Abstract 3750: Inhibition of phosphorylation of nestin decrease pancreatic cancer cell growth.
Authors: Yoko Matsuda, Hisashi Yoshimura, Taeko Suzuki, Zenya Naito, Kiyoko Kawahara, Yuji Yanagisawa, Yoko Kawamoto, Toshiyuki Ishiwata
Journal: Cancer Research (2013)

Abstract Background: Nestin is a class VI intermediate filament, first described as a central nervous system (CNS) progenitor/stem cell marker and recently reported as a tumor stem cell marker in several malignant tumors, including glioblastoma and melanoma. In the normal pancreas, nestin is expressed in exocrine progenitor cells. Previously, we have reported that 30% of pancreatic cancer cases expressed nestin in cancer cells, and its expression correlated with nerve invasion and the presence of cancer cells in the tumor resection margins (Kawamoto, et al. Hum Pathol 2009). Knockdown of nestin exhibited a sheet-like appearance with tight cell-cell adhesion, increased expression of filamentous-actin and E-cadherin, and attenuated migration and invasion, both of which recovered following nestin re-expression (Matsuda, et al. Cancer Biol Ther 2011). Furthermore, knockdown of nestin suppressed sphere-forming ability, and altered the expression levels of stemness-related genes such as NANOG and transcription factor 4 (Matsuda, et al. AACR 2012). Therefore, nestin is considered to be a novel therapeutic target for pancreatic cancer via suppression of cell migration, invasion, metastasis and stemness. To develop nestin-targeting therapy, we focused on the phosphorylation of nestin protein. Phosphorylation of nestin at two different sites of progenitor cells of the CNS and myocytes has been reported to regulate the cell cycle via activation of cyclin-dependent kinase 5 and cdc2 kinase (Sahlgren, et al. Mol Cell Biol 2003). In this study, we analyzed the expression and roles of nestin phosphorylation in pancreatic cancer cells. Methods: Expression levels of phosphorylated nestin in pancreatic cancer cells were determined by flow cytometer, fluorescent staining and Western blot analyses. To assess the functions of phosphorylated nestin, we prepared three mutated nestin expression vectors which possess mutations at two different phosphorylated sites, both in nestin. A human pancreatic cancer cell line, MIA PaCa-2, which express very low levels of nestin was transfected with the mutated or wild-type nestin-expression vector. Results: Expression level of phosphorylated nestin was higher in M-phase cells than S- or G0/G1-phase cells. Phosphorylated nestin was strongly and diffusely localized in M-phase cells, while it was weakly detected in the nuclei of cells in other phases. Mutated nestin in phosphorylated site-transfected cells showed lower levels of cell growth, migration and invasion than wild-type nestin-transfected cells. Furthermore, mutated nestin-transfected cells had a tendency towards increasesed multinuclear cells. Conclusion: Phosphorylation of nestin affects the cell growth, migration and invasion of pancreatic cancer cells, and phosphorylated nestin may play important roles in the regulation of the cell cycle. Inhibition of phosphorylated nestin may be a novel target for pancreatic cancer treatment. Citation Format: Yoko Matsuda, Hisashi Yoshimura, Taeko Suzuki, Zenya Naito, Kiyoko Kawahara, Yuji Yanagisawa, Yoko Kawamoto, Toshiyuki Ishiwata. Inhibition of phosphorylation of nestin decrease pancreatic cancer cell growth. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3750. doi:10.1158/1538-7445.AM2013-3750