In:
Proceedings of the National Academy of Sciences, Proceedings of the National Academy of Sciences, Vol. 109, No. 17 ( 2012-04-24)
Abstract:
Our results have broad clinical implications. Small-molecule inhibitors targeting the Smo signaling molecule are being evaluated currently for the treatment of pancreatic cancer ( 5 ). However, the activity of these Smo inhibitors likely is limited to the stromal compartment, having no effect on epithelial cells. Our genetic data identifying an alternative genetic pathway involving Gli argue strongly for the clinical development of Gli inhibitors, because these molecules contribute critical function in the tumor epithelium as well as the tumor stroma. We next investigated which downstream genes mediated Gli-driven phenotypes in pancreatic cancer cells. We performed gene-expression profiling analysis in human pancreatic ductal adenocarcinoma cells in which Gli-mediated transcription had been inhibited by Gli3T. Interestingly, we found that Gli activation in pancreatic cancer cells imposed a unique transcriptional program. Specifically, we found that the expression of the IκB kinase IKBKE (IKKε) was regulated by Gli proteins. These results suggest that IKBKE may play a critical role in pancreatic tumorigenesis. In general, kinases are enzymes that affect other proteins by adding phosphate molecules to these proteins. IKBKE is a noncanonical IkB kinase involved in stimulating the NF-κB signaling pathway. We showed that IKBKE is highly expressed in pancreatic tumors that are characterized by both Kras and Gli activation. We also found that Gli regulates NF-κB activity in vitro and in vivo. Furthermore, we demonstrated that Gli-dependent IKBKE expression in pancreatic cancer cells is essential for maintaining their cell survival and transformation characteristics, which are fundamental properties of cancerous cells. These data establish a connection between Gli transcription factors and the IKBKE and NF-κB activity in tumorigenesis, providing a molecular basis for the observed requirement of Gli transcription in the development of pancreatic ductal adenocarcinoma in vivo. In addition, we evaluated the tumorigenic potential of Gli1 in pancreatic cancer by ectopically expressing Gli1 in the mouse pancreas. We found that Gli1 activation significantly enhanced the ability of Kras to promote tumor development. Thus, both our loss-of-function and gain-of-function analyses highlight the functional importance of epithelial Gli activity in Kras-induced pancreatic tumorigenesis in vivo. However, using a primary pancreatic ductal epithelial cell (PDEC) culture, we showed that Gli-mediated transcription activation is required for Kras-induced PDEC proliferation and survival. Additionally, we combined this model with another in which oncogenic Kras activation in the mouse pancreatic epithelium drove tumor initiation and progression, mimicking the pathogenesis of human pancreatic cancer. Significantly, when these mice were genetically modified to exhibit both activation of Kras and inhibition of Gli transcription in the pancreatic epithelium, we found that the development of pancreatic precursor lesions and pancreatic ductal adenocarcinoma initiated by Kras activation was blocked. This inhibition happened even when the p53 tumor suppressor, which commonly is inactivated in this disease, was deleted. These results provide genetic evidence that Gli activity is required for Kras-driven transformation of the pancreatic epithelium. To investigate the role of Gli in pancreatic tumorigenesis, we generated a mouse model of pancreatic epithelium-specific inhibition of Gli-mediated transcription. This model was achieved by the ectopic expression of a dominant Gli3 repressor allele, Gli3T, which is capable of inhibiting Gli1- and Gli2-mediated activation of transcription. We found that inhibition of Gli transcriptional activity did not affect differentiation of the pancreatic epithelial cell lineages. This lack of effect suggests that epithelial Gli activity is largely dispensable for the proper development of the mouse pancreas. The Gli proteins Gli1, Gli2, and Gli3 are transcription factors. They operate in a cellular signaling pathway called the “Hedgehog” (Hh) pathway that often is activated in human tumors. The Hh pathway is mediated by activation of a cell-surface molecule, Smoothened (Smo) ( 3 ). Stimulation of the Hh pathway results in the activation of Gli proteins. However, Gli proteins may be regulated by other oncogenic pathways. For example, Gli1 gene expression is maintained in pancreatic cancer cells despite deletion of the Smo gene and is regulated in part by Kras independently of Hh ligand input ( 4 ). However, Gli's role in pancreatic tumorigenesis in vivo remains unexplored. Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer-related mortality in the United States ( 1 ). This disease arises from precursor lesions in pancreatic epithelium, characterized by mutations in a potential cancer-causing gene called the “ KRAS oncogene” ( 1 , 2 ). However, limited understanding of the molecular and genetic mechanisms underlying Kras-dependent pancreatic tumorigenesis has impeded the development of early diagnostic and treatment tools. Here, using mouse genetics and cellular biology, we demonstrated that the ability of Gli proteins to activate gene transcription is critical for the development of Kras-driven pancreatic cancer ( Fig. P1 ). Significantly, we identified a key mechanism that operates after Gli transcription activity during pancreatic tumorigenesis: the induction of the expression of the atypical IkB kinase IKBKE and the activity of the NF-κB pathway in the tumor epithelium ( Fig. P1 ).
Type of Medium:
Online Resource
ISSN:
0027-8424
,
1091-6490
DOI:
10.1073/pnas.1114168109
Language:
English
Publisher:
Proceedings of the National Academy of Sciences
Publication Date:
2012
detail.hit.zdb_id:
209104-5
detail.hit.zdb_id:
1461794-8
SSG:
11
SSG:
12
Permalink
