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Open Access Research

TFPIα and TFPIβ are expressed at the surface of breast cancer cells and inhibit TF-FVIIa activity

Benedicte Stavik123, Mari Tinholt1234, Marit Sletten1, Grethe Skretting23, Per Morten Sandset234 and Nina Iversen1*

Author Affiliations

1 Department of Medical Genetics, Oslo University Hospital and University of Oslo, BOX 4956 Nydalen, Oslo, N-0424, Norway

2 Department of Haematology, Oslo University Hospital, BOX 4956 Nydalen, Oslo, N-0424, Norway

3 Research Institute of Internal Medicine, Oslo University Hospital, BOX 4956 Nydalen, Oslo, N-0424, Norway

4 Institute of Clinical Medicine, University of Oslo, Oslo, Norway

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Journal of Hematology & Oncology 2013, 6:5  doi:10.1186/1756-8722-6-5

Published: 15 January 2013

Abstract

Background

Tissue factor (TF) pathway inhibitor-1 (TFPI) is expressed in several malignant tissues- and cell lines and we recently reported that it possesses anti-tumor effects in breast cancer cells, indicating a biological role of TFPI in cancer. The two main splice variants of TFPI; TFPIα and TFPIβ, are both able to inhibit TF-factor VIIa (FVIIa) activity in normal cells, but only TFPIα circulates in plasma. The functional importance of TFPIβ is therefore largely unknown, especially in cancer cells. We aimed to characterize the expression and function of TFPIα, TFPIβ, and TF in a panel of tumor derived breast cancer cell lines in comparison to normal endothelial cells.

Methods

TFPIα, TFPIβ, and TF mRNA and protein measurements were conducted using qRT-PCR and ELISA, respectively. Cell-associated TFPI was detected after phosphatidylinositol-phospholipase C (PI-PLC) and heparin treatment by flow cytometry, immunofluorescence, and Western blotting. The potential anticoagulant activity of cell surface TFPI was determined in a factor Xa activity assay.

Results

The expression of both isoforms of TFPI varied considerably among the breast cancer cell lines tested, from no expression in Sum149 cells to levels above or in the same range as normal endothelial cells in Sum102 and MDA-MB-231 cells. PI-PLC treatment released both TFPIα and TFPIβ from the breast cancer cell membrane and increased TF activity on the cell surface, showing TF-FVIIa inhibitory activity of the glycosylphosphatidylinositol- (GPI-) anchored TFPI. Heparin treatment released TFPIα without decreasing the cell surface levels, thus indicating the presence of intracellular storage pools of TFPIα in the breast cancer cells.

Conclusion

GPI-attached TFPI located at the surface of breast cancer cells inhibited TF activity and could possibly reduce TF signaling and breast cancer cell growth locally, indicating a therapeutic potential of the TFPIβ isoform.

Keywords:
TFPIα; TFPIβ; TF-FVIIa; Breast cancer cells; Endothelial cells; Heparin; PI-PLC; Anticoagulant activity