Genetically-modified T cells comprising a modified intron in the T cell receptor alpha gene

Patent metadata

Field	Value
Publication number	US-12448613-B2
Application number	US-202117330365-A
Country	US
Kind code	B2
Filing date	May 25, 2021
Priority date	Jun 30, 2017
Publication date	Oct 21, 2025
Grant date	Oct 21, 2025

Abstract

The present invention provides a genetically-modified T cell comprising in its genome a modified human T cell receptor alpha gene. The modified T cell receptor alpha gene comprises an exogenous sequence of interest inserted into an intron within the T cell receptor alpha gene that is positioned 5′ upstream of TRAC exon 1. The exogenous sequence of interest can comprise an exogenous splice acceptor site and/or a poly A signal, which disrupts expression of the T cell receptor alpha subunit. The sequence of interest can also include a coding sequence for a polypeptide, such as a chimeric antigen receptor. Additionally, the endogenous splice donor site and the endogenous splice acceptor site flanking the intron are unmodified and/or remain functional. The invention further provides compositions and methods for producing the genetically-modified cell, and populations of the cell, and methods for the treatment of a disease, such as cancer, using such cells.

First claim

What is claimed is:

1. An isolated genetically-modified human T cell comprising in its genome a modified human T cell receptor alpha gene, wherein said modified human T cell receptor alpha gene comprises an exogenous sequence of interest inserted into an intron within the human T cell receptor alpha gene that is positioned 5′ upstream of the T cell receptor alpha constant region (TRAC) exon 1, and wherein said exogenous sequence of interest comprises an exogenous splice acceptor site or an exogenous splice acceptor site and a poly A signal, and wherein an endogenous splice donor site and an endogenous splice acceptor site flanking said intron are unmodified, and wherein said genetically-modified human T cell does not express an endogenous T cell receptor on the cell surface.

2. The isolated genetically-modified human T cell of claim 1 , wherein said intron comprises SEQ ID NO: 3.

3. The isolated genetically-modified human T cell of claim 1 , wherein said exogenous sequence of interest comprises, from 5′ to 3′, an exogenous splice acceptor site, a 2A element or IRES element, a coding sequence for a protein of interest, and a polyA signal.

4. The isolated genetically-modified human T cell of claim 3 , wherein said 2A element is a T2A element.

5. The isolated genetically-modified human T cell of claim 1 , wherein said sequence of interest comprises a coding sequence for a chimeric antigen receptor or an exogenous T cell receptor.

6. The isolated genetically-modified human T cell of claim 1 , wherein said exogenous sequence of interest is inserted into said intron at an engineered meganuclease recognition site.

7. The isolated genetically-modified human T cell of claim 1 , wherein said exogenous sequence of interest comprises, from 5′ to 3′, an exogenous splice acceptor site, a 2A element or IRES element, a coding sequence for a protein of interest, and a polyA signal, and wherein said sequence of interest comprises a coding sequence for a chimeric antigen receptor or an exogenous T cell receptor.

8. The isolated genetically-modified human T cell of claim 7 , wherein said exogenous sequence of interest is inserted into said intron at an engineered meganuclease recognition site.

9. An isolated population of genetically-modified human T cells comprising a plurality of said isolated genetically-modified human T cell of claim 1 .

10. An isolated population of genetically-modified human T cells comprising a plurality of said isolated genetically-modified human T cell of claim 3 .

11. An isolated population of genetically-modified human T cells comprising a plurality of said isolated genetically-modified human T cell of claim 5 .

12. An isolated population of genetically-modified human T cells comprising a plurality of said isolated genetically-modified human T cell of claim 7 .

13. A pharmaceutical composition useful for treatment of a disease in a subject in need thereof, wherein said pharmaceutical composition comprises a pharmaceutically-acceptable carrier and said isolated genetically-modified human T cell of claim 1 .

14. A pharmaceutical composition useful for treatment of a disease in a subject in need thereof, wherein said pharmaceutical composition comprises a pharmaceutically-acceptable carrier and said isolated genetically-modified human T cell of claim 3 .

15. A pharmaceutical composition useful for treatment of a disease in a subject in need thereof, wherein said pharmaceutical composition comprises a pharmaceutically-acceptable carrier and said isolated genetically-modified human T cell of claim 5 .

16. A pharmaceutical composition useful for treatment of a disease in a subject in need thereof, wherein said pharmaceutical composition comprises a pharmaceutically-acceptable carrier and said isolated genetically-modified human T cell of claim 7 .

17. A method of treating a disease in a subject in need thereof, said method comprising administering to said subject said isolated genetically-modified human T cell of claim 1 .

18. A method of treating a disease in a subject in need thereof, said method comprising administering to said subject said isolated genetically-modified human T cell of claim 3 .

19. A method of treating a disease in a subject in need thereof, said method comprising administering to said subject said isolated genetically-modified human T cell of claim 5 .

20. A method of treating a disease in a subject in need thereof, said method comprising administering to said subject said isolated genetically-modified human T cell of claim 7 .

Assignees

• Prec Biosciences Inc

Inventors

• Jantz Derek
• Smith James Jefferson
• Beard Clayton

Classifications

• A61K40/40

  characterised by antigens that are targeted or presented by cells of the immune system · CPC title

• A61K40/32

  T-cell receptors [TCR] · CPC title

• A61K40/31

  Chimeric antigen receptors [CAR] · CPC title

• A61K40/11

  T-cells, e.g. tumour infiltrating lymphocytes [TIL] or regulatory T [Treg] cells; Lymphokine-activated killer [LAK] cells · CPC title

• C12N2800/60

  Vectors containing traps for, e.g. exons, promoters · CPC title