title: Pancreatic Premalignant Lesions Secrete Tissue Inhibitor of Metalloproteinases-1, Which Activates Hepatic Stellate Cells Via CD63 Signaling to Create a Premetastatic Niche in the Liver.
abstract: OBJECTIVE
  Pancreatic ductal adenocarcinoma (PDAC) metastasizes to liver at early stages, making this disease highly lethal. Tissue inhibitor of metalloproteinases-1 (TIMP1) creates a metastasis-susceptible environment in the liver. We investigated the role of TIMP1 and its receptor CD63 in metastasis of early-stage pancreatic tumors using mice and human cell lines and tissue samples.
  
  
  METHODS
  We obtained liver and plasma samples from patients in Germany with chronic pancreatitis, pancreatic intra-epithelial neoplasia, or PDAC, as well as hepatic stellate cells (HSCs). We performed studies with Ptf1a+/Cre;Kras+/LSL-G12D;Trp53loxP/loxP (CPK) mice, Pdx-1+/Cre;Kras+/LSL-G12D;Trp53+/LSL-R172H (KPC) mice, and their respective healthy littermates as control, and Cd63-/-mice with their wild-type littermates. KPC mice were bred with Timp1-/-mice to produce KPCxTimp1-/-mice. TIMP1 was overexpressed and CD63 was knocked down in mice using adenoviral vectors AdTIMP1 or AdshCD63, respectively. Hepatic susceptibility to metastases was determined after intravenous inoculation of syngeneic 9801L pancreas carcinoma cells. Pancreata and liver tissues were collected and analyzed by histology, immunohistochemical, immunoblot, enzyme-linked immunosorbent assay, and quantitative polymerase chain reaction analyses. We analyzed the effects of TIMP1 overexpression or knockdown and CD63 knockdown in transduced human primary HSCs and HSC cell lines.
  
  
  RESULTS
  Chronic pancreatitis, pancreatic intra-epithelial neoplasia, and PDAC tissues from patients expressed higher levels of TIMP1 protein than normal pancreas. The premalignant pancreatic lesions that developed in KPC and CPK mice expressed TIMP1 and secreted it into the circulation. In vitro and in vivo, TIMP1 activated human or mouse HSCs, which required interaction between TIMP1 and CD63 and signaling via phosphatidylinositol 3-kinase, but not TIMP1 protease inhibitor activity. This signaling pathway induced expression of endogenous TIMP1. TIMP1 knockdown in HSCs reduced their activation. Cultured TIMP1-activated human and mouse HSCs began to express stromal-derived factor-1, which induced neutrophil migration, a marker of the premetastatic niche. Mice with pancreatic intra-epithelial neoplasia-derived systemic increases in TIMP1 developed more liver metastases after injections of pancreatic cancer cells than mice without increased levels of TIMP1. This increase in formation of liver metastases from injected pancreatic cancer cells was not observed in TIMP1 or CD63 knockout mice.
  
