Humanized T cell co-receptor mice
AssigneeRegeneron Pharmaceuticals, Inc.
InventorsLynn Macdonald, Andrew J. Murphy, Naxin Tu, Cagan Gurer
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Abstract
The invention provides genetically modified non-human animals that express chimeric human/non-human T cell co-receptor polypeptides (e.g., CD4, CD8α, CD8β), as well as embryos, cells, and tissues comprising the same. Also provided are constructs for making said genetically modified animals and methods of making the same.
Claims28
A genetically modified non-human animal comprising a first nucleotide sequence encoding a chimeric human/non-human CD8α polypeptide and a second nucleotide sequence encoding a chimeric human/non-human CD8β polypeptide,
- wherein the amino acid sequence of the human portion of the chimeric human/non-human CD8α polypeptide is set forth in SEQ ID NO:59 and is operably linked to non-human transmembrane and cytoplasmic domains of a non-human CD8α polypeptide,
- wherein the second nucleotide sequence comprises a nucleic acid sequence encoding a human CD8β polypeptide extracellular domain comprising at least the immunoglobulin V-like domain of the human CD8β polypeptide,
- wherein the nucleic acid sequence encoding the human CD8β polypeptide extracellular domain is operably linked to a nucleic acid sequence encoding transmembrane and cytoplasmic domains of an endogenous non-human CD8β polypeptide, and
- wherein the non-human animal expresses a chimeric human/non-human CD8 protein comprising the chimeric human/non-human CD8α and CD8β polypeptides.
A genetically modified mouse comprising:
- (i) a first nucleotide sequence that encodes a chimeric human/mouse CD8α polypeptide, wherein the amino acid sequence of the human portion of the chimeric human/mouse CD8α polypeptide is set forth in SEQ ID NO:59 and is operably linked to mouse transmembrane and cytoplasmic domains of a mouse CD8α polypeptide, and
- (ii) a second nucleotide sequence that comprises a nucleic acid sequence encoding a human CD8β polypeptide extracellular domain comprising at least the immunoglobulin V-like domain of the human CD8β polypeptide,
- wherein the nucleic acid sequence encoding the human CD8β polypeptide extracellular domain is operably linked to a nucleic acid sequence encoding transmembrane and cytoplasmic domains of a mouse CD8β polypeptide, and
- wherein the second nucleotide sequence encodes a chimeric human/mouse CD8β polypeptide;
- and
- wherein the mouse expresses a chimeric human/mouse CD8 protein comprising the chimeric human/mouse CD8α and CD8β polypeptides.
A method of modifying a mouse to express a chimeric human/mouse CD8 polypeptide comprising replacing at an endogenous mouse CD8 locus (i) a nucleotide sequence encoding a CD8α polypeptide extracellular domain with a first orthologous nucleotide sequence encoding the human amino acid sequence set forth in SEQ ID NO:59 and (ii) a nucleotide sequence encoding a CD8β polypeptide extracellular domain and comprising at least exons 2-3 of an endogenous mouse CD8β locus with a second orthologous nucleotide sequence encoding a human CD8β polypeptide extracellular domain.
A method for determining whether an antigen will generate an HLA class I-restricted immune response comprising
- (i) exposing a genetically modified non-human animal to the antigen, wherein the non-human animal comprises
- (a) a first nucleotide sequence that encodes a chimeric human/non-human CD8α polypeptide of which the amino acid sequence of the human portion is set forth in SEQ ID NO:59 and is operably linked to non-human transmembrane and cytoplasmic domains of a non-human CD8α polypeptide,
- (b) a second nucleotide sequence that comprises a nucleic acid sequence encoding a human/CD8β polypeptide extracellular domain comprising at least the immunoglobulin V-like domain of the human CD8β polypeptide,
- wherein the nucleic acid sequence encoding the CD8β polypeptide extracellular domain is operably linked to a nucleic acid sequence encoding transmembrane and cytoplasmic domains of a non-human CD8β polypeptide, and
- the second nucleotide sequence encodes a chimeric human/non-human CD8β polypeptide, and
- (c) a third nucleotide sequence comprising a sequence that encodes a human or humanized MHC I polypeptide, and
- wherein the non-human animal expresses a chimeric human/non-human CD8 protein comprising the chimeric human/non-human CD8α and CD8β polypeptides and the human or humanized MHC I polypeptide;
- and
- (ii) detecting the absence or presence of an antigen-specific HLA class I-restricted immune response in the non-human animal;
- wherein the presence of an antigen-specific HLA class I-restricted response determines that the antigen will generate an HLA class I-restricted immune response.
Dependencies are inferred from the claim text.