Humanized C3 animals

Patent metadata

Field	Value
Publication number	US-9795121-B2
Application number	US-201514703818-A
Country	US
Kind code	B2
Filing date	May 4, 2015
Priority date	May 5, 2014
Publication date	Oct 24, 2017
Grant date	Oct 24, 2017

Abstract

Non-human animals comprising a human or humanized C3 and/or C5 nucleic acid sequence are provided as well as methods for using the same to identify compounds capable of modulating the complement system. Non-human animals that comprise a replacement of the endogenous C5 gene and/or C3 gene with a human or humanized C5 gene and/or C3 gene, and methods for making and using the non-human animals, are described. Non-human animals comprising a human or humanized C5 gene under control of non-human C5 regulatory elements is also provided, including non-human animals that have a replacement of non-human C5-encoding sequence with human C5-encoding sequence at an endogenous non-human C5 locus. Non-human animals comprising a human or humanized C3 gene under control of non-human C3 regulatory elements is also provided, including non-human animals that have a replacement of non-human C3 protein-encoding sequence with human or humanized C3 protein-encoding sequence at an endogenous non-human C3 locus. Non-human animals comprising human or humanized C3 and/or C5 sequences, wherein the non-human animals are rodents, e.g., mice or rats, are provided.

First claim

The invention claimed is:

1. A rodent, comprising a replacement at an endogenous rodent C3 locus of a rodent gene sequence encoding an exon of a C3 gene with a nucleic acid sequence encoding at least one exon of a human C3 gene to form a modified C3 gene, wherein expression of the modified C3 gene is under control of rodent regulatory elements at the endogenous rodent C3 locus, wherein the rodent is a mouse or a rat.

2. The rodent of claim 1 , wherein the human C3 gene encoding human C3 protein comprises exon 1 through exon 41 of the human C3 gene.

3. The rodent of claim 1 , wherein the human C3 gene encoding human C3 protein comprises exon 2 through exon 41 of the human C3 gene.

4. The rodent of claim 1 , wherein the rodent is a mouse that is incapable of expressing a mouse C3 protein.

5. The rodent of claim 1 , wherein the rodent is a mouse that expresses a mouse C5 protein encoded by an endogenous mouse C5 gene.

6. The rodent of claim 4 , wherein the mouse expresses a human or humanized C5 protein.

7. The rodent of claim 6 , wherein the mouse comprises a replacement at an endogenous mouse C5 locus of a mouse gene encoding C5 protein with a human C5 gene encoding human C5 protein.

8. A method for making a humanized rodent, comprising replacing a rodent C3 gene sequence encoding rodent C3 protein with a human C3 gene sequence comprising one or more exons of the human C3 gene sequence encoding human or humanized C3 protein, wherein the replacement is at an endogenous rodent C3 locus and the human C3 gene sequence comprising one or more exons of the human C3 gene sequence encoding human or humanized C3 protein is operably linked to rodent regulatory elements or sequences at the endogenous rodent C3 locus, wherein the rodent is a mouse or a rat.

9. The method of claim 8 , wherein the rodent regulatory elements or sequences are endogenous rodent regulatory elements or sequences at the rodent C3 locus.

10. The method of claim 8 , wherein the human C3 gene sequence replacing the rodent C5 gene sequence comprises at least one exon of the human C3 gene sequence.

11. The method of claim 8 , wherein the human C3 gene sequence replacing the rodent C3 gene sequence comprises all 41 exons of the human C3 gene sequence.

12. The method of claim 8 , wherein the replacement is at an endogenous rodent C3 locus and the human C3 gene sequence comprising one or more exons of the human C3 gene sequence encoding human or humanized C3 protein is operably linked to endogenous rodent regulatory elements or sequences at the endogenous rodent C3 locus.

13. The rodent of claim 7 , wherein expression of the human C5 gene is under control of mouse regulatory elements at the endogenous mouse C5 locus.

14. The rodent of claim 1 , wherein the rodent is a mouse that expresses human C3 protein in serum at a concentration of at least about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 110%, 120%, 130%, 140%, 150%, 160%, 170%, 180%, 190% or 200% of the level of mouse C3 protein present in the serum of an age-matched mouse that expresses functional endogenous mouse C3 protein, but does not comprise said replacement.

15. The rodent of claim 1 , wherein the rodent is a mouse that expresses human C3 protein in serum at a concentration of between about 10% and about 200%, between about 20% and about 150%, or between about 30% and about 100% of the level of mouse C3 protein present in the serum of an age-matched mouse that expresses functional endogenous mouse C3 protein, but does not comprise said replacement.

16. The rodent of claim 1 , wherein the rodent is a mouse that expresses human C3 protein in serum at a concentration of between about 100 μg ml and about 1500 μg/ml, between about 200 μg/ml, and about 1250 μg/ml, or between about 300 μg/ml and about 1000 μg/ml.

17. The rodent of claim 1 , wherein the rodent is a mouse that expresses human C3 protein in serum at a concentration of at least about 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1250 or 1500 μg/ml.

18. The rodent of claim 1 , wherein the human C3 protein is able to cross-react with murine C5 protein to recapitulate complement activity.

19. The rodent of claim 2 , wherein the human C3 protein is able to cross-react with murine C5 protein to recapitulate complement activity.

20. The rodent of claim 3 , wherein the human C3 protein is able to cross-react with murine C5 protein to recapitulate complement activity.

21. The rodent of claim 1 , wherein a portion of the endogenous mouse C3 locus, including 5′ regulatory elements upstream of exon 1 through exon 41, is deleted and replaced with a human C3 gene sequence comprising 5′ regulatory elements upstream of exon 1 through exon 41 of the human C3 gene.

22. The rodent of claim 1 , further comprising replacement of the 3′ untranslated region of the endogenous rodent C3 locus with the 3′ untranslated region of the human C3 gene.

23. The rodent of claim 21 , further comprising replacement of the 3′ untranslated region of the endogenous rodent C3 locus with the 3′ untranslated region of the human C3 gene.

Assignees

• Regeneron Pharma

Inventors

• Hu Ying
• Latuszek Adrianna
• Cao Jingtai
• Mujica Alexander
• Wiegand Stanley
• Mcwhirter John
• Murphy Andrew
• Macdonald Lynn

Classifications

• A01K67/0278 Primary

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

• C12N15/8509

  for producing genetically modified animals, e.g. transgenic · CPC title

• A01K2267/03

  Animal model, e.g. for test or diseases · CPC title

• C07K14/472

  Complement proteins, e.g. anaphylatoxin, C3a, C5a · CPC title