title: Pancreatic cancer cells surviving gemcitabine treatment express markers             of stem cell differentiation and epithelial-mesenchymal transition.
abstract: Objective response rates to standard chemotherapeutic regimens remain low in pancreatic cancer. Subpopulations of cells have been identified in various solid tumors which express stem cell-associated markers and are associated with increased resistance against radiochemotherapy. We investigated the expression of stem cell genes and markers of epithelial-mesenchymal transition in pancreatic cancer cells that survived high concentrations of gemcitabine treatment. Capan-1 and Panc-1 cells were continuously incubated with 1 and 10 µM gemcitabine. Surviving cells were collected after 1, 3 and 6 days. Expression of PDX-1, SHH, CD24, CD44, CD133, EpCAM, CBX7, OCT4, SNAIL, SLUG, TWIST, Ki-67, E-cadherin, β-catenin and vimentin were quantified by qPCR or immunocytochemistry. Migration was assessed by wound‑healing assay. SHH was knocked down using RNA interference. Five primary pancreatic cancer cell lines were used to validate the qPCR results. All investigated genes were upregulated after 6 days of gemcitabine incubation. Highest relative expression levels were observed for OCT4 (13.4-fold), CD24 (47.3-fold) and EpCAM (15.9-fold) in Capan-1 and PDX-1 (13.3‑fold), SHH (24.1-fold), CD44 (17.4-fold), CD133 (20.2-fold) and SLUG (15.2-fold) in Panc-1 cells. Distinct upregulation patterns were observed in the primary cells. Migration was increased in Panc-1 cells and changes in the expression of E-cadherin and β-catenin were typical of epithelial-mesenchymal transition in both cell lines. SHH knockdown reduced IC(50) from 30.1 to 27.6 nM in Capan-1 while it strongly inhibited proli-feration in Panc-1 cells. Cells surviving high-dose gemcitabine treatment express increased levels of stem cell genes, show characteristics associated with epithelial-mesenchymal transition and retain their proliferative capacity.
authors.affiliation: Institute for Surgical Research, Philipps University of Marburg, D-35043 Marburg, Germany. karl.quint@med.uni-marburg.de
authors.forename: Karl
authors.initials: K
authors.lastname: Quint
authors.forename: Manuel
authors.initials: M
authors.lastname: Tonigold
authors.forename: Pietro
authors.initials: P
authors.lastname: Di Fazio
authors.forename: Roberta
authors.initials: R
authors.lastname: Montalbano
authors.forename: Susanne
authors.initials: S
authors.lastname: Lingelbach
authors.forename: Felix
authors.initials: F
authors.lastname: Rückert
authors.forename: Beate
authors.initials: B
authors.lastname: Alinger
authors.forename: Matthias
authors.initials: M
authors.lastname: Ocker
authors.forename: Daniel
authors.initials: D
authors.lastname: Neureiter
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D015415:Biomarkers / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects*; D045744:Cell Line, Tumor; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D058750:Epithelial-Mesenchymal Transition / Q000187:drug effects* / Q000235:genetics*; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D053823:Hedgehog Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D014411:Neoplastic Stem Cells / Q000378:metabolism*; D010190:Pancreatic Neoplasms / Q000235:genetics* / Q000378:metabolism; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics; D015203:Reproducibility of Results; D014945:Wound Healing / Q000235:genetics; D000093542:Gemcitabine
chemical_list: D000964:Antimetabolites, Antineoplastic; D015415:Biomarkers; D053823:Hedgehog Proteins; D012333:RNA, Messenger; C509239:SHH protein, human; D003841:Deoxycytidine; D000093542:Gemcitabine