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SMAD4 Loss triggers the phenotypic changes of pancreatic ductal adenocarcinoma cells

Yu-Wen Chen1, Pi-Jung Hsiao2, Ching-Chieh Weng1, Kung-Kai Kuo3, Tzu-Lei Kuo1, Deng-Chyang Wu45, Wen-Chun Hung6 and Kuang-Hung Cheng1*

Author Affiliations

1 Institute of Biomedical Sciences, National Sun Yat-Sen University, Kaohsiung 80424, Taiwan

2 Division of Endocrinology and Metabolism, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan

3 Division of Hepatobiliary Pancreatic Surgery, Department of Surgery, Kaohsiung Medical University, Kaohsiung, Taiwan

4 Division of Internal Medicine, Kaohsiung Municipal Hsiao-Kang Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan

5 Division of Gastroenterology, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan

6 National Institute of Cancer Research, National Health Research Institutes, Tainan 704, Taiwan

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BMC Cancer 2014, 14:181  doi:10.1186/1471-2407-14-181

Published: 14 March 2014



SMAD4 is a gastrointestinal malignancy-specific tumor suppressor gene found mutated in one third of colorectal cancer specimens and half of pancreatic tumors. SMAD4 inactivation by allelic deletion or intragenic mutation mainly occurs in the late stage of human pancreatic ductal adenocarcinoma (PDAC). Various studies have proposed potential SMAD4-mediated anti-tumor effects in human malignancy; however, the relevance of SMAD4 in the PDAC molecular phenotype has not yet been fully characterized.


The AsPC-1, CFPAC-1 and PANC-1 human PDAC cell lines were used. The restoration or knockdown of SMAD4 expression in PDAC cells were confirmed by western blotting, luciferase reporter and immunofluorescence assays. In vitro cell proliferation, xenograft, wound healing, quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR), Western blotting, and immunohistochemistry analysis were conducted using PDAC cells in which SMAD4 was either overexpressed or knocked down.


Here, we report that re-expression of SMAD4 in SMAD4-null PDAC cells does not affect tumor cell growth in vitro or in vivo, but significantly enhances cells migration in vitro. SMAD4 restoration transcriptionally activates the TGF-β1/Nestin pathway and induces expression of several transcriptional factors. In contrast, SMAD4 loss in PDAC leads to increased expression of E-cadherin, vascular endothelial growth factor (VEGF), epidermal growth factor receptor (EGFR) and CD133. Furthermore, SMAD4 loss causes alterations to multiple kinase pathways (particularly the phosphorylated ERK/p38/Akt pathways), and increases chemoresistance in vitro. Finally, PDAC cells with intact SMAD4 are more sensitive to TGF-β1 inhibitor treatment to reduced cell migration; PDAC cells lacking SMAD4 showed decreased cell motility in response to EGFR inhibitor treatment.


This study revealed the molecular basis for SMAD4-dependent differences in PDAC with the aim of identifying the subset of patients likely to respond to therapies targeting the TGF-β or EGFR signaling pathways and of identifying potential therapeutic interventions for PDAC patients with SMAD4 defects.

SMAD4/DPC4; TGFβ1 pathway; CD133; EGFR; Pancreatic cancer