Purpose: We have previously reported that actin bundling protein fascin overexpression was associated with poor prognosis, and its stable downregulation reduces cell motility and invasiveness in the ESCC (Clin Cancer Res 2005). In this study, we have examined whether fascin is a potential tumor target using a mouse model and in vitro studies.
Experimental design: Previously, having used a specific shRNA vector, we have established a stable transfectant (F4) with down-regulated fascin from KYSE 170 ESCC cell line. In the present study, using the stable cell line F4, we elucidated the role of fascin in cell growth, adhesion to extracellular matrix and induction of apoptosis in vitro experiments. In addition, we revealed the effect of down-regulation of fascin on the tumor growth in vivo tumor formation assay.
Results: The stable transfectant F4 showed a significantly slower growth pattern by 40.3% (p<0.01) and more frequent detachment from collagen-coated plates by 53.6% (p<0.01), compared to mock transfectants, indicating that fascin plays a role in cell growth by maintaining the proper adhesion of cells to extracellular matrix. In addition, F4 cells showed an increase in apoptotic cells by FACS analysis. Subsequently, we confirmed that the loss of adhesion was not induced by apoptosis, