IP Library › Granted Patent US 12,258,557
Granted Patent B2
US 12,258,557 · App. 18/525,460 · Granted Mar 25, 2025

Humanized cell line

Inventors: Yoshihiro Kawaoka (Middleton, WI); Kosuke Takada (Tokyo, JP); Masaki Imai (Kawasaki, JP)
Assignees: Wisconsin Alumni Research Foundation (WARF); The University of Tokyo
C12N15/102C12N5/0602C12N9/10C12Q1/04C12Y204/99001
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Quick Facts
Patent No.
US 12,258,557
App. No.
18/525,460
Granted
Mar 25, 2025
Kind
B2
Abstract

A mammalian or avian cell line that expresses high levels of human influenza virus receptors is provided. In one embodiment, the cell line supports human influenza virus, e.g., human A/H3 influenza virus, isolation and growth much more effectively than corresponding conventional (unmodified) cells or in corresponding human virus receptor-overexpressing cells, and the propagated viruses may maintain higher genetic stability than in the corresponding cells.

Claims (22)

1. An isolated recombinant mammalian cell, comprising a reduced amount of cell surface β-galactoside α2,3 sialyl residues relative to a corresponding non-recombinant mammalian cell, wherein β-galactoside α2,3 sialyltransferase (ST3Gal) genes are mutated so as to reduce the amount of the cell surface β-galactoside α2,3 sialyl residues, wherein the ST3Gal genes include ST3Gal-I, ST3Gal-II, ST3Gal-III, ST3Gal-IV, ST3Gal-V, ST3Gal-VI, and ST3Gal-II-like genes, and wherein the recombinant mammalian cell comprises an expression cassette encoding human β-galactoside α2,6 sialyltransferase (ST6Gal), wherein the reduced amount of cell surface 3-galactoside a2,3 sialyl residues is the result of mutation of the ST3Gal genes in the recombinant cell.

2. The isolated recombinant mammalian cell of claim 1 , further comprising an increased amount of human β-galactoside α2,6 sialyl residues relative to a corresponding non-recombinant mammalian cell.

3. The isolated recombinant mammalian cell of claim 1 , which comprises an expression cassette encoding human β-galactoside α2,6 sialyltransferase I (ST6Gal-I) or II (ST6Gal-II).

4. A method of modifying the amount of cell surface β-galactoside α2,3 sialyl residues and human β-galactoside α2,6 sialyl residues on a mammalian cell, comprising:

mutating β-galactoside α2,3 sialyltransferase (ST3Gal) genes, wherein the ST3Gal genes include ST3Gal-I, ST3Gal-II, ST3Gal-III, ST3Gal-IV, ST3Gal-V, ST3Gal-VI, and ST3Gal-II-like genes, and

overexpressing a human β-galactoside α2,6 sialyltransferase (ST6Gal) gene, in a parental mammalian cell so as to result in a modified mammalian cell having a reduced amount of cell surface β-galactoside α2,3 sialyl residues and an increased amount of human p β-galactoside α2,6 sialyl residues on the surface of the modified cell relative to the corresponding parental mammalian cell.

5. The method of claim 4 , wherein the mutations include one or more nucleotide insertions or one or more nucleotide deletions, or both, in the ST3Gal genes.

6. The method of claim 4 wherein the modified cell comprises an expression cassette comprising a ST6Gal open reading frame.

7. A method of detecting or propagating an influenza virus, comprising:

infecting the recombinant mammalian cell of claim 2 with a sample having or suspected of having an influenza virus.

8. The method of claim 7 , further comprising collecting progeny virus.

9. The method of claim 7 , wherein the sample is from an avian or a mammal suspected of being infected with an influenza virus.

10. The method of claim 7 , wherein the influenza virus is a human influenza virus.

11. The method of claim 7 , wherein the influenza virus is an influenza A or B virus.

12. The method of claim 7 , wherein the influenza virus is a H3 virus.

13. The method of claim 7 , wherein the influenza virus is A/H1N1, A/H3N2, a B/Yamagata-lineage influenza B virus or a B/Victoria-lineage influenza B virus.

14. The method of claim 7 , further comprising detecting whether the sample is infected with an influenza virus.

15. The method of claim 14 , further comprising identifying the HA and/or NA subtype of the virus.

16. The isolated recombinant mammalian cell of claim 1 , which is infected with human influenza virus.

17. The isolated recombinant mammalian cell of claim 1 , wherein the mammalian cell is a canine, nonhuman primate cell or a rodent cell.

18. The isolated recombinant mammalian cell of claim 17 , wherein the nonhuman primate cell is a chimpanzee, ape or monkey cell.

19. The isolated recombinant mammalian cell of claim 17 , wherein the rodent cell is a mouse, rat and guinea pig cell.

Assignments (2)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Jul 1, 2024
From: KAWAOKA, YOSHIHIRO
To: WISCONSIN ALUMNI RESEARCH FOUNDATION (WARF); THE UNIVERSITY OF TOKYO
Reel/Frame 067882/0930 →
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Jul 1, 2024
From: TAKADA, KOSUKE; IMAI, MASAKI
To: THE UNIVERSITY OF TOKYO
Reel/Frame 067883/0788 →
Continuity (3)
Continuation 16785449 · Feb 7, 2020
Provisional Application 62803266 · Feb 8, 2019
Related Publication 20240318167A1 · Sep 26, 2024
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