title: The P2X7 receptor regulates cell survival, migration and invasion of pancreatic ductal adenocarcinoma cells.
abstract: BACKGROUND
  Pancreatic ductal adenocarcinoma (PDAC) is presently one of the cancers with the worst survival rates and least effective treatments. Moreover, total deaths due to PDAC are predicted to increase in the next 15 years. Therefore, novel insights into basic mechanism of PDAC development and therapies are needed. PDAC is characterized by a complex microenvironment, in which cancer and stromal cells release different molecules, such as ATP. ATP can be transported and/or exocytosed from active cancer cells and released from dying cells in the necrotic core of the cancer. We hypothesized that one of the ATP receptors, the P2X7 receptor (P2X7R) could be an important player in PDAC behaviour.
  
  
  METHODS
  We determined the expression (real time PCR and Western blot) and localization (immunofluorescence) of P2X7R in human PDAC cell lines (AsPC-1, BxPC-3, Capan-1, MiaPaCa-2, Panc-1) and a "normal" human pancreatic duct epithelial cell line (HPDE). The function of P2X7R in proliferation (BrdU assay), migration (wound assay) and invasion (Boyden chamber with matrigel) was characterized. Furthermore, we studied P2X7R-dependent pore formation (YoPro-1 assay) and cell death (caspase and annexin V / propidium iodide assays).
  
  
  RESULTS
  We found higher expression of P2X7R protein in PDAC compared to HPDE cells. P2X7R had notable disparate effects on PDAC survival. Firstly, high concentrations of ATP or the specific P2X7R agonist, BzATP, had cytotoxic effects in all cell lines, and cell death was mediated by necrosis. Moreover, the P2X7R-pore antagonist, A438079, prevented ATP-induced pore formation and cell death. Second, in basal conditions and with low concentrations of ATP/BzATP, the P2X7R allosteric inhibitor AZ10606120 reduced proliferation in all PDAC cell lines. P2X7R also affected other key characteristics of cancer cell behavior. AZ10606120 reduced cell migration and invasion in PDAC cell lines compared to that of untreated/vehicle-treated control cells, and stimulation with sub-millimolar concentrations of ATP or BzATP substantially increased cell invasion.
  
  
  CONCLUSIONS
  PDAC cell lines overexpress P2X7R and the receptor plays crucial roles in cell survival, migration and invasion. Therefore, we propose that drugs targeting P2X7R could be exploited in therapy of pancreatic cancer.
authors.affiliation: Department of Biology, Section of Cell Biology and Physiology, University of Copenhagen, August Krogh Building, Universitetsparken 13, DK-2100, Copenhagen, Denmark. andrea.giannuzzo@bio.ku.dk.
authors.forename: Andrea
authors.initials: A
authors.lastname: Giannuzzo
authors.affiliation: Department of Biology, Section of Cell Biology and Physiology, University of Copenhagen, August Krogh Building, Universitetsparken 13, DK-2100, Copenhagen, Denmark. sfpedersen@bio.ku.dk.
authors.forename: Stine Falsig
authors.initials: SF
authors.lastname: Pedersen
authors.affiliation: Department of Biology, Section of Cell Biology and Physiology, University of Copenhagen, August Krogh Building, Universitetsparken 13, DK-2100, Copenhagen, Denmark. inovak@bio.ku.dk.
authors.forename: Ivana
authors.initials: I
authors.lastname: Novak
mesh_terms: D000218:Adamantane / Q000031:analogs & derivatives / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000634:Aminoquinolines / Q000494:pharmacology; D017209:Apoptosis; D021441:Carcinoma, Pancreatic Ductal / Q000378:metabolism* / Q000473:pathology; D053148:Caspase 3 / Q000378:metabolism; D053179:Caspase 7 / Q000378:metabolism; D045744:Cell Line, Tumor; D002465:Cell Movement; D002470:Cell Survival; D006801:Humans; D009361:Neoplasm Invasiveness; D010190:Pancreatic Neoplasms / Q000378:metabolism* / Q000473:pathology; D058911:Purinergic P2X Receptor Agonists / Q000494:pharmacology; D058920:Purinergic P2X Receptor Antagonists / Q000494:pharmacology; D011725:Pyridines / Q000494:pharmacology; D058486:Receptors, Purinergic P2X7 / Q000502:physiology*; D013777:Tetrazoles / Q000494:pharmacology
chemical_list: C523668:3-(5-(2,3-dichlorophenyl)-1H-tetrazol-1-yl)methylpyridine; C000602458:AZ10606120; D000634:Aminoquinolines; C546468:P2RX7 protein, human; D058911:Purinergic P2X Receptor Agonists; D058920:Purinergic P2X Receptor Antagonists; D011725:Pyridines; D058486:Receptors, Purinergic P2X7; D013777:Tetrazoles; D000255:Adenosine Triphosphate; C505407:CASP3 protein, human; C505520:CASP7 protein, human; D053148:Caspase 3; D053179:Caspase 7; D000218:Adamantane