Funktionelle Charakterisierung der Rolle von TMPRSS4 und TFPI2 im Pankreaskarzinom

In der Tumorbiologie des hochmalignen Pankreaskarzinoms sind bis heute wenige Schlüsselmutationen bekannt. Die Suche nach Zielproteinen für einen molekularen Therapieansatz stellt eine aktuelle wissenschaftliche Herausforderung dar. TMPRSS4 ist eine transmembranäre Serinprotease, für die - wie für...

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Bibliographic Details
Main Author: Heers, Hendrik
Contributors: Buchholz, Malte (PD Dr.) (Thesis advisor)
Format: Doctoral Thesis
Published: Philipps-Universität Marburg 2013
Online Access:PDF Full Text
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Table of Contents: The tumour biology of pancreatic cancer features few established key mutations until now. The search for novel target proteins in molecular therapy is an important scientific challenge of our days. TMPRSS4 is a transmembranous serine protease with a proven inductive influence on the malignant behaviour of cancer cells, just like many proteins of its class. Pancreatic cancer cells and cells of precursor lesions show an increased gene expression of TMPRSS4. TFPI2 is a protease inhibitor commonly down-regulated in pancreatic cancer. Its function as a tumour suppressor gene is well established for many tumour entities. The strikingly synchronous overexpression of TMPRSS4 and suppression of TFPI2 provoked the assumption of a direct interaction of the two gene products. A corresponding binding site between the proteins was calculated by means of bioinformatics. The assignment on hand was to examine the functional significance of TMPRSS4 and TFPI2 in pancreatic cancer and to determine their hypothetical interaction. The DNA of both genes was cloned into doxycycline-inducible overexpression vectors. Also, shRNA constructs to enable permanent knock-down of expression were provided. Several pancreatic cancer and epithelial cell lines were stably transfected with these constructs in order to conduct functional experiments on proliferation and migration in vitro. Quantitative real time PCR was used to determine the success of transfection at mRNA level. None of the mentioned constructs showed a significant influence on the proliferative cell behaviour during the MTT and BrdU assays that were carried out. To investigate the cell migration the transfected cell lines went through a series of time lapse microscopy experiments. A trend towards inhibition of migration through TMPRSS4 and induction of migration through TFPI2 was found. This effect was only significant for TFPI2 knock-down cells, though. Evidence for a direct interaction of the two genes in question could not be found at all. A significant influence of TMPRSS4 or TFPI2 on the proliferative and migratory potential of pancreatic cancer cells can be negated. Experiments on tissue invasion and metastasis are desirable in this context.