Abstract 3355: Signaling networks in pancreatic cancer cells with stem cell characteristics
Authors: Matthias Ilmer, Jody Vykoukal, Eckhard Alt
Journal: Cancer Research (2012)

Abstract Despite considerable efforts, the 5-year survival rate for pancreatic ductal adenocarcinoma (PDAC) has not changed in the last several decades. This poor prognosis is a function of the propensity of PDAC towards early metastatic spread and drug resistance. An ever-growing body of evidence attributes these characteristics to the presence of cancer stem cells (CSCs) within the tumor, the specific origin and nature of which is yet to be fully understood. Conserved pathways, e.g. TGF-β and Wnt, have been shown in general to regulate CSC hallmarks of self-renewal and differentiation, but these effects seem restricted to specialized cancer cells within the tumor mass. Notwithstanding, the function of Wnt signaling alone or in combination with other pathways in pancreatic CSCs remains elusive. We are elucidating whether microenvironmental cues are able to induce stemness in so-called facultative stem cells associated with the tumor mass. In light of mounting evidence, we are investigating canonical Wnt signaling and its interplay with other pathways as well as its function in regulating stemness-associated characteristics _ anoikis, epithelial-mesenchymal plasticity, drug-resistance, and tumorigenesis. To enrich for CSCs, we established sphere culture in serum-depleted media that yields cells exhibiting high levels of embryonic stem cell markers (Bmi1, Nanog, Oct3/4, Sox2), and increased colony-formation and drug-resistance relative to adherent cultures of the same line. Early immunofluorescence staining data reveal heterogeneous cellular β-Catenin distribution in the parental population of an established pancreatic cancer line (Panc1), increased nuclear translocation after Wnt3a stimulation, and higher nuclear β-Catenin staining in sphere cells. Known Wnt target genes (CyclinD1, Axin2) detected by qRT-PCR are significantly upregulated in spheroid cultures, further indicating that canonical Wnt-signaling could be intrinsically up regulated. In a sphere-forming, surrogate in vitro, assay for evaluation of stemness, constant stimulation with rhWnt3a increased the self-renewal capacity of floating pancreatic cancer cells. Together, these data support the hypothesis that activation of Wnt-signaling in an autocrine/paracrine manner might lead to stemness in susceptible CSC. We are deciphering patterns of multiple heterogeneously activated pathways, specifically the signaling sequence and chronology, that drives sphere-formation. Western Blot analysis shows enhanced activating phosphorylation of β-Catenin in sphere cells whereas SMAD3, a critical component of TGF-β-signaling, was activated in more differentiated pancreatic cancer cells. In conclusion, we show evidence for differential regulation of Wnt/TGF-β-signaling in pancreatic cancer cells, suggesting a reconsideration of current clinical strategies to include combining standard therapy with inhibitors of these signaling pathways. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3355. doi:1538-7445.AM2012-3355