Abstract 3325: Cancer stem cell like properties of 5-fluorouracil resistant Panc-1 cell

Abstract

Abstract Cancer stem cells (CSCs), a small fraction of tumor-initiating cells, are known to be resistant to chemotherapy-induced apoptosis. CSCs are considered to be responsible for self-renewal/differentiation, invasion, and metastasis of malignant tumor. We have previously reported that pancreatic cancer derived side population (SP) cells, a cancer stem cell enriched fraction, were more resistant to chemotherapeutic agents and more likely to cause epithelial-to-mesenchymal transition (EMT) compare to major population (MP) cells. Therefore, we formulated the HYPOTHESIS that 5-fuluorouracil (5-FU) resistant population cells are enriched with CSCs. The AIM of this study is to evaluate whether apoptosis-resistant cancer cells have CSC-like properties, i.e., self-renewal, local invasion and EMT. METHODS: Seventy to eighty percent confluent pancreatic cancer cells, PANC-1, were incubated in the presence of 5-FU (30mg/ml) for 24 hours, and further incubated for one to five weeks without 5-FU. To assess the capacity of self-renewing, cells were planted using a method for anchorage-independent culturing to form spheroids. EMT was induced with TGF-beta (7.5 ng/ml, for 3 days), then evaluated mRNA expression by real time PCR for E-cadherin, Snail, and vimentin. E-cadherin protein level was also examined for immunoblot analysis. Activity of local invasion was analyzed by Matrigel invasion assay. To evaluate tumorigenicity, 5-FU resistant cells were injected into NOD/SCID mice. RESULTS: 5-FU induced significant apoptosis in PANC-1 cells, reducing the number of surviving cells to less than one percent. The survived cells showed a colony-forming activity as they started to re-proliferate at four weeks after 5-FU removal. There was a group of cells among the 5-FU resistant surviving cells, that gradually regains sensitivity to 5-FU by re-plating. The frequency of cells that were capable of initiating spheres was higher in 5-FU resistant cells and they over expressed stem cell marker genes, Oct 4 and Nanog. Incubation with TGF-beta induced EMT-associated gene alterations. These alterations were greater in 5-FU resistant surviving cells as compared to untreated cells. Matrigel invasion activity of apoptosis-resistant PANC-1 cells was greater by seven fold than controlled cells. In addition, in vivo tumor experiments showed 5-FU resistant cells to be more tumorigenic than conventional Panc-1 cells. In CONCLUSION, apoptosis-resistant cancer cells have CSC-like properties, i.e., initiating sphere formation, expressing stem cell genes, and respond to EMT stimulation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3325.