IgL) locus, 
 said IgL locus comprising human IgL chain λ variable region gene segments consisting of one human Vλ gene segment and one human Jλ gene segment, said human Vλ gene segment is selected from the group consisting of functional Vλ gene segments positioned from 22,375,609 to 23,327,884 from human chromosome 22, with reference to the GRCH37/hg19 sequence database of a human λ light chain locus, 
 said human Jλ gene segment is selected from the group consisting of functional Jλ gene segments positioned from 22,375,609 to 23,327,884 from human chromosome 22, with reference to the GRCH37/hg19 sequence database of a human λ light chain locus, 
 wherein said human λ V region light chain gene segments are positioned
 (i) at an endogenous mouse immunoglobulin light chain locus comprising a light chain locus enhancer and 
 (ii) upstream of a constant region, 
 
 said IgL locus being functional to express an immunoglobulin light chain comprising a human λ variable region, 
 wherein said transgenic mouse comprises splenic B cells expressing immunoglobulin light chains comprising human λ variable regions, 
 wherein said transgenic mouse comprises immunoglobulin IgM antibody comprising said human λ variable regions. 
 
     
     
         2 . The method of  claim 1 , wherein said human Vλ gene segment is selected from the group consisting of functional Vλ gene segments from Vλ4-69 to Vλ3-1 of a human λ light chain locus, and wherein said human Jλ gene segment is selected from the group consisting of functional J gene segments of Jλ1, Jλ2, Jλ3 and Jλ7 of a human λ light chain locus, 
     
     
         3 . The method of  claim 2 , the method, further comprising:
 recovering a substance selected from the group consisting of: said antibody or antigen-binding fragment thereof; said nucleic acid encoding said human IgL chain variable region; and said cell expressing said antibody or antigen-binding fragment thereof.   
     
     
         4 . The method of  claim 1  above, further comprising the step of providing a pharmaceutical composition comprising said antigen-specific antibody, antigen-binding fragment thereof, said human IgL chain λ variable region, said nucleic acid encoding said human IgL chain λ variable region, or said cell expressing said antibody or antigen-binding fragment thereof, and a pharmaceutically acceptable carrier or excipient. 
     
     
         5 . The method of  claim 2 , wherein said human IgL chain λ variable region of said cell comprising nucleic acid encoding human IgL chain λ variable region and human IgL chain constant region of said antibody is obtained from a second cell comprising nucleic acid encoding said human IgL chain λ variable region. 
     
     
         6 . The method of  claim 5 , wherein said second cell is selected from the group consisting of:
 a B cell of said transgenic mouse contacted with said antigen; a cell comprising nucleic acid encoding said human IgL chain λ variable region and a mouse IgL chain constant region; a hybridoma expressing said human IgL chain λ variable region; a plurality of cells comprising nucleic acid encoding said human IgL chain λ variable region; and an immortalized cell comprising nucleic acid encoding said human IgL chain λ variable region.   
     
     
         7 . The method of  claim 2 , wherein said enhancer comprises a mouse λ enhancer. 
     
     
         8 . The method of  claim 2 , wherein said enhancer is a mouse κ enhancer. 
     
     
         9 . The method of  claim 2 , wherein at least 60% of immunoglobulin light chains expressed by said mouse comprise human IgL chain λ variable regions derived from recombination of human Vλ and Jλ gene segments. 
     
     
         10 . The method of  claim 2 , wherein said human IgL chain λ variable region gene segments are unrearranged and comprise a 3′ human Vλ gene segment which is positioned within 100 kb of said light chain locus enhancer. 
     
     
         11 . The method of  claim 2 , wherein the genome comprises endogenous V light chain DNA comprising endogenous Vλ or Vκ chain DNA. 
     
     
         12 . The method of  claim 2 , wherein said mouse comprises splenic B cells which express immunoglobulin gamma (IgG) antibody. 
     
     
         13 . The method of  claim 12 , wherein said immunoglobulin gamma (IgG) antibody comprises said human IgL chain λ variable regions comprising antigen specific human IgL light chain λ variable regions. 
     
     
         14 . The method of  claim 2  wherein said human Vλ and Jλ gene segments are positioned upstream to an Igκ constant region. 
     
     
         15 . The method of  claim 14 , wherein at least 60% of immunoglobulin light chains expressed by said mouse comprise human V regions derived from recombination of human Vλ and Jλ gene segments. 
     
     
         16 . The method of  claim 14 , wherein at least 85% of immunoglobulin light chains expressed by said mouse comprise human V regions derived from recombination of human Vλ and Jλ gene segments. 
     
     
         17 . The method of  claim 2 , wherein at least 85% of immunoglobulin light chains expressed by said mouse comprise human V regions derived from recombination of human Vλ and Jλ gene segments. 
     
     
         18 . The method of  claim 2 , wherein said one human Vλ gene segment is rearranged with said one human Jλ gene segment to encode a variable region. 
     
     
         19 . The method of  claim 2 , wherein said one human Vλ gene segment and said one human Jλ gene segment are unrearranged and are functional to rearrange to express an immunoglobulin light chain comprising a human variable region. 
     
     
         20 . The method of  claim 2 , wherein the germline of said transgenic mouse comprises a homozygous immunoglobulin light chain (IgL) locus. 
     
     
         21 . The method of  claim 20 , wherein all light chains expressed by said mouse are encoded by said homozygous IgL locus comprising human lambda V region light chain gene segments consisting of one human Vλ gene segment and one human Jλ gene segment. 
     
     
         22 . The method of  claim 2 , wherein endogenous light chain expression in said IgL locus is inactive. 
     
     
         23 . The method of  claim 2 , wherein endogenous kappa light chain expression in said IgL locus is inactive. 
     
     
         24 . The method of  claim 2 , wherein said light chain locus enhancer is endogenous. 
     
     
         25 . The method of  claim 2 , wherein a plurality of said immunoglobulin light chains comprises a plurality of human λ variable regions, wherein said plurality of said immunoglobulin light chains further comprises a plurality of mouse λ variable regions, and wherein the mouse expresses more immunoglobul in light chain comprising a human Vλ region than immunoglobulin light chain comprising a mouse Vλ region. 
     
     
         27 . The method of  claim 2 , wherein said IgL locus functional to express an immunoglobulin light chain comprising a human λ variable region is homozygous. 
     
     
         28 . The method of  claim 2 , wherein said transgenic mouse comprises immunoglobulin IgM and IgD antibody comprising human λ variable regions. 
     
     
         29 . A method of treating a disease or condition in a human, the method comprising administering the subject said antibody, antigen-binding fragment thereof or said human IgL chain λ variable region obtained by the method of  claim 2 . 
     
     
         30 . The method of  claim 2 , wherein said genome further comprises an immunoglobulin heavy chain (IgH) locus comprising unrearranged human heavy chain gene segments comprising a plurality of each of human VH gene segments, human D segments and human JH segments, wherein said human VH, D and JH gene segments are at an endogenous IgH locus upstream of a constant region comprising an endogenous CH gene segment, said IgH locus being functional to express an IgH chain comprising a human VH. 
     
     
         31 . The method of  claim 29 , where said light chain constant region of said IgL locus of the genome of said transgenic mouse is a λ constant region.