P2-01-23: Seprase Promotes the Growth and Impairs the Migratory Ability of Breast Cancer Cells.

Abstract

Abstract Background: Seprase or Fibroblast Activation Protein alpha (FAP-a) is an integral membrane serine peptidase. It has been shown that it plays an important role in tumour proliferation, migration, invasion and angiogenesis. However, previous work has not satisfactorily explained both the suppression and promotion effects that have been observed. Furthermore, the interaction of Seprase with other membrane molecules or pathway had not been well investigated. This study aimed to investigate the role of Seprase in human breast cancer. Methods: The full length of Seprase cDNA was isolated from normal human prostate tissue using RT-PCR and then cloned into an expression vector (pEF6-V5 His, TOPO). Following confirmation of successful cloning, the plasmid was transfected into the to human breast cancer cell lines, MCF-7 and MDA-MB-231. The function of Separase in these two cell lines was then analysed using cell function assays and ECIS, and the potential interaction with other signalling pathways explored. Results: Compared with the MCF-7 wild type cells (MCF-7wt) and the MCF-7 cells transfected with pEF6/V5 empty vector (MCF-7pef), the MCF7 cells transfected with Seprase (MCF-7exp) had increased growth ability and impaired migratory ability. The growth rate of MCF-7exp cells was 1132.72% on day 5 while those of MCF-7wt and MCF-7pef control cells were 897.24% and 990.99% respectively (p=0.011 and 0.032). The resistance increase of MCF-7exp cells at 2 hours after wounding in ECIS assays was 478.22ohms compare with 1063.75 ohms and 1563.09 ohms in MCF-7wt and MCF-7pef control cells (p<0.05). In MDA-MB-231 cells, over-expression of Seprase resulted in impaired motility. The resistance increase of the MDA-MB-231 wild type cells (MDA-MB-231wt), MDA-MB-231 cells tranfected with pEF6/V5 vector (MDA-MB-231pef), and the MDA-MB-231 cells transfected with Seprase (MDA-MB-231exp) at 3 hours after wounding in ECIS assay were 216.75ohms, 216.00ohms and 120.16ohms respectively. The growth of MDA-MB-231 cells and the adhesion and invasion ability of both MCF-7 cells and MDA-MB-231 cells were not dramatically influenced with Seprase expression. An inhibitor to focal adhesion kinase (FAK) restored the reduced motility ability of both MCF-7exp cells and MDA-MB-231 exp cells. However, inhibitors to PI3K, ERK and ROCK had no influence on it. Conclusion: These results suggest that Seprase promotes proliferation but inhibits the migration of breast cancer cells by potentially regulating the FAK pathway. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P2-01-23.