title: Anterior gradient 2 downregulation in a subset of pancreatic ductal adenocarcinoma is a prognostic factor indicative of epithelial-mesenchymal transition.
abstract: Anterior gradient 2 (AGR2), a member of the protein disulfide isomerase family, has been implicated in various cancers including pancreatic ductal adenocarcinoma (PDAC) and is known to promote cancer progression. However, the prognostic value of AGR2 expression and the interaction with epithelial-mesenchymal transition (EMT) remain unclear. We investigated the clinical significance of AGR2 and EMT markers in PDAC patients by immunohistochemical analyses. Although AGR2 expression was not observed in normal pancreas, all pancreatic precursor neoplastic lesions were positive for AGR2, even at the earliest stages, including pancreatic intraepithelial neoplasia-1A, AGR2 expression was reduced in 27.7% (54/195 cases) of PDAC patients. AGR2 downregulation correlated with EMT markers (vimentin overexpression and reduced membranous E-cadherin expression), high Union for International Cancer Control stage (P<0.0001), high histological cellular grade (P<0.0001), and adverse outcome (P<0.0001). In vitro, targeted silencing of AGR2 in cancer cells using siRNA reduced cell proliferation, colony formation, cell invasiveness, and migration, but did not alter EMT markers. To confer a more aggressive phenotype and induce EMT in PDAC cells, we co-cultured PDAC cell lines with primary-cultured pancreatic stellate cells (PSCs) and found that AGR2 was downregulated in co-cultured PDAC cells compared with PDAC monocultures. Treatment with transforming growth factor beta-1 (TGF-β), secreted from PSCs, decreased AGR2 expression, whereas inhibition of TGF-β signaling using recombinant soluble human TGF-β receptor type II and TGF-β-neutralizing antibodies restored AGR2 expression. We conclude that AGR2 downregulation is a useful prognostic marker, induced by EMT, and that secreted TGF-β from PSCs may partially contribute to AGR2 downregulation in PDAC patients. AGR2 downregulation does not induce EMT or a more aggressive phenotype, but is a secondary effect of these processes in advanced PDAC.
authors.affiliation: 1] Department of Anatomic Pathology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan [2] Reserch Fellow of Japan Society for the Promotion of Science, Tokyo, Japan.
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Mizuuchi
authors.affiliation: Department of Anatomic Pathology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Shinichi
authors.initials: S
authors.lastname: Aishima
authors.affiliation: Department of Surgery and Oncology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Kenoki
authors.initials: K
authors.lastname: Ohuchida
authors.affiliation: 1] Department of Anatomic Pathology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan [2] Reserch Fellow of Japan Society for the Promotion of Science, Tokyo, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Shindo
authors.affiliation: Department of Anatomic Pathology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Minoru
authors.initials: M
authors.lastname: Fujino
authors.affiliation: Department of Anatomic Pathology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Masami
authors.initials: M
authors.lastname: Hattori
authors.affiliation: Department of Anatomic Pathology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Tetsuyuki
authors.initials: T
authors.lastname: Miyazaki
authors.affiliation: Kyushu University Hospital Cancer Center, Fukuoka, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Mizumoto
authors.affiliation: Department of Surgery and Oncology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Masao
authors.initials: M
authors.lastname: Tanaka
authors.affiliation: Department of Anatomic Pathology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Yoshinao
authors.initials: Y
authors.lastname: Oda
mesh_terms: D014408:Biomarkers, Tumor / Q000378:metabolism*; D015153:Blotting, Western; D015820:Cadherins / Q000378:metabolism; D021441:Carcinoma, Pancreatic Ductal / Q000378:metabolism*; D002465:Cell Movement / Q000502:physiology; D049109:Cell Proliferation / Q000502:physiology; D003114:Colony-Forming Units Assay; D017931:DNA Primers / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D058750:Epithelial-Mesenchymal Transition / Q000235:genetics / Q000502:physiology*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D007150:Immunohistochemistry; D018061:Infectious pancreatic necrosis virus / Q000378:metabolism*; D053208:Kaplan-Meier Estimate; D009088:Mucoproteins; D009361:Neoplasm Invasiveness / Q000503:physiopathology; D015513:Oncogene Proteins; D011379:Prognosis; D011506:Proteins / Q000378:metabolism*; D034622:RNA Interference; D060888:Real-Time Polymerase Chain Reaction; D016212:Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000494:pharmacology; D014746:Vimentin / Q000378:metabolism
chemical_list: C502296:AGR2 protein, human; D014408:Biomarkers, Tumor; D015820:Cadherins; D017931:DNA Primers; D009088:Mucoproteins; D015513:Oncogene Proteins; D011506:Proteins; D016212:Transforming Growth Factor beta; D014746:Vimentin