title: FAP-overexpressing fibroblasts produce an extracellular matrix that enhances invasive velocity and directionality of pancreatic cancer cells.
abstract: BACKGROUND
  Alterations towards a permissive stromal microenvironment provide important cues for tumor growth, invasion, and metastasis. In this study, Fibroblast activation protein (FAP), a serine protease selectively produced by tumor-associated fibroblasts in over 90% of epithelial tumors, was used as a platform for studying tumor-stromal interactions. We tested the hypothesis that FAP enzymatic activity locally modifies stromal ECM (extracellular matrix) components thus facilitating the formation of a permissive microenvironment promoting tumor invasion in human pancreatic cancer.
  
  
  METHODS
  We generated a tetracycline-inducible FAP overexpressing fibroblastic cell line to synthesize an in vivo-like 3-dimensional (3D) matrix system which was utilized as a stromal landscape for studying matrix-induced cancer cell behaviors. A FAP-dependent topographical and compositional alteration of the ECM was characterized by measuring the relative orientation angles of fibronectin fibers and by Western blot analyses. The role of FAP in the matrix-induced permissive tumor behavior was assessed in Panc-1 cells in assorted matrices by time-lapse acquisition assays. Also, FAP+ matrix-induced regulatory molecules in cancer cells were determined by Western blot analyses.
  
  
  RESULTS
  We observed that FAP remodels the ECM through modulating protein levels, as well as through increasing levels of fibronectin and collagen fiber organization. FAP-dependent architectural/compositional alterations of the ECM promote tumor invasion along characteristic parallel fiber orientations, as demonstrated by enhanced directionality and velocity of pancreatic cancer cells on FAP+ matrices. This phenotype can be reversed by inhibition of FAP enzymatic activity during matrix production resulting in the disorganization of the ECM and impeded tumor invasion. We also report that the FAP+ matrix-induced tumor invasion phenotype is β1-integrin/FAK mediated.
  
  
  CONCLUSIONS
  Cancer cell invasiveness can be affected by alterations in the tumor microenvironment. Disruption of FAP activity and β1-integrins may abrogate the invasive capabilities of pancreatic and other tumors by disrupting the FAP-directed organization of stromal ECM and blocking β1-integrin dependent cell-matrix interactions. This provides a novel preclinical rationale for therapeutics aimed at interfering with the architectural organization of tumor-associated ECM. Better understanding of the stromal influences that fuel progressive tumorigenic behaviors may allow the effective future use of targeted therapeutics aimed at disrupting specific tumor-stromal interactions.
authors.affiliation: Department of Medical Oncology, Fox Chase Cancer Center, 333 Cottman Avenue, Philadelphia, Pennsylvania 19111, USA.
authors.forename: Hyung-Ok
authors.initials: HO
authors.lastname: Lee
authors.forename: Stefanie R
authors.initials: SR
authors.lastname: Mullins
authors.forename: Janusz
authors.initials: J
authors.lastname: Franco-Barraza
authors.forename: Matthildi
authors.initials: M
authors.lastname: Valianou
authors.forename: Edna
authors.initials: E
authors.lastname: Cukierman
authors.forename: Jonathan D
authors.initials: JD
authors.lastname: Cheng
mesh_terms: D000230:Adenocarcinoma / Q000201:enzymology / Q000473:pathology*; D000818:Animals; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000473:pathology; D018929:Cell Culture Techniques; D045744:Cell Line, Tumor / Q000201:enzymology / Q000473:pathology; D002465:Cell Movement; D024042:Collagen Type I / Q000378:metabolism; D010450:Endopeptidases; D005109:Extracellular Matrix / Q000502:physiology* / Q000648:ultrastructure; D016326:Extracellular Matrix Proteins / Q000378:metabolism*; D005347:Fibroblasts / Q000201:enzymology*; D005353:Fibronectins / Q000378:metabolism / Q000648:ultrastructure; D051417:Focal Adhesion Kinase 1 / Q000502:physiology; D018093:Gelatinases / Q000235:genetics / Q000502:physiology*; D006801:Humans; D019012:Integrin beta1 / Q000502:physiology; D008565:Membrane Proteins / Q000235:genetics / Q000502:physiology*; D051379:Mice; D008813:Mice, Inbred ICR; D016513:Mice, SCID; D041681:NIH 3T3 Cells / Q000201:enzymology; D009361:Neoplasm Invasiveness / Q000473:pathology*; D009363:Neoplasm Proteins / Q000502:physiology*; D010190:Pancreatic Neoplasms / Q000201:enzymology / Q000473:pathology*; D011993:Recombinant Fusion Proteins / Q000502:physiology; D012697:Serine Endopeptidases / Q000235:genetics / Q000502:physiology*; D059008:Time-Lapse Imaging; D014183:Transplantation, Heterologous; D059016:Tumor Microenvironment / Q000502:physiology*; D000099167:Fibroblast Activation Protein Alpha
chemical_list: D024042:Collagen Type I; D010450:Endopeptidases; D016326:Extracellular Matrix Proteins; D005353:Fibronectins; D051417:Focal Adhesion Kinase 1; D018093:Gelatinases; D019012:Integrin beta1; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D011993:Recombinant Fusion Proteins; D012697:Serine Endopeptidases; D000099167:Fibroblast Activation Protein Alpha; C496050:PTK2 protein, human