Genetically modified non-human animals and methods of use thereof

Patent metadata

Field	Value
Publication number	US-11778995-B2
Application number	US-202017000140-A
Country	US
Kind code	B2
Filing date	Aug 21, 2020
Priority date	Nov 5, 2012
Publication date	Oct 10, 2023
Grant date	Oct 10, 2023

Abstract

Genetically modified non-human animals are provided that may be used to model human hematopoietic cell development, function, or disease. The genetically modified non-human animals comprise a nucleic acid encoding human IL-6 operably linked to an IL-6 promoter. In some instances, the genetically modified non-human animal expressing human IL-6 also expresses at least one of human M-CSF, human IL-3, human GM-CSF, human SIRPa or human TPO. In some instances, the genetically modified non-human animal is immunodeficient. In some such instances, the genetically modified non-human animal is engrafted with healthy or diseased human hematopoietic cells. Also provided are methods for using the subject genetically modified non-human animals in modeling human hematopoietic cell development, function, and/or disease, as well as reagents and kits thereof that find use in making the subject genetically modified non-human animals and/or practicing the subject methods.

First claim

The invention claimed is:

1. A method of screening a candidate agent for the ability to treat a hematopoietic cancer, the method comprising,
contacting a first genetically modified, immunodeficient mouse with a candidate agent, wherein the first genetically modified, immunodeficient mouse is a Rag-2 −/− IL-2rg null mouse comprising
a genome comprising a nucleic acid encoding human IL-6, wherein the nucleic acid encoding human IL-6 replaces the endogenous mouse IL-6 gene at the endogenous mouse IL-6 gene locus, and wherein the nucleic acid encoding human IL-6 is operably linked to the endogenous mouse IL-6 promoter, and
an engraftment of human hematopoietic cancer cells; and

comparing the viability and/or proliferative rate of human hematopoietic cancer cells in the contacted first genetically modified, immunodeficient mouse to the viability and/or proliferative rate of human hematopoietic cancer cells in a second genetically modified, immunodeficient mouse not contacted with the candidate agent, wherein the second genetically modified, immunodeficient mouse is a Rag-2 −/− IL-2rg null mouse comprising
a genome comprising a nucleic acid encoding human IL-6, wherein the nucleic acid encoding human IL-6 replaces the endogenous mouse IL-6 gene at the endogenous mouse IL-6 gene locus, and wherein the nucleic acid encoding human IL-6 is operably linked to the endogenous mouse IL-6 promoter, and
an engraftment of human hematopoietic cancer cells,

wherein a decrease in the viability and/or rate of proliferation of the human hematopoietic cancer cells in the contacted first genetically modified,
immunodeficient mouse is indicative of the ability of the candidate agent to treat the hematopoietic cancer.

2. The method according to claim 1 , wherein the hematopoietic cancer is multiple myeloma.

3. The method according to claim 1 , wherein the first and second genetically modified, immunodeficient mice do not express mouse IL-6.

4. The method according to claim 1 , wherein the genomes of the first and second genetically modified mice comprise at least one additional nucleic acid selected from the group consisting of:
i) a nucleic acid encoding human SIRPa operably linked to a SIRPa promoter, wherein the first and second genetically modified, immunodeficient mice express a human SIRPa polypeptide;
ii) a nucleic acid encoding human M-CSF operably linked to a M-CSF promoter, wherein the first and second genetically modified, immunodeficient mice express a human M-CSF polypeptide;
iii) a nucleic acid encoding human IL-3 operably linked to an IL-3 promoter, wherein the first and second genetically modified, immunodeficient mice express a human IL-3 polypeptide;
iv) a nucleic acid encoding human GM-CSF operably linked to a GM-CSF promoter, wherein the first and second genetically modified, immunodeficient mice express a human GM-CSF polypeptide; and
v) a nucleic acid encoding human TPO operably linked to a TPO promoter, wherein the first and second genetically modified, immunodeficient mice express a human TPO polypeptide.

5. The method according to claim 4 , wherein the nucleic acid encoding human SIRPa is randomly inserted into the genomes, the nucleic acid encoding human M-CSF is at the rodent M-CSF locus, the nucleic acid encoding human IL-3 is at the rodent IL-3 locus, the nucleic acid encoding human GM-CSF is at the rodent GM-CSF locus, and the nucleic acid encoding human TPO is at the rodent TPO locus.

6. The method according to claim 5 , wherein the first and second genetically modified, immunodeficient mice do not express mouse IL-6.

7. The method according to claim 1 , wherein the genomes of the first and second genetically modified, immunodeficient mice comprise a nucleic acid encoding human SIRPa or a nucleic acid encoding a fusion protein comprising a biologically active fragment of a full length human SIRPA polypeptide randomly inserted into the genomes, and wherein the nucleic acid is operably linked to a promoter.

8. The method according to claim 7 , wherein the promoter is a mouse SIRPa promoter.

9. The method according to claim 7 , wherein the first and second genetically modified, immunodeficient mice do not express mouse IL-6.

10. The method according to claim 1 , wherein the candidate agent is a polypeptide.

11. The method according to claim 10 , wherein the polypeptide is an antibody.

Assignees

• Regeneron Pharma
• Univ Yale
• Institute For Res In Biomedicine Irb

Inventors

• Flavell Richard
• Strowig Till
• Manz Markus G
• Borsotti Chiara
• Dhodapkar Madhav
• Murphy Andrew J
• Stevens Sean
• Yancopoulos George D

Classifications

• C12N5/10

  Cells modified by introduction of foreign genetic material · CPC title

• A61K49/0008

  Screening agents using (non-human) animal models or transgenic animal models or chimeric hosts, e.g. Alzheimer disease animal model, transgenic model for heart failure · CPC title

• A01K67/0278 Primary

  Knock-in vertebrates, e.g. humanised vertebrates · CPC title

• C07K14/524

  Thrombopoietin, i.e. C-MPL ligand · CPC title

• A01K67/0276 Primary

  Knock-out vertebrates · CPC title