Antibody producing non-human animals

Abstract

Described are transgenic, non-human animals comprising a nucleic acid encoding an immunoglobulin light chain, whereby the immunoglobulin light chain is human, human-like, or humanized. The nucleic acid is provided with a means that renders it resistant to DNA rearrangements and/or somatic hypermutations. In one embodiment, the nucleic acid comprises an expression cassette for the expression of a desired molecule in cells during a certain stage of development in cells developing into mature B cells. Further provided is methods for producing an immunoglobulin from the transgenic, non-human animal.

1 . A method for producing a composition comprising at least two different antibodies or antigen-binding fragments thereof comprising paired variable regions, said at least two antibodies or antigen-binding fragments thereof having different binding specificities, comprising contacting at least one light chain variable region with at least two heavy chain variable regions under conditions allowing for pairing of light chain variable regions with heavy chain variable regions and harvesting antibodies or antigen-binding fragments thereof having binding specificities resulting from said pairing,
 wherein said contacting takes place inside a cell; and   said variable regions are produced by expression of nucleic acid sequences encoding said variable regions, and   wherein said nucleic acids encode heavy chain variable regions expressed by B cells of transgenic non-human animals that have been immunized with antigens,   wherein the genome of the transgenic animals comprises a transgene comprising a immunoglobulin light chain germline V gene segment fused to a immunoglobulin light chain germline J gene segment such that there is no mutation due to said fusion, wherein the fused V/J gene segments encode a rearranged immunoglobulin light chain variable region and wherein the transgene comprises a light chain constant region gene segment or is operatively linked to an endogenous light chain constant region gene segment and   wherein said transgenic animals produce populations of B cells producing antigen-specific antibodies, wherein said antibodies comprise the rearranged light chain immunoglobulin variable region encoded by the fused V/J gene segments paired with a diversity of heavy chain variable regions.   
     
     
         2 . The method of  claim 1 , wherein said light chain variable region upon pairing does not significantly contribute to the resulting binding specificity of the resulting antibodies or antigen-binding fragments thereof. 
     
     
         3 . The method of  claim 1  for producing a composition of antibodies or antigen binding fragments thereof comprising at least three different binding specificities. 
     
     
         4 . The method of  claim 1 , wherein at least two different heavy chain variable regions are expressed from one or more nucleic acids encoding these variable regions, whereby expression of said different variable regions is under the direction of different control elements. 
     
     
         5 . The method of  claim 4 , wherein said differential expression is different in levels of expression and/or time of expression. 
     
     
         6 . An expression system for carrying out the method of  claim 1 , comprising nucleic acids encoding said variable regions together with elements required for gene expression and pairing. 
     
     
         7 . Recombinant host cells that comprise the expression system of  claim 6 . 
     
     
         8 . The cells of  claim 7  which are eukaryotic cells. 
     
     
         9 . A method for identifying antibodies or antigen-binding fragments thereof having specific affinity for at least one target epitope, comprising contacting antibodies or antigen-binding fragments thereof produced by the cells of  claim 7  with said target epitope and identifying antibodies or antigen-binding fragments thereof showing said specific affinity. 
     
     
         10 . The method of  claim 9 , wherein antibodies or antigen-binding fragments thereof having specific affinity for at least two target epitopes, are identified. 
     
     
         11 . A composition comprising antibodies or antigen binding fragments thereof obtainable by the method of  claim 1 . 
     
     
         12 . The composition of  claim 11 , wherein said antibodies or antigen-binding fragments thereof are directed to at least two target antigens and/or two target epitopes. 
     
     
         13 . The composition of  claim 12 , wherein the target epitopes are associated with a disease or disorder. 
     
     
         14 . The composition of  claim 13 , wherein antibodies or antigen-binding fragments thereof are human antibodies. 
     
     
         15 . A pharmaceutical composition comprising the composition of  claim 13 . 
     
     
         16 . The composition of  claim 11  comprising at least one monospecific antibody and at least one bispecific antibody. 
     
     
         17 . The composition of  claim 11  comprising at least two monospecific antibodies. 
     
     
         18 . The composition of  claim 13 , wherein at least one target antigen is a protein or carbohydrate expressed on the surface of bacteria, viruses, or other pathogen or is an endogenous animal protein. 
     
     
         19 . The composition of  claim 18 , wherein the target antigen comprises two or more epitopes derived from or two or more targets selected from proteins and carbohydrates expressed on the surface of bacteria, viruses, or other pathogen or are endogenous animal proteins. 
     
     
         20 . The composition of  claim 18 , wherein the target antigen is an EGF receptor family member including erbB1 (EGF-R; HER 1), erbB2, (HER2), erbB3 (HER3) or erbB4 (HER4), or is c-Met (HGF-R or is a vascular endothelial growth factor receptor (VEGFR) family member or is VEGF, CD20, CD38, CD33, CEA, EpCAM, PSMA, CD19, CTLA4 or TGFbeta. 
     
     
         21 . The composition of  claim 19 , wherein the target antigen comprises two or more epitopes derived from or two or more targets which are selected from an EGF receptor family member including erbB1 (EGF-R; HER 1), erbB2, (HER2), erbB3 (HER3), and erbB4 (HER4); c-Met (HGF-R; a vascular endothelial growth factor receptor (VEGFR) family member; VEGF, CD20, CD38, CD33, CEA, EpCAM, PSMA, CD19, CTLA4 and TGFbeta.