Genetically modified non-human animals and methods of use thereof
The invention relates generally to genetically modified non-human animals expressing human polypeptides and their methods of use.
1. A genetically modified mouse comprising in its genome a recombination activating gene 2 (Rag-2) gene knock-out, an IL2 receptor gamma chain (IL2rg) gene knock-out, a replacement of a mouse M-CSF gene with a nucleic acid encoding a human M-CSF polypeptide at a mouse M-CSF gene locus, a replacement of a mouse IL-3 gene with a nucleic acid encoding a human IL-3 polypeptide at a mouse IL-3 gene locus, a replacement of a mouse GM-CSF gene with a nucleic acid encoding a human GM-CSF polypeptide at a mouse GM-CSF gene locus, an insertion of a nucleic acid encoding a human SIRPA polypeptide, and a replacement of a mouse TPO gene with a nucleic acid encoding a human TPO polypeptide at a mouse TPO gene locus, wherein each of the nucleic acids encoding the human M-CSF polypeptide, the human IL-3 polypeptide, the human GM-CSF polypeptide, the human SIRPA polypeptide, and the human TPO polypeptide is operably linked to a promoter, and wherein the mouse expresses the human M-CSF polypeptide, the human IL-3 polypeptide, the human GM-CSF polypeptide, the human SIRPA polypeptide, and the human TPO polypeptide.
2. The genetically modified mouse of claim 1 , further comprising human hematopoietic cells.
3. The genetically modified mouse of claim 1 , further comprising a human cancer cell.
4. The genetically modified mouse of claim 3 , wherein the human cancer cell is a leukemia cell or a melanoma cell.
5. A method of hematopoietic stem and progenitor cell (HSPC) engraftment comprising a step of administering at least one HSPC to the genetically modified mouse of claim 1 .
6. The method of claim 5 , wherein the genetically modified mouse comprises a human cancer cell.
7. The method of claim 6 , wherein the human cancer cell is a leukemia cell or a melanoma cell.
8. A genetically modified mouse comprising in its genome
a recombination activating gene 2 (Rag-2) gene knock-out,
an IL2 receptor gamma chain (IL2rg) gene knock-out,
a replacement of a mouse M-CSF gene with a nucleic acid encoding a human M-CSF polypeptide at a mouse M-CSF gene locus,
a replacement of a mouse IL-3 gene with a nucleic acid encoding a human IL-3 polypeptide at a mouse IL-3 gene locus,
a replacement of a mouse GM-CSF gene with a nucleic acid encoding a human GM-CSF polypeptide at a mouse GM-CSF gene locus,
an insertion of a nucleic acid encoding a human SIRPA polypeptide, and
a replacement of a mouse TPO gene with a nucleic acid encoding a human TPO polypeptide at a mouse TPO gene locus,
wherein each of the nucleic acids encoding the human M-CSF polypeptide, the human IL-3 polypeptide, the human GM-CSF polypeptide, the human SIRPA polypeptide, and the human TPO polypeptide is operably linked to a promoter,
wherein the mouse expresses the human M-CSF polypeptide, the human IL-3 polypeptide, the human GM-CSF polypeptide, the human SIRPA polypeptide, and the human TPO polypeptide,
and wherein, when engrafted with human CD34+ cells, the mouse exhibits a significantly higher proportion of human CD33+ myeloid cells in blood and bone marrow relative to a genetically modified mouse engrafted with human CD34+ cells which does not express the human M-CSF polypeptide, the human IL-3 polypeptide, the human GM-CSF polypeptide, the human SIRPA polypeptide, and the human TPO polypeptide.
9. The genetically modified mouse of claim 8 , wherein the mouse is engrafted with human CD34+ cells.
10. The method of claim 5 , wherein the administering step is not preceded by preconditioning irradiation of the genetically modified mouse.
11. A genetically modified mouse comprising in its genome
a recombination activating gene 2 (Rag-2) gene knock-out,
an IL2 receptor gamma chain (IL2rg) gene knock-out,
a replacement of a mouse M-CSF gene with a nucleic acid encoding a human M-CSF polypeptide at a mouse M-CSF gene locus,
a replacement of a mouse IL-3 gene with a nucleic acid encoding a human IL-3 polypeptide at a mouse IL-3 gene locus,
a replacement of a mouse GM-CSF gene with a nucleic acid encoding a human GM-CSF polypeptide at a mouse GM-CSF gene locus,
an insertion of a nucleic acid encoding a human SIRPA polypeptide, and
a replacement of a mouse TPO gene with a nucleic acid encoding a human TPO polypeptide at a mouse TPO gene locus,
wherein each of the nucleic acids encoding the human M-CSF polypeptide, the human IL-3 polypeptide, the human GM-CSF polypeptide, the human SIRPA polypeptide, and the human TPO polypeptide is operably linked to a promoter,
wherein the mouse expresses the human M-CSF polypeptide, the human IL-3 polypeptide, the human GM-CSF polypeptide, the human SIRPA polypeptide, and the human TPO polypeptide,
and wherein, when engrafted with human CD34+ cells and a human tumor, the mouse exhibits increased human myeloid cell infiltration of the engrafted tumor relative to a genetically modified mouse engrafted with human CD34+ cells and a human tumor, which does not express the human M-CSF polypeptide, the human IL-3 polypeptide, the human GM-CSF polypeptide, the human SIRPA polypeptide, and the human TPO polypeptide.
12. The genetically modified mouse of claim 11 , wherein the mouse is engrafted with human CD34+ cells and a human tumor.
13. The genetically modified mouse of claim 1 , wherein the human SIRPA polypeptide is a biologically active fragment of a full-length human SIRPA polypeptide.
14. The genetically modified mouse of claim 8 , wherein the human SIRPA polypeptide is a biologically active fragment of a full-length human SIRPA polypeptide.
15. The genetically modified mouse of claim 11 , wherein the human SIRPA polypeptide is a biologically active fragment of a full-length human SIRPA polypeptide.