Abstract
Pancreatic ductal adenocarcinoma (PDAC) is the fourth most common cause of cancer death in North America. Activating KRAS mutations and Smad4 loss occur in approximately 90% and 55% of PDAC, respectively. While their roles in the early stages of PDAC development have been confirmed in genetically modified mouse models, their roles in the multistep malignant transformation of human pancreatic duct cells have not been directly demonstrated. Here, we report that Smad4 represents a barrier in KRAS-mediated malignant transformation of the near normal immortalized human pancreatic duct epithelial (HPDE) cell line model. Marked Smad4 downregulation by shRNA in KRAS G12V expressing HPDE cells failed to cause tumorigenic transformation. However, KRAS-mediated malignant transformation occurred in a new HPDE-TGF-β resistant (TβR) cell line that completely lacks Smad4 protein expression and is resistant to the mito-inhibitory activity of TGF-β. This transformation resulted in tumor formation and development of metastatic phenotype when the cells were implanted orthotopically into the mouse pancreas. Smad4 restoration re-established TGF-β sensitivity, markedly increased tumor latency by promoting apoptosis, and decreased metastatic potential. These results directly establish the critical combination of the KRAS oncogene and complete Smad4 inactivation in the multi-stage malignant transformation and metastatic progression of normal human HPDE cells.
Highlights
Pancreatic cancer is the fourth leading cause of cancer death in North America with an overall five year survival rate of
We have demonstrated that KRASG12V expression is insufficient for the full malignant transformation of H6c7 cells when residual Smad4 signaling remains
As H6c7 cells have already manifested deregulated Rb and p53 pathways, we conclude that Smad4 absence is obligatory and serves as a restriction point for KRAS-mediated malignant transformation of the human pancreatic duct epithelial (HPDE) cell line
Summary
Pancreatic cancer is the fourth leading cause of cancer death in North America with an overall five year survival rate of 90% of invasive PDAC and during the multi-stage PDAC carcinogenesis, its occurrence has been shown to precede the inactivation of tumor suppressors p16 (95%), p53 (75%), and Smad (55%) [3]. Modified mouse models (GEMMs) engineered to express the KRASG12D oncogene in the developing pancreas can recapitulate the advancement of PanIN lesions seen in patients, only a subset of mice develop invasive and metastatic PDAC [5,6]. The incomplete progression to invasive adenocarcinoma indicates that KRAS alone is insufficient for malignant transformation of the pancreatic duct epithelium
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