A Novel Binding Protein for Fibroblast Growth Factors (FGF-BP2):Cloning, Expression Profile, Tumorigenic Activity and Regulation of Gene Expression by Fetal Bovine Serum and Retinoic Acid.

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Bibliographic Details
Main Author: Schmidt, Joachim
Format: Doctoral Thesis
Published: Philipps-Universität Marburg 2003
Online Access:PDF Full Text
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Angiogenesis is one of the essential alterations in cell physiology that dictate malignant growth and metastasis and has long been a major focus in molecular cancer research. More than a dozen distinct proteins are currently known to induce proliferation of endothelial cells in vitro and/or angiogenesis in vivo. Some of the most effective and best-studied angiogenic factors are fibroblast growth factors (FGFs) that are potent stimulators of new blood vessel formation during tumor growth. However, some FGFs (aFGF and bFGF) are upon secretion immobilized in the extracellular matrix (ECM) and unable to reach their high affinity receptors. There are several possible mechanisms by which bFGF can be released from its matrix storage site and thus activated. One established mechanism is the action of an FGF binding protein (FGF-BP1). FGF-BP1 activates bFGF and is thus able to stimulate tumor cell proliferation and angiogenesis. Recently a human cDNA clone containing an open reading frame for a protein, which shows amino acid sequence similarity of 21% and homology of 41 % to FGF-BP1 was discovered. This novel molecule was named FGF-BP2. In this study the FGF-BP2 expression profile under physiological conditions in normal human tissue and in the pathological setting of tumor cell lines was evaluated by Northern Blotting analysis. Following its in vitro gene regulation by various drugs and growth factors was studied. Finally the FGF-BP2 cDNA was cloned into an expression vector, the gene was overexpressed in the human adrenal carcinoma cell line SW-13 and then analyzed for its tumorigenic potential in soft agar growth assays. This study revealed a widespread FGF-BP2 mRNA expression in normal human tissue. Among 36 tumor cell lines tested though, FGF-BP2 mRNA expression was limited to all of the melanoma cell lines. Interestingly, no FGF-BP2 mRNA expression was detected in normal human neonatal melanocytes. Furthermore it was evaluated if some of the well defined mechanisms for gene regulation that are known for FGF-BP1 also exist for FGF-BP2 as tested in two melanoma cell lines. However, in contrast to what is known for FGF-BP1 it was neither possible to show a significant up-regulation of the FGF-BP2 mRNA expression by Fetal Bovine Serum, EGF and TPA, nor was any down-regulation inducible by all-trans retinoic acid. Finally it was shown that FGF-BP2 overexpression in the human adrenal carcinoma cell line SW-13 promotes a bFGF-dependent colony formation in vitro, indicating its tumorigenic potential. This study demonstrates that the novel FGF-binding protein has not only structural but also functional similarities to FGF-BP1. Its biological activity, together with other data that are not presented here suggest that FGF-BP2 is like FGF-BP1 a tumor promoting agent that may solubilize matrix bound bFGF. However, FGF-BP2 shows a different expression pattern from FGF-BP1 and does not seem to follow similar mechanisms of transcriptional or posttranscriptional gene regulation. Most interesting though seems to be the function that FGF-BP2 might have in melanoma progression since it is known that bFGF and its activated signal pathway play a crucial role in the development of melanoma.