Gamma delta T cells derived from induced pluripotent stem cells, and production method therefor

Patent metadata

Field	Value
Publication number	US-12576149-B2
Application number	US-202519019930-A
Country	US
Kind code	B2
Filing date	Jan 14, 2025
Priority date	Feb 5, 2021
Publication date	Mar 17, 2026
Grant date	Mar 17, 2026

Abstract

Provided is a γδ T cell for securing the purity and number of cells sufficient for treatment. Also provided is a method of generating the γδ T cell. More specifically, provided are homogeneous γδ T cells excellent in that the γδ T cells are not affected by exhaustion of the cells. The foregoing is achieved by γδ T cells obtained by subjecting induced pluripotent stem cells (iPS cells) to differentiation induction treatment. Specifically, the foregoing is achieved by γδ T cells generated by subjecting iPS cells having a rearranged γδ TCR gene (γδ TCR-type iPS cells) to differentiation induction treatment. According to the method of generating the γδ T cell of the present invention, there can be provided γδ T cells and a cell population of γδT cells that have an excellent function of having antigen-specific cytotoxic activity in an MHC-unrestricted manner, and that are more homogeneous and have a higher effect than γδ T cells separated from peripheral blood.

First claim

The invention claimed is:

1 . A method of generating an iPS cell-derived γδ T cell, comprising:
(a) culturing a hematopoietic progenitor cell in a medium obtained by supplementing a basal medium with one kind or a plurality of kinds selected from FMS-like tyrosine kinase 3 ligand (FLT3L), stem cell factor (SCF), IL-2, IL-7, thrombopoietin (TPO), and L-ascorbic acid,
wherein the hematopoietic progenitor cell is obtained by differentiation induction treatment of an iPS cell having a rearranged γδ TCR gene; and
(b) culturing the cell resulted from step (a) in a medium containing a γδ T cell stimulant, thereby generating an iPS cell-derived γδ T cell,
wherein the γδ T cell stimulant is a phosphoric acid compound or a derivative thereof, which is a metabolite of an isoprenoid biosynthesis pathway, or a specific inhibitor of a farnesyl pyrophosphate (FPP) synthase serving as a rate-limiting enzyme of the isoprenoid biosynthesis pathway, and
wherein step (b) is initiated 17 to 31 days after the start of the differentiation induction treatment.

2 . The method according to claim 1 , wherein step (a) comprises co-culturing the hematopoietic progenitor cell with a feeder cell.

3 . The method according to claim 1 , wherein step (a) comprises culturing the hematopoietic progenitor cell without a feeder cell.

4 . The method according to claim 3 , wherein step (a) comprises culturing the hematopoietic progenitor cell using a culture substrate coated with: (i) vascular cell adhesion molecule-1 (VCAM1) and (ii) delta-like protein 4 (DLL4) or delta-like protein 1 (DLL1).

5 . The method according to claim 3 , wherein step (a) further comprises culturing the hematopoietic progenitor cell using a medium containing DKK1 and/or azelaic acid (AZA).

6 . The method according to claim 1 , wherein the medium containing a γδ T cell stimulant further comprises one kind or a plurality of kinds selected from IL-2 and IL-15.

7 . The method according to claim 1 , wherein step (a) is performed under a serum-free condition.

8 . The method according to claim 1 , wherein step (a) is performed under a hypoxic condition.

9 . A cell population, comprising the iPS cell-derived γδ T cell generated by the method according to claim 1 , wherein 10% or less of the γδ T cells in the cell population are undifferentiated iPS cells.

10 . The cell population according to claim 9 , having a higher cytotoxic activity in an antigen-specific manner than a cell population of γδ T cells separated from peripheral blood.

11 . The cell population according to claim 9 , wherein 90% or more of the iPS cell-derived γδ T cells in the cell population have base sequences identical to each other in a CDR3 region of a TCR gene.

12 . The cell population according to claim 11 , wherein the cell population comprises 1×10 5 or more iPS cell-derived γδ T cells.

13 . A cell population generated according to the method of claim 1 comprising IPS cell-derived γδ T cells, wherein 90% or more of the iPS cell-derived γδ T cells exhibit a higher expression amount of CD7 and/or CD8a than γδ T cells separated from peripheral blood.

14 . A method of antigen-specific cellular immunotherapy, comprising administering to a subject in need thereof a therapeutically effective amount of the iPS cell-derived γδ T cell generated according to the method of claim 1 .

15 . The method according to claim 1 , wherein the medium in step (a) contains a bead-like carrier coated with VCAM1 and DLL4.

16 . A method for treating a disease, comprising administering to a subject in need thereof a therapeutically effective amount of the cell population according to claim 9 , wherein the disease is selected from the group consisting of cancer, infectious diseases, and autoimmune disorders.

17 . A pharmaceutical composition, comprising the cell population according to claim 9 as an active ingredient.

18 . The method according to claim 3 , wherein step (a) is performed under a serum-free condition.

19 . The method according to claim 3 , wherein step (a) is performed under a hypoxic condition.

20 . The method according to claim 1 , wherein step (b) is initiated 17 to 24 days after the start of the differentiation induction treatment.

Assignees

• Univ Kobe Nat Univ Corp

Inventors

• Aoi Takashi
• Murai Nobuyuki

Classifications

• A61K40/42

  Cancer antigens · CPC title

• A61K40/32

  T-cell receptors [TCR] · CPC title

• A61K40/11

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

• C12N5/0636 Primary

  T lymphocytes · CPC title

• C12N5/0037

  Serum-free medium, which may still contain naturally-sourced components · CPC title