title: Identification of genes regulating migration and invasion using a new model of metastatic prostate cancer.
abstract: BACKGROUND
  Understanding the complex, multistep process of metastasis remains a major challenge in cancer research. Metastasis models can reveal insights in tumor development and progression and provide tools to test new intervention strategies.
  
  
  METHODS
  To develop a new cancer metastasis model, we used DU145 human prostate cancer cells and performed repeated rounds of orthotopic prostate injection and selection of subsequent lymph node metastases. Tumor growth, metastasis, cell migration and invasion were analyzed. Microarray analysis was used to identify cell migration- and cancer-related genes correlating with metastasis. Selected genes were silenced using siRNA, and their roles in cell migration and invasion were determined in transwell migration and Matrigel invasion assays.
  
  
  RESULTS
  Our in vivo cycling strategy created cell lines with dramatically increased tumorigenesis and increased ability to colonize lymph nodes (DU145LN1-LN4). Prostate tumor xenografts displayed increased vascularization, enlarged podoplanin-positive lymphatic vessels and invasive margins. Microarray analysis revealed gene expression profiles that correlated with metastatic potential. Using gene network analysis we selected 3 significantly upregulated cell movement and cancer related genes for further analysis: EPCAM (epithelial cell adhesion molecule), ITGB4 (integrin β4) and PLAU (urokinase-type plasminogen activator (uPA)). These genes all showed increased protein expression in the more metastatic DU145-LN4 cells compared to the parental DU145. SiRNA knockdown of EpCAM, integrin-β4 or uPA all significantly reduced cell migration in DU145-LN4 cells. In contrast, only uPA siRNA inhibited cell invasion into Matrigel. This role of uPA in cell invasion was confirmed using the uPA inhibitors, amiloride and UK122.
  
  
  CONCLUSIONS
  Our approach has identified genes required for the migration and invasion of metastatic tumor cells, and we propose that our new in vivo model system will be a powerful tool to interrogate the metastatic cascade in prostate cancer.
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Banyard
authors.forename: Ivy
authors.initials: I
authors.lastname: Chung
authors.forename: Matthew
authors.initials: M
authors.lastname: Migliozzi
authors.forename: Derek T
authors.initials: DT
authors.lastname: Phan
authors.forename: Arianne M
authors.initials: AM
authors.lastname: Wilson
authors.forename: Bruce R
authors.initials: BR
authors.lastname: Zetter
authors.affiliation: Vascular Biology Program, Boston Children's Hospital, Karp Family Research Laboratories, 300 Longwood Avenue, 02115 Boston, MA, USA. diane.bielenberg@childrens.harvard.edu.
authors.forename: Diane R
authors.initials: DR
authors.lastname: Bielenberg
mesh_terms: D045744:Cell Line, Tumor; D002465:Cell Movement / Q000235:genetics*; D003094:Collagen; D004338:Drug Combinations; D006801:Humans; D007797:Laminin; D008207:Lymphatic Metastasis; D008297:Male; D009361:Neoplasm Invasiveness / Q000235:genetics*; D009389:Neovascularization, Pathologic / Q000235:genetics* / Q000473:pathology; D011471:Prostatic Neoplasms / Q000235:genetics* / Q000473:pathology; D011509:Proteoglycans; D014568:Urokinase-Type Plasminogen Activator / Q000037:antagonists & inhibitors / Q000235:genetics
chemical_list: D004338:Drug Combinations; D007797:Laminin; D011509:Proteoglycans; C059485:matrigel; D003094:Collagen; D014568:Urokinase-Type Plasminogen Activator