title: Targeting Epithelial-Mesenchymal Transition for Identification of Inhibitors for Pancreatic Cancer Cell Invasion and Tumor Spheres Formation.
abstract: BACKGROUND
  Pancreatic cancer has an enrichment of stem-like cancer cells (CSCs) that contribute to chemoresistant tumors prone to metastasis and recurrence. Drug screening assays based on cytotoxicity cannot identify specific CSC inhibitors, because CSCs comprise only a small portion of cancer cell population, and it is difficult to propagate stable CSC populations in vitro for high-throughput screening (HTS) assays. Based on the important role of cancer cell epithelial-to-mesenchymal transition (EMT) in promoting CSCs, we hypothesized that inhibition of EMT can be a useful strategy for inhibiting CSCs, and therefore a feasible approach for HTS can be built for identification of CSC inhibitors, based on assays detecting EMT inhibition.
  
  
  METHODS
  An immunofluorescent assay was established and optimized for HTS to identify compounds that enhance E-cadherin expression, as a hallmark of inhibition of EMT. Four chemical libraries containing 41,472 compounds were screened in PANC-1 pancreatic cancer cell line. Positive hits were validated for EMT and CSC inhibition in vitro using sphere formation assay, western blotting, immune fluorescence, and scratch assay.
  
  
  RESULTS
  Initial hits were refined to 73 compounds with a secondary screening, among which 17 exhibited concentration dependent induction of E-cadherin expression. Six compounds were selected for further study which belonged to 2 different chemical structural clusters. A novel compound 1-(benzylsulfonyl) indoline (BSI, Compound #38) significantly inhibited pancreatic cancer cell migration and invasion. BSI inhibited histone deacetylase, increased histone 4 acetylation preferably, resulting in E-cadherin up-regulation. BSI effectively inhibited tumor spheres formation. Six more analogues of BSI were tested for anti-migration and anti-CSC activities.
  
  
  CONCLUSIONS
  This study demonstrated a feasible approach for discovery of agents targeting EMT and CSCs using HTS, and identified a class of novel chemicals that could be developed as anti-EMT and anti-CSC drug leads.
authors.affiliation: Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, 3901 Rainbow Blvd., Kansas City, Kansas, United States of America.
authors.forename: Kishore
authors.initials: K
authors.lastname: Polireddy
authors.affiliation: Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, 3901 Rainbow Blvd., Kansas City, Kansas, United States of America.
authors.forename: Ruochen
authors.initials: R
authors.lastname: Dong
authors.affiliation: High-Throughput Screening Core Facility, Structural Biology Center, University of Kansas, Lawrence, Kansas, United States of America.
authors.forename: Peter R
authors.initials: PR
authors.lastname: McDonald
authors.affiliation: Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, 3901 Rainbow Blvd., Kansas City, Kansas, United States of America.
authors.forename: Tao
authors.initials: T
authors.lastname: Wang
authors.affiliation: Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, 3901 Rainbow Blvd., Kansas City, Kansas, United States of America.
authors.forename: Brendan
authors.initials: B
authors.lastname: Luke
authors.affiliation: Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, 3901 Rainbow Blvd., Kansas City, Kansas, United States of America.
authors.forename: Ping
authors.initials: P
authors.lastname: Chen
authors.affiliation: High-Throughput Screening Core Facility, Structural Biology Center, University of Kansas, Lawrence, Kansas, United States of America.
authors.forename: Melinda
authors.initials: M
authors.lastname: Broward
authors.affiliation: High-Throughput Screening Core Facility, Structural Biology Center, University of Kansas, Lawrence, Kansas, United States of America.
authors.forename: Anuradha
authors.initials: A
authors.lastname: Roy
authors.affiliation: Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, 3901 Rainbow Blvd., Kansas City, Kansas, United States of America.
authors.forename: Qi
authors.initials: Q
authors.lastname: Chen
mesh_terms: D051927:CD24 Antigen / Q000378:metabolism; D015820:Cadherins / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D000071858:Epithelial Cell Adhesion Molecule / Q000378:metabolism; D058750:Epithelial-Mesenchymal Transition / Q000187:drug effects*; D057166:High-Throughput Screening Assays; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism; D008856:Microscopy, Fluorescence; D010190:Pancreatic Neoplasms / Q000378:metabolism / Q000473:pathology; D054852:Small Molecule Libraries / Q000737:chemistry / Q000633:toxicity*; D000071250:Snail Family Transcription Factors / Q000037:antagonists & inhibitors / Q000378:metabolism; D062826:Zonula Occludens-1 Protein / Q000235:genetics / Q000378:metabolism
chemical_list: D051927:CD24 Antigen; D015820:Cadherins; C499379:EPCAM protein, human; D000071858:Epithelial Cell Adhesion Molecule; D018960:Hyaluronan Receptors; D054852:Small Molecule Libraries; D000071250:Snail Family Transcription Factors; C569042:TJP1 protein, human; D062826:Zonula Occludens-1 Protein