Rekombinante Expression und Untersuchungen zur Funktion eines Fibroblasten-Wachstumsfaktor-bindenden Proteins(FGF-BP)

Nach Herz-Kreislauferkrankungen stellen maligne Tumoren die zweithäufigste Todesursache in den Industriestaaten dar. Therapeutisch stehen die operative Therapie sowie die Chemo- und Strahlentherapie zur Verfügung. Der Heilungserfolg ist jedoch gerade bei fortgeschrittenen Tumorstadien begrenzt. Ein...

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Bibliographic Details
Main Author: Hagenbusch, Julia
Contributors: Aigner, Achim (Prof. Dr.) (Thesis advisor)
Format: Dissertation
Language:German
Published: Philipps-Universität Marburg 2010
Pharmakologisches Institut
Subjects:
FGF
Online Access:PDF Full Text
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Table of Contents: Malignant tumors are the second weighted cause of death in the western industrial nations after cardiovascular diseases. Therapeutic possibilities include operative methods, chemotherapy and therapeutic radiology. The healing outcome of the advanced stage of tumors however is limited. In the last years a further therapeutic strategy arised based on the specific inhibition of growth factors, receptors or other mitogen factors, which are highly expressed in malignant cells. Accordingly it is important to identify and analyse these target factors to allow a specific therapy of tumor diseases. The focus of this thesis is to study a gene product, which is overexpressed in different malignant tumors: the Fibroblast Growth Factor- binding Protein (FGF-BP). FGF-BP is a heparin binding protein, which interacts with different Fibroblast Growth factors (FGFs), especially FGF-2. It mobilizes FGF-2 from the extracellular matrix and plays an important role in the activation of this growth factor. Using immunhistochemical methods used in this thesis could be shown, that FGF-BP is overexpressed about 30% in ovarial carcinomas. This percentage is comparable with the overexpression ratio of the Her2-Receptor in breast cancer. This finding is background for further studies, which specify the overexpression of FGF-BP in reference to the subtypes and stages of ovarian carcinoma. For the further analysis FGF-BP was recombinantly expressed (rFGF-BP). In this study it was possible to express recombinant FGF-BP at high levels. This rFGF-BP binds FGF-2 as described in previous studies and novel binding partners: FGF-4 and FGF-9. In proliferation assays with SW13-cells a growth promoting effect was presented, which was comparable with stable transfected FGF-BP-SW13 cells. In tumor cells stably transfected with the fusion protein FGF-BP-CFP analysed with the laser scanning microscopy a cytoplasmatic localisation in the absence of FGF-2 was observed. Under coexpression with FGF-2, there was a translocalization from FGF-BP in the nucleus. On the basis of the analysis of different N- respectively C-terminal truncated FGF-BP-mutants it was shown, that the translocation of FGF-BP in the nucleus is dependent from the N-terminus of FGF-BP. To evaluate the biological relevance of these observations proliferation studies with different cell lines were performed. In these studies the dependence of the endogenous expression of FGF-2 – both inhibitory and stimulating effects of FGF-BP in reference to cell growth could be determined. Taken together with the results of the confocal laser microscopy this finding correlates with the balance between FGF-BP and FGF-2. FGF-BP migratesin the presence of FGF-2 into the nucleus, where it stimulates the proliferation. In the absence of FGF-2 it remains in the cytoplasma and displays inhibitory effects. The proliferative effects of FGF-BP thus are dependend on the intact N-terminus, the presence of FGF-2 and the balance between cytoplasmatic and nuclear localized FGF-BP. Based on the proliferation stimulating effects of FGF-BP in FGF-2 positive tumors and the overexpression in ovarian carcinoma demonstrated in this thesis, there is a chance to develop therapeutic strategies e.g. using RNAi-based knockdown strategies.