Overexpression of the urokinase receptor splice variant uPAR-del4/5 in breast cancer cells affects cell adhesion and invasion in a dose-dependent manner and modulates transcription of tumor-associated genes

mRNA levels of the urokinase receptor splice variant uPAR-del4/5 are associated with prognosis in breast cancer. Its overexpression in cancer cells affects tumor biologically relevant processes. In the present study, individual breast cancer cell clones displaying low vs. high uPAR-del4/5 expression were analyzed demonstrating that uPAR-del4/5 leads to reduced cell adhesion and invasion in a dose-dependent manner. Additionally, matrix metalloproteinase-9 (MMP-9) was found to be strongly upregulated in uPAR-del4/5 overexpressing compared to vector control cells. uPAR-del4/5 may thus play an important role in the regulation of the extracellular proteolytic network and, by this, influence the metastatic potential of breast cancer cells

Rab31 expression levels modulate tumor-relevant characteristics of breast cancer cells

BACKGROUND: Rab proteins constitute a large family of monomeric GTP-binding proteins that regulate intracellular vesicle transport. Several Rab proteins, including rab31, have been shown to affect cancer progression and are related with prognosis in various types of cancer including breast cancer. Recently, the gene encoding rab31 was found to be overexpressed in estrogen receptor-positive breast cancer tissue. In a previous study we found a significant association of high rab31 mRNA expression with poor prognosis in node-negative breast cancer patients. In the present study, we aimed to investigate the impact of rab31 (over)-expression on important aspects of tumor progression in vitro and in vivo. METHODS: Breast cancer cells displaying low (MDA-MB-231) or no (CAMA-1) endogenous rab31 expression were stably transfected with a rab31 expression plasmid. Batch-transfected cells as well as selected cell clones, expressing different levels of rab31 protein, were analyzed with regard to proliferation, cell adhesion, the invasive capacity of tumor cells, and in vivo in a xenograft tumor model. Polyclonal antibodies directed to recombinantly expressed rab31 were generated and protein expression analyzed by immunohistochemistry, Western blot analysis, and a newly developed sensitive ELISA. RESULTS: Elevated rab31 protein levels were associated with enhanced proliferation of breast cancer cells. Interestingly, weak to moderate overexpression of rab31 in cell lines with no detectable endogenous rab31 expression was already sufficient to elicit distinct effects on cell proliferation. By contrast, increased expression of rab31 in breast cancer cells led to reduced adhesion towards several extracellular matrix proteins and decreased invasive capacity through Matrigel(TM). Again, the rab31-mediated effects on cell adhesion and invasion were dose-dependent. Finally, in a xenograft mouse model, we observed a significantly impaired metastatic dissemination of rab31 overexpressing MDA-MB-231 breast cancer cells to the lung. CONCLUSIONS: Overexpression of rab31 in breast cancer cells leads to a switch from an invasive to a proliferative phenotype as indicated by an increased cell proliferation, reduced adhesion and invasion in vitro, and a reduced capacity to form lung metastases in vivo

Analyse der tumorbiologischen Bedeutung der Urokinaserezeptor-Spleißvariante uPAR-del4/5 und des GTP-bindenden Proteins rab31 in Brustkrebszelllinien

The mRNA expression of the urokinase receptor splice variant uPAR-del4/5 and the GTP binding protein rab31 are prognostic factors in breast cancer. In order to investigate the tumor-biological role of this proteins in vitro and in vivo, two different breast cancer cell lines were stably transfected with expression plasmids encoding uPAR-del4/5 or rab31, respectively. Cell lines expressing high levels of rab31 showed enhanced cellular growth. In contrast to this, expression of uPAR-del4/5 did not have effects on cell proliferation. The adhesive capacity towards components of the extracellular matrix, as well as the invasive capacity was significantly reduced when uPAR-del4/5 or rab31 was overexpressed. In addition to this, a highly reduced ability of the cells overexpressing either uPAR-del4/5 or rab31 to colonize the lungs in a xenograft mouse-model was observed. These data indicate that both proteins affect tumor-biologically relevant processes in a similar way. But as both represent independent prognostic factors, they may be components of different but possibly associated tumor-relevant signaling pathways.Die mRNA Expression einer Spleißvariante des Urokinase-Rezeptors, uPAR-del4/5, und des GTP-bindenden Proteins rab31 sind prognostische Faktoren für das Mammakarzinom. In der vorliegenden Arbeit wurden beide Proteine einzeln in verschiedenen Brustkrebszelllinien überexprimiert und deren Effekt auf tumorbiologisch relevante Prozesse in vitro und in vivo untersucht. Hier zeigte sich, dass das Wachstum von Brustkrebszelllinien durch eine hohe rab31 Expression angeregt wird, wohingegen die Überexpression von uPAR-del4/5 keinen Effekt zeigte. Für beide Proteine konnte sowohl eine dosisabhängige Reduktion der Adhäsion gegenüber Komponenten der extrazellulären Matrix, als auch ein verringertes Invasionsvermögen gezeigt werden. In einem Xenograft-Mausmodell führte die Überexpression in Brustkrebszelllinien in beiden Fällen zu einer verringerten Lungenkolonisation. Diese Ergebnisse zeigen, dass beide Proteine tumorbiologisch relevante Prozesse in vitro und in vivo in ähnlicher Weise beeinflussen. Da es sich aber um unabhängige prognostische Faktoren handelt, sind diese vermutlich in unterschiedlichen aber möglicherweise assoziierten Signalwegen involviert

Overexpression of the urokinase receptor splice variant uPAR-del4/5 in breast cancer cells affects cell adhesion and invasion in a dose-dependent manner and modulates transcription of tumor-associated genes

mRNA levels of the urokinase receptor splice variant uPAR-del4/5 are associated with prognosis in breast cancer. Its overexpression in cancer cells affects tumor biologically relevant processes. In the present study, individual breast cancer cell clones displaying low vs. high uPAR-del4/5 expression were analyzed demonstrating that uPAR-del4/5 leads to reduced cell adhesion and invasion in a dose-dependent manner. Additionally, matrix metalloproteinase-9 (MMP-9) was found to be strongly upregulated in uPAR-del4/5 overexpressing compared to vector control cells. uPAR-del4/5 may thus play an important role in the regulation of the extracellular proteolytic network and, by this, influence the metastatic potential of breast cancer cells