Ligand-dependent Notch Signaling Is Involved in Tumor Initiation and Tumor Maintenance in Pancreatic Cancer

Abstract

Purpose: Aberrant activation of the Notch signaling pathway is commonly observed in human pancreatic cancer, although the mechanism(s) for this activation has not been elucidated. Experimental Design: A panel of 20 human pancreatic cancer cell lines was profiled for the expression of Notch pathway-related ligands, receptors, and target genes. Disruption of intracellular Notch signaling, either genetically by RNA interference targeting NOTCH1 or pharmacologically by means of the g-secretase inhibitor GSI-18, was used for assessing requirement of Notch signaling in pancreatic cancer initiation and maintenance. Results: Striking overexpression of Notch ligand transcripts was detectable in the vast majority of pancreatic cancer cell lines, most prominently JAGGED2 (18 of 20 cases, 90%) and DLL4 (10 of 20 cases, 50%). In two cell lines, genomic amplification of the DLL3 locus was observed, mirrored by overexpression of DLL3 transcripts. In contrast, coding region mutations of NOTCH1 or NOTCH2 were not observed. Genetic and pharmacologic inhibition of Notch signaling mitigated anchorage-independent growth in pancreatic cancer cells, confirming that sustained Notch activation is a requirement for pancreatic cancer maintenance. Further, transient pretreatment of pancreatic cancer cells with GSI-18 resulted in depletion in the proportion of tumor-initiating aldehyde dehydrogenase^expressing subpopulation and was associated with inhibition of colony formation in vitro and xenograft engraftment in vivo, underscoring a requirement for the Notch-dependent aldehyde dehydrogenase^expressing cells in pancreatic cancer initiation. Conclusions: Our studies confirm that Notch activation is almost always ligand dependent in pancreatic cancer, and inhibition of Notch signaling is a promising therapeutic strategy in this malignancy.Pancreatic cancer is an almost uniformly lethal disease with an overall 5-year survival of f5%, and this dire prognosis has not markedly improved over the last few decades (1). In the United States, f34,000 individuals succumb to this malignancy each year. To date, the only potentially curative therapeutic option is complete surgical resection, but unfortunately, the majority of patients are diagnosed at a locally advanced or distant metastatic stage, thus precluding surgical cure (2). Currently available treatment options for advanced pancreatic cancer, such as gemcitabine, have had minimal effect in ameliorating survival. Identification of aberrant signaling pathways that can also form the substrate for targeted therapies has thus become an area of foremost priority.The reactivation of embryonic signal transduction pathways, such as Notch and Hedgehog, have been reported in a variety of human cancers (3, 4); further, the availability of potent smallmolecule inhibitors has meant that these pathways can be targeted in these cancers, as we and others have recently shown

Notch1‐Snail1‐E‐cadherin pathway in metastatic hepatocellular carcinoma

“…Using over-expression and knock-down of Notch signaling by siRNA or inhibitors, activated Notch1 signaling has been reported to be involved in tumor development, 32 tumor growth, 33 tumor angiogenesis, 34 chemotherapy sensitivity 35 and invasion of tumor cells by limiting the migration of tumor cells. 17,19,20,36 For the first time, our study demonstrated that knocking-down Notch1 was able to inhibit liver tumor metastasis in vivo in a mouse model. Therefore, targeting the Notch pathway could be a potential treatment for tumor metastasis.…”

“…High levels of Notch1 expression were significantly associated with advanced TNM staging and blood vessel infiltration in HCC (Table 1); Jagged1 expression significantly correlated with the presence of tumor nodules (data not shown). The roles of aberrant Notch signaling in tumors have primarily been linked to tumor initiation and maintenance, 20 the promotion of tumor growth and progression 17 and tumor angiogenesis. 34 Previous observations of Notch signaling mediating tumor invasion and metastasis have been limited to only cancer cell lines.…”

“…17 In addition, Notch signaling also regulates NFjB-DNA binding, 18 which affects genes that are critical for metastasis. Some recent studies have demonstrated the transcriptional activation of MMP9 and uPA by Notch1 in human prostate cancer 19 and the participation of Notch1 signaling in pancreatic cancer 20 and osteosarcoma. 21 In breast cancer, the suppression of E-cadherin expression occurs through the Jagged1-Notch1-Slug EMT pathway.…”

Chemoresistance in Cancer Stem Cells and Strategies to Overcome Resistance

“…Additional toxicity may be observed due to goblet cell metaplasia [114,115]. A third GSI, GSI-18 been shown to deplete the Aldefluor+ CSCs and decrease colony formation and xenograft engraftment in pancreatic cancer models [116]. Similar results were seen in DAOY medulloblastoma cells [117].…”

Hey Factors at the Crossroad of Tumorigenesis and Clinical Therapeutic Modulation of Hey for Anticancer Treatment

“…The levels of Hey factors are strikingly elevated in high-grade glioma, malignant osteosarcoma, high-grade esophageal squamous cell carcinoma, aggressive pancreatic adenocarcinoma rhabdomyosarcoma, as well as colorectal carcinoma (17)(18)(19)(20)(21)(22)(23). In these malignant carcinomas, aberrant Hey expression has been associated with poor prognosis, overall survival (OS), tumor grade, chemotherapy resistance, lymphatic metastasis, and vascular proliferative properties (17,(23)(24)(25).…”