  
  CONCLUSIONS
  Expression of TIMP1 is increased in chronic pancreatitis, pancreatic intra-epithelial neoplasia, and PDAC tissues from patients. TIMP1 signaling via CD63 leads to activation of HSCs, which create an environment in the liver that increases its susceptibility to pancreatic tumor cells. Strategies to block TIMP1 signaling via CD63 might be developed to prevent PDAC metastasis to the liver.
authors: Institut für Molekulare Immunologie und Experimentelle Onkologie, Technische Universität München, München, Germany.
authors: Barbara
authors: B
authors: Grünwald
authors: Institut für Molekulare Immunologie und Experimentelle Onkologie, Technische Universität München, München, Germany.
authors: Veronika
authors: V
authors: Harant
authors: Institut für Molekulare Immunologie und Experimentelle Onkologie, Technische Universität München, München, Germany.
authors: Susanne
authors: S
authors: Schaten
authors: Institut für Molekulare Immunologie und Experimentelle Onkologie, Technische Universität München, München, Germany.
authors: Monika
authors: M
authors: Frühschütz
authors: Institut für Molekulare Immunologie und Experimentelle Onkologie, Technische Universität München, München, Germany.
authors: Ria
authors: R
authors: Spallek
authors: Institut für Molekulare Immunologie und Experimentelle Onkologie, Technische Universität München, München, Germany.
authors: Bastian
authors: B
authors: Höchst
authors: Institut für Molekulare Immunologie und Experimentelle Onkologie, Technische Universität München, München, Germany.
authors: Katharina
authors: K
authors: Stutzer
authors: Institut für Molekulare Immunologie und Experimentelle Onkologie, Technische Universität München, München, Germany.
authors: Sonja
authors: S
authors: Berchtold
authors: Chirurgische Klinik Technische Universität München, München, Germany.
authors: Mert
authors: M
authors: Erkan
authors: Chirurgische Klinik Technische Universität München, München, Germany.
authors: Olga
authors: O
authors: Prokopchuk
authors: Chirurgische Klinik Technische Universität München, München, Germany.
authors: Marc
authors: M
authors: Martignoni
authors: Institut für Pathologie, Klinikum rechts der Isar, Technische Universität München, München, Germany.
authors: Irene
authors: I
authors: Esposito
authors: Institut für Virologie, Technische Universität München, München, Germany.
authors: Mathias
authors: M
authors: Heikenwalder
authors: II. Medizinische Klinik und Poliklinik, Klinikum rechts der Isar, Technische Universität München, München, Germany.
authors: Aayush
authors: A
authors: Gupta
authors: II. Medizinische Klinik und Poliklinik, Klinikum rechts der Isar, Technische Universität München, München, Germany.
authors: Jens
authors: J
authors: Siveke
authors: Institut für Biochemie, Christian-Albrechts-Universität zu Kiel, Kiel, Germany.
authors: Paul
authors: P
authors: Saftig
authors: Institut für Molekulare Immunologie und Experimentelle Onkologie, Technische Universität München, München, Germany.
authors: Percy
authors: P
authors: Knolle
authors: Institut für Molekulare Immunologie und Experimentelle Onkologie, Technische Universität München, München, Germany.
authors: Dirk
authors: D
authors: Wohlleber
authors: Institut für Molekulare Immunologie und Experimentelle Onkologie, Technische Universität München, München, Germany. Electronic address: achim.krueger@tum.de.
authors: Achim
authors: A
authors: Krüger
mesh_terms: D000818:Animals; D014408:Biomarkers, Tumor / Q000378:metabolism*; D002278:Carcinoma in Situ / Q000378:metabolism / Q000473:pathology; D021441:Carcinoma, Pancreatic Ductal / Q000378:metabolism / Q000473:pathology / Q000556:secondary; D016022:Case-Control Studies; D045744:Cell Line, Tumor; D005260:Female; D055166:Hepatic Stellate Cells / Q000378:metabolism; D006801:Humans; D008113:Liver Neoplasms / Q000378:metabolism / Q000556:secondary; D051379:Mice; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D018345:Mice, Knockout; D009362:Neoplasm Metastasis; D010179:Pancreas / Q000378:metabolism / Q000473:pathology; D010190:Pancreatic Neoplasms / Q000378:metabolism* / Q000473:pathology; D050500:Pancreatitis, Chronic / Q000378:metabolism / Q000473:pathology; D011230:Precancerous Conditions / Q000378:metabolism* / Q000473:pathology; D015398:Signal Transduction; D060149:Tetraspanin 30 / Q000378:metabolism*; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000378:metabolism*; D059016:Tumor Microenvironment
chemical_list: D014408:Biomarkers, Tumor; C554586:CD63 protein, human; C554587:Cd63 protein, mouse; C560822:TIMP1 protein, human; D060149:Tetraspanin 30; C560825:Timp1 protein, mouse; D019715:Tissue Inhibitor of Metalloproteinase-1
keywords: Microenvironment; Signal Transduction; Tissue Inhibitor of Metalloproteinases; Tumor Progression