Genetically modified t cell receptor mice

Abstract

The invention provides a genetically modified non-human animal that comprises in its genome unrearranged T cell receptor variable gene loci, as well as embryos, cells, and tissues comprising the same. Also provided are constructs for making said genetically modified non-human animal and methods of making the same. Various methods of using the genetically modified non-human animal are also provided.

1 . A genetically modified mouse whose genome comprises, at an endogenous T cell receptor (TCR) α locus and in operable linkage from 5′ to 3′:
 (i) at least one unrearranged human TCR Vα gene segment, 
 (ii) at least one unrearranged human TCR Jα gene segment, and 
 (iii) a mouse TCR constant α (Cα) gene sequence that encodes a mouse TCR Cα domain, 
 wherein the at least one unrearranged human TCR Vα gene segment and the at least one unrearranged human TCR Jα gene segment rearrange, in a T cell of the genetically modified mouse, to form a rearranged human TCR Vα/Jα sequence that encodes a human TCR α variable domain and that is operably linked to the mouse TCR Cα gene sequence such that the rearranged human TCR Vα/Jα sequence operably linked to the mouse TCR Cα gene sequence encode a chimeric TCR α polypeptide comprising the human TCR α variable domain operably linked to the mouse TCR Cα domain, and 
 wherein the mouse expresses the chimeric TCR α polypeptide on the surface of the T cell. 
 
     
     
         2 . The genetically modified mouse of  claim 1 , wherein:
 (i) the at least one unrearranged human TCR Vα gene segment replaces one or more endogenous TCR Vα gene segments, and   (ii) the at least one unrearranged human TCR Jα gene segment replaces one or more endogenous TCR Jα gene segments.   
     
     
         3 .- 7 . (canceled) 
     
     
         8 . The genetically modified mouse of  claim 2 , wherein T cells of the genetically modified mouse undergo thymic T cell development to produce CD4 and CD8 single positive T cells. 
     
     
         9 . The genetically modified mouse of  claim 2 , wherein the genetically modified mouse comprises a normal ratio of splenic CD3+ T cells to total splenocytes. 
     
     
         10 . The genetically modified mouse  claim 2 , wherein the genetically modified mouse generates a population of central and effector memory T cells to an antigen of interest. 
     
     
         11 . The genetically modified mouse of  claim 2 , wherein the at least one unrearranged human TCR Vα gene segment comprises a complete repertoire of unrearranged human TCR Vα gene segments and the at least one unrearranged human TCR Jα gene segment comprises a complete repertoire of unrearranged human TCR Jα gene segments. 
     
     
         12 .- 14 . (canceled) 
     
     
         15 . A genetically modified mouse whose genome comprises, at an endogenous TCR β locus and in operable linkage from 5′ to 3′:
 (i) at least one unrearranged human TCR Vβ gene segment, 
 (ii) at least one unrearranged human TCR Dβ gene segment, 
 (iii) at least one unrearranged human TCR Jβ gene segment, and 
 (iv) a mouse TCR constant β (Cβ) gene sequence that encodes a mouse TCR Cβ domain; 
 wherein the at least one unrearranged human TCR Vβ gene segment, the at least one unrearranged human TCR Dβ gene segment and the at least one unrearranged human TCR J gene segment rearrange, in a T cell of the genetically modified mouse, to form a rearranged human TCR Vβ/Dβ/Jβ sequence that encodes a human TCR β variable domain and that is operably linked to the mouse TCR Cβ gene sequence such that the rearranged human TCR Vβ/Dβ/Jβ sequence operably linked to the mouse TCR Cβ gene sequence encode a chimeric TCR polypeptide comprising the human TCR β variable domain operably linked to the mouse TCR CP domain, and 
 wherein the mouse expresses the chimeric TCR β polypeptide on the surface of the T cell. 
 
     
     
         16 . The genetically modified mouse of  claim 15 , wherein:
 (i) the at least one unrearranged human TCR Vβ gene segment replaces one or more endogenous TCR Vβ gene segments,   (ii) the at least one unrearranged human TCR Dβ gene segment replaces one or more endogenous TCR Dβ gene segments, and   (iii) the at least one unrearranged human TCR Jβ gene segment replaces one or more endogenous TCR Jβ gene segments.   
     
     
         17 .- 21 . (canceled) 
     
     
         22 . The genetically modified mouse of  claim 16 , wherein T cells of the genetically modified mouse undergo thymic T cell development to produce CD4 and CD8 single positive T cells. 
     
     
         23 . The genetically modified mouse of  claim 16 , wherein the genetically modified mouse comprises a normal ratio of splenic CD3+ T cells to total splenocytes. 
     
     
         24 . The genetically modified mouse of  claim 16 , wherein the genetically modified mouse generates a population of central and effector memory T cells to an antigen of interest. 
     
     
         25 . The genetically modified mouse of  claim 16 , wherein the at least one unrearranged human TCR Vβ gene segment comprises a complete repertoire of unrearranged human Vβ gene segments, the at least one unrearranged human Dβ gene segment comprises a complete repertoire of unrearranged human Dβ segments, and the at least one unrearranged human TCR Jβ gene segment comprises a complete repertoire of unrearranged human TCR Jβ gene segments. 
     
     
         26 .- 28 . (canceled) 
     
     
         29 . A genetically modified mouse whose genome comprises:
 (a) unrearranged human TCR Vα gene segments that replace endogenous TCR Vα gene segments and unrearranged human TCR Jα gene segments that replace endogenous TCR Jα gene segments,   wherein the unrearranged human TCR Vα gene segments and the unrearranged human TCR Jα gene segments are operably linked to each other and an endogenous TCR Cα gene sequence that encodes an endogenous TCR Cα domain,   wherein the unrearranged human TCR Vα gene segments and the unrearranged human TCR Jα gene segments rearrange, in a T cell in the genetically modified mouse, to form a rearranged human TCR Vα/Jα sequence that encodes a human TCR α variable domain and that is operably linked to the endogenous TCR Cα gene sequence such that rearranged human TCR Vα/Jα sequence operably linked to the endogenous TCR Cα gene sequence encode a chimeric TCR α polypeptide comprising the human TCR α variable domain operably linked to the endogenous TCR Cα domain; and,   (b) unrearranged human TCR Vβ gene segments that replace endogenous TCR Vβ gene segments, unrearranged human TCR Dβ gene segments that replace endogenous TCR Dβ gene segments, and at least one unrearranged human TCR Jβ gene segments that replace endogenous TCR Jβ gene segments,   wherein the unrearranged human TCR Vβ gene segments, the unrearranged human TCR Dβ gene segments, and the unrearranged human TCR Jβ gene segments are operably linked to each other and an endogenous TCR Cβ gene sequence that encodes an endogenous TCR CP domain,   wherein the unrearranged human TCR Vβ gene segments, the unrearranged human TCR Dβ gene segments and the unrearranged human TCR Jβ gene segments rearrange, in the T cell in the genetically modified mouse, to form a rearranged human TCR Vβ/Dβ/Jβ sequence that encodes a human TCR β variable domain and that is operably linked to the endogenous TCR gene sequence such that the rearranged human TCR Vβ/Dβ/Jβ sequence operably linked to the endogenous TCR Cβ gene sequence encode a chimeric TCR β polypeptide comprising the human TCR δ variable domain operably linked to the endogenous TCR Cβ domain, and   wherein the mouse expresses, on the surface of the T cell, a T cell receptor comprising the chimeric TCR α polypeptide and the chimeric TCR β polypeptide.   
     
     
         30 .- 35 . (canceled) 
     
     
         36 . The genetically modified mouse of  claim 29 , wherein T cells of the genetically modified mouse undergo thymic T cell development to produce CD4 and CD8 single positive T cells. 
     
     
         37 . The genetically modified mouse of  claim 29 , wherein the genetically modified mouse comprises a normal ratio of splenic CD3+ T cells to total splenocytes. 
     
     
         38 . The genetically modified mouse of  claim 29 , wherein the genetically modified mouse generates a population of central and effector memory T cells to an antigen of interest. 
     
     
         39 .- 68 . (canceled)