마우스 위암 모델을 이용한 위암의 발생 및 전이에서 E-cadherin, Smad4, 그리고 p53의 역할

Abstract

Loss of E-cadherin (Cdh1), Smad4 and p53 have all been shown to play important roles in gastric cancer formation. Compound conditional knockout mice for Smad4, p53, and E-cadherin were generated to further define and compare the roles of these genes in gastric, intestinal and breast cancer development by crossing them with Pdx-1-Cre, Villin-Cre and MMTV-Cre transgenic mice. We have determined that gastric adenocarcinoma was more frequent in Pdx-1-Cre

Smad4F/F

Trp53F/F

Cdh1F/+ mice than in Pdx-1-Cre

Trp53F/F mice [P<0.001]. Pdx-1-Cre

Cdh1F/+ mice developed gastric adenocarcinomas without E-cadherin expression. However, intestinal and mammary adenocarcinomas with the same genetic background retained the E-cadherin expression and were phenotypically similar to mice with both wild-type Cdh1 alleles. Pdx-1-Cre

Cdh1F/+ mice developed gastric adenocarcinomas more rapidly than Pdx-1-Cre

F/F

Trp53 F/F

Cdh1F/F mice, demonstrating that Cdh1 heterozygosity accelerates the development and progression of gastric cancer, in combination with loss of Smad4 and p53. Lung metastases were identified in 14.2% Pdx-1-Cre

Cdh1F/+ mice, but not in the other genotypes. Nuclear β-catenin accumulation was identified at the invasive tumor front of gastric adenocarcinomas arising in Pdx-1-Cre

Cdh1F/+ mice. This phenotype was less prominent in mice with intact E-cadherin or Smad4, indicating that the inhibition of β-catenin signaling by E-cadherin or Smad4 may down-regulate signaling pathways involved in metastases in Pdx-1-Cre

Cdh1F/+ mice. Knockdown of β-catenin significantly inhibited migratory activity of Pdx-1-Cre

Cdh1F/+ cell lines. Thus, loss of E-cadherin and Smad4 cooperate with p53 loss to promote the development and metastatic progression of gastric adenocarcinomas, with similarities to human diffuse-type gastric cancer. Furthermore, a primary cell line, designated NCC-S1 was primarily cultured from a gastric cancer developed from a Villin-Cre

Cdh1F/+ mouse to establish a metastatic gastric cancer model. A cancer subpopulation of NCC-S1 cells was isolated from lung metastases that developed from heterotopic tumors transplanted into severe combined immunodeficiency mice. The metastasis derived cells, designated NCC-S1M, formed orthotopic tumors and exhibited a significant improvement in in vivo metastatic potential compared with the parental cell line, showing epithelial to mesenchymal transition (EMT) features. NCC-S1M cells demonstrated many features of cancer stem cells, including the ability to initiate tumor formation and chemoresistance. Importantly, Sca-1 was found to be over-expressed in NCC-S1M cells than NCC-S1 cells. Sca-1 positive cells demonstrated increased ability to initiate tumor formation and in vivo chemoresistance than Sca-1 negative cells, showing up-regulation of interleukin 1 signaling pathway. Over-expressed genes in Sca-1 positive NCC-S1 cells clustered in 123 metastatic gastric cancer patients according to overall survival following cisplatin/fluorouracil chemotherapy. Taken together, we have identified Sca-1 as a murine gastric cancer stem cell marker using this unique metastatic cell line models.