Cancer stem cell population and method for production thereof

Patent metadata

Field	Value
Publication number	US-11965180-B2
Application number	US-202016994388-A
Country	US
Kind code	B2
Filing date	Aug 14, 2020
Priority date	Oct 6, 2010
Publication date	Apr 23, 2024
Grant date	Apr 23, 2024

Abstract

Provided is a cancer stem cell mass from which cells incapable of forming cancer are substantially removed and which has a characteristic property of reproducing a layered structure of a cancer tissue; a process for producing the cancer stem cell mass; and use of the cancer stem cell mass. A human cancer tissue was repeatedly grown in a NOG mouse, separated cancer cells from the grown cancer tissue, and tested and compared various cancer cell culture processes. As a result, a cancer stem cell composition which is homogeneous and is substantially free of the coexistence of cells capable of forming cancer and cells incapable of forming cancer in a mixed state can be produced successively by employing an attached culture process using a serum-free stem cell culture medium rather than a generally employed floating culture process.

First claim

The invention claimed is:

1. A method of identifying a drug candidate, comprising:
(i) transplanting a sample comprising human colon cancer tissue into NOD/SCID/gamma c null or NOD-scid/IL-2Rg null (NOG) mice to produce a human colon cancer cell mass;
(ii) isolating the human colon cancer cell mass from the NOD/SCID/gamma c null or NOD-scid/IL-2Rg null (NOG) mice;
(iii) preparing cells from the human colon cancer cell mass;
(iv) culturing the cells prepared from the human colon cancer cell mass in an in vitro adherent culture system using a serum-free stem cell medium, wherein culturing the cells comprises removing floating cells, and wherein adherent cells from the human colon cancer cell mass produce a population of human colon cancer stem cells from which cells with no cancer-forming ability have been substantially removed,
wherein the population of human colon cancer stem cells are positive for LGR5;
(v) culturing the population of human colon cancer stem cells from which cells with no cancer-forming ability have been substantially removed, wherein the population is characterized by reproducing a hierarchical structure of a cancer tissue, and the population is cultured as a spheroid culture under in vitro conditions to reproduce a characteristic structure of a cancer development process originating from human colon cancer stem cells or a biological property of human colon cancer stem cells;
(vi) treating the cultured human colon cancer stem cells of (v) with a test substance;
(vii) observing a change in the hierarchical structure formed from the human colon cancer stem cells, a cancer development process originating from the human colon cancer stem cells, or a biological property of the human colon cancer stem cells; and
(viii) identifying a test substance that inhibits formation of the hierarchical structure formed from the human colon cancer stem cells, a cancer development process originating from the human colon cancer stem cells, or a biological property of the human colon cancer stem cells, as the drug candidate.

2. The method of claim 1 , wherein the population of human colon cancer stem cells is substantially homogeneous.

3. The method of claim 1 , wherein the frequency of human colon cancer stem cells in the population of human colon cancer stem cells in extreme limiting dilution analysis is 1/20 or higher.

4. The method of claim 1 , wherein the population of human colon cancer stem cells comprises 1×10 4 or more human colon cancer stem cells.

Assignees

• Chugai Pharmaceutical Co Ltd

Inventors

• Yamazaki Tatsumi
• Okabe Hisafumi
• Kobayashi Shinta
• Chen Yu Jau
• Kato Atsuhiko
• Suzuki Masami
• Matsubara Koichi

Classifications

• C12N5/0695 Primary

  Stem cells; Progenitor cells; Precursor cells · CPC title

• G01N33/5011

  for testing antineoplastic activity · CPC title

• G01N33/5073

  Stem cells · CPC title

• A61P35/00

  Antineoplastic agents · CPC title