Method for searching and screening for target of anti-cancer agent using non-human animal model having NOG established cancer cell line transplanted therein

Patent metadata

Field	Value
Publication number	US-11536713-B2
Application number	US-201013519059-A
Country	US
Kind code	B2
Filing date	Dec 24, 2010
Priority date	Dec 25, 2009
Publication date	Dec 27, 2022
Grant date	Dec 27, 2022

Abstract

An objective of the present invention is to provide non-human animal models of cancer pathology, which mimic the hierarchical organization, cancer progression process, or biological property of human cancer tissues, and uses thereof. To achieve the objective described above, first, the present inventors transplanted cells of NOG-established cancer lines into NOG mice and morphologically observed the resulting tissue organization. As a result, the non-human animal models were demonstrated to exhibit pathologies (the hierarchical organization, cancer progression process, or biological properties of the cancer cells) similar to that of human cancer. Specifically, the present inventors succeeded in preparing non-human animal models exhibiting pathologies more similar to a human cancer, and cell culture systems using NOG-established cancer cell lines where the in vitro cell morphology is more similar to that of human cancer.

First claim

What is claimed is:

1. A method of searching for an anti-cancer agent target, which comprises the steps of:
(1) preparing a mouse model of human colon cancer by transplanting an NOD/SCID/gamma(c)(null) NOG-established colon cancer cell line into a mouse,
wherein the NOD/SCID/gamma(c)(null) (NOG)-established cancer cell line was established by transplanting a human colon cancer tissue into a severely immunodeficient NOG mouse, and
wherein the mouse model of human colon cancer comprises:
(a) cancer stem cells in an epithelium-like glandular area, and in an invasive area, detected using continuous BrdU labelling, Ki67 staining, or Lgr5 staining, and
(b) partial epithelial-mesenchymal transition (EMT) cells derived from cancer stem cells in the invasive area, detected by a lack of E-cadherin expression at a contact surface of a tumor cell with stroma cells or presence of fibronectin expression,

(2) identifying a tissue piece from the mouse model of human colon cancer comprising fibroblasts derived from human colon cancer cells through complete epithelial-mesenchymal transition (complete-EMT) in a tumor stroma using IHC staining,
(3) collecting the tissue piece identified in step (2);
(4) assessing the expression of DNA, RNA, or protein in the collected tissue piece of (3); and
(5) identifying DNA, RNA, or protein that changes in a manner dependent of hierarchical organization and cancer progression process of a cancer cell in the tissue piece, thereby identifying the anti-cancer agent target,
wherein the hierarchical organization comprises a two-layer structure comprising an epithelium-like glandular structure and an invasive area, and
wherein the cancer progression process comprises:
(a) an epithelial duct-forming pattern or
(b) an invasive EMT pattern, while including an epithelial duct-forming pattern.

2. The method of claim 1 , further comprising in step (2) detecting morphologies comprising budding, clustering tumor cells, dissociated single tumor cells, or reconstruction of tubules in the invasive area.

3. A method of searching for an anti-cancer agent target, which comprises the steps of:
(1) preparing a mouse model of human colon cancer by subcutaneously transplanting an NOD/SCID/gamma(c)(null) (NOG)-established colon cancer cell line into a mouse,
wherein the NOD/SCID/gamma(c)(null) (NOG)-established cancer cell line was established by transplanting a human colon cancer tissue into a severely immunodeficient NOG mouse, and
wherein the mouse model of human colon cancer comprises:
(a) cancer stem cells in an epithelium-like glandular area, and in an invasive area, detected using continuous BrdU labelling, Ki67 staining, or Lgr5 staining, and
(b) partial epithelial-mesenchymal transition (EMT) cells derived from cancer stem cells in the invasive area, detected by a lack of E-cadherin expression at a contact surface of a tumor cell with stroma cells or presence of fibronectin expression,

( 2 ) identifying a tissue piece from the mouse model of human colon cancer comprising fibroblasts derived from human colon cancer cells through complete epithelial-mesenchymal transition (complete-EMT) in a tumor stroma using IHC staining
(3) collecting the tissue piece identified in step (2);
(4) assessing the expression of DNA, RNA, or protein in the collected tissue piece of (3); and
(5) identifying DNA, RNA, or protein that changes in a manner dependent of hierarchical organization and cancer progression process of a cancer cell in the tissue piece, thereby identifying the anti-cancer agent target,
wherein the hierarchical organization comprises a two-layer structure comprising an epithelium-like glandular structure and an invasive area, and
wherein the cancer progression process comprises:
(a) an epithelial duct-forming pattern or
(b) an invasive EMT pattern, while including an epithelial duct-forming pattern.

4. The method of claim 3 , further comprising in step (2) detecting morphologies comprising budding, clustering tumor cells, dissociated single tumor cells, or reconstruction of tubules in the invasive area.

5. The method of claim 1 , further comprising in step (2) detecting cancer stem cells with continuous BrdU labelling, Ki67 staining, and Lgr5 staining.

6. The method of claim 3 , further comprising in step (2) detecting cancer stem cells with continuous BrdU labelling, Ki67 staining, and Lgr5 staining.

Assignees

• Suzuki Masami
• Matsubara Koichi
• Kato Atsuhiko
• Kato Chie
• Kobayashi Shinta
• Chen Yu Jau
• Yamazaki Masaki
• Chugai Pharmaceutical Co Ltd

Inventors

• Suzuki Masami
• Matsubara Koichi
• Kato Atsuhiko
• Kato Chie
• Kobayashi Shinta
• Chen Yu Jau
• Yamazaki Masaki

Classifications

• A01K2227/105

  Murine · CPC title

• A01K67/0271

  Chimeric vertebrates, e.g. comprising exogenous cells · CPC title

• A01K2207/12

  Animals modified by administration of exogenous cells · CPC title

• A01K2267/0331

  Animal model for proliferative diseases · CPC title

• G01N33/5011 Primary

  for testing antineoplastic activity · CPC title