Immune cell organoid co-cultures

Patent metadata

Field	Value
Publication number	US-12517112-B2
Application number	US-201816954506-A
Country	US
Kind code	B2
Filing date	Dec 21, 2018
Priority date	Dec 21, 2017
Publication date	Jan 6, 2026
Grant date	Jan 6, 2026

Abstract

The present invention provides co-cultures of organoids and immune cells, and methods of using these to identify agents for treating diseases.

First claim

The invention claimed is:

1 . A method for identifying an agent suitable for treating a cancer, wherein the method comprises:
contacting a tumouroid co-culture with one or more agents, wherein the tumouroid co-culture comprises immune cells and at least one tumouroid, detecting the presence or absence of one or more change in the tumouroid co-culture that is indicative of agent suitability for treating the cancer, and identifying the agent suitable for treating the cancer if the presence or absence of one or more of said changes in the tumouroid co-culture is detected, wherein the method is preceded by: preparing the at least one tumouroid by culturing tumour epithelial cells in a tumouroid culture medium; preparing the immune cells by culturing the immune cells in an immune cell expansion medium, wherein the tumouroid culture medium comprises IL-2 and/or the immune cell expansion medium comprises IL-2; and preparing the tumouroid co-culture by mixing the at least one tumouroid with the immune cells in an in vitro culture.

2 . The method of claim 1 , wherein the method is preceded by one or more of the following steps:
preparing the immune cells by separating immune cells from an immune sample from a subject; and/or preparing the tumouroid co-culture by removing the tumouroid culture medium from the at least one tumouroid, and mixing the at least one tumouroid with the immune cells in a tumouroid co-culture medium.

3 . The method of claim 2 , wherein the tumour epithelial cells are obtained from a sample from a cancer patient.

4 . The method of claim 1 , wherein the agent suitable for treating the cancer is identified if the presence or absence of one or more said changes is detected in the tumouroid co-culture but not in a reference organoid co-culture or a reference tumouroid co-culture; and/or
wherein the method is preceded by one or more of the following steps: preparing at least one organoid by culturing normal epithelial cells in an organoid culture medium; and/or preparing the immune cells by separating immune cells from an immune sample from a subject, and culturing the immune cells in an immune cell expansion medium; and/or preparing the reference tumouroid co-culture or the reference organoid co-culture, optionally by removing the tumouroid culture medium or the organoid culture medium from the at least one tumouroid or the at least one organoid, and subsequently mixing the at least one reference organoid or the at least one reference tumouroid with the immune cells in an organoid co-culture medium or a tumouroid co-culture medium, optionally wherein the immune sample is a tumour sample, normal colon tissue and/or peripheral blood.

5 . The method of claim 4 , wherein the normal epithelial cells are autologous with the tumour epithelial cells.

6 . The method of claim 4 , wherein (i) the tumouroid co-culture medium and/or (ii) the reference organoid co-culture medium or the reference tumouroid co-culture medium, comprises extracellular matrix, optionally selected from collagen, or any animal-derived or synthetic basement membrane matrix, optionally wherein the collagen is rat tail collagen I.

7 . The method of claim 4 , wherein:
(a) the immune cells of the tumouroid co-culture have a motility of at least 40 μm/day; and/or (b) at least 20% of the immune cells in the tumouroid co-culture moves a distance of at least 200 μm in 80 hours; and/or (c) the immune cells remain active for at least 4 h; and/or (d) the one or more agents are of known suitability for treating cancer and the method further comprises identifying the one or more agents as suitable agents for treating cancer in a particular patient.

8 . The method of claim 7 , wherein both the tumouroid co-culture and the reference organoid co-culture or the reference tumouroid co-culture are derived from the particular patient.

9 . The method of claim 4 , wherein the immune cells:
(a) are allogeneic with the tumouroid and/or organoid, optionally wherein the immune cells and tumouroid and/or organoid are derived from either peripheral blood or tissue biopsy of a different patient or healthy control; and/or (b) are HLA matched with the at least one tumouroid and/or the at least one organoid.

10 . The method of claim 4 , wherein the at least one tumouroid and/or at least one organoid:
(a) comprises or consists of autologous cells; and/or (b) are separated into populations sharing one or more genotypes.

11 . The method of claim 4 , wherein:
(a) the at least one tumouroid or at least one organoid comprises or consists of mammalian cells or human cells; and/or (b) the tumouroid co-culture is cultured in (i) immune cell expansion medium or (ii) a 50:50 (v/v) mixture of immune cell expansion medium and tumouroid culture medium; and/or (c) the organoid co-culture is cultured in (i) immune cell expansion medium or (ii) a 50:50 (v/v) mixture of immune cell expansion medium and organoid culture medium; and/or (d) the reference organoid co-culture or reference tumouroid co-culture is cultured for at least 4 h.

12 . The method of claim 4 , wherein the tumour epithelial cells or the normal epithelial cells are obtained from a cancer patient; or the tumour epithelial cells and the normal epithelial cells are obtained from different cancer patients; or the tumour epithelial cells and the normal epithelial cells are obtained from a single cancer patient, optionally from the same sample.

13 . The method of claim 1 , wherein the immune cells:
(a) comprise one or more cell types selected from the group consisting of intra-epithelial lymphocytes (IELs), tumour-infiltrating lymphocytes (TILs), peripheral blood mononuclear cells (PBMCs), peripheral blood lymphocytes (PBLs), T cells, and cytotoxic T lymphocytes (CTLs), αβ T cells, γδ T cells, B cells, NK cells, and mononuclear phagocytes; and/or (b) are obtained from a sample from a cancer patient; and/or (c) are obtained from a peripheral blood sample and/or a tissue biopsy; and/or (d) are obtained from the same patient as the tumour epithelial cells; and/or (e) are allogeneic with the at least one tumouroid, optionally wherein the immune cells and the at least one tumouroid are derived from either peripheral blood or tissue biopsy of a different patient or healthy control; and/or (f) are HLA matched with the at least one tumouroid; and/or (g) are cultured in the immune cell expansion medium for at least 4 h.

14 . The method of claim 1 , wherein the at least one tumouroid:
(a) comprises or consists of autologous cells; and/or (b) are separated into populations sharing one or more genotypes, phenotypes, and/or epigenetic markers, prior to the mixing with immune cells.

15 . The method of claim 1 , wherein:
(a) the at least one tumouroid comprises or consists of mammalian cells or human cells; and/or (b) the at least one tumouroid co-culture is cultured in immune cell expansion medium or in a 50:50 (v/v) mixture of immune cell expansion medium and tumouroid culture medium; and/or (c) the tumouroid co-culture is cultured for at least 4 h.

16 . The method of claim 1 , wherein: (a) the tumouroid culture medium comprises IL-2 and the immune cell expansion medium comprises IL 2; and/or (b) the tumouroid co-culture comprises IL-2.

17 . The method of claim 1 , wherein the immune cells are antigenically compatible with a patient from whom the at least one tumouroid is derived.

18 . A method of testing a CAR-T immunotherapy, TCR transgenic T cells, neoantigen, or checkpoint inhibitor, for eff

Assignees

• Koninklijke Nederlandse Akademie Van Wetenschappen
• Hub Organoids Ip B V

Inventors

• Kretzschmar Kai
• Bar-Ephraim Jotam Elazar
• Clevers Johannes Carolus
• Fernandez-Boj Sylvia
• Vries Robert Gerhardus Jacob

Classifications

• A61K40/4243

  Wilms tumor 1 [WT1] · CPC title

• A61K40/46

  Viral 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

• A61K2239/50

  Colon · CPC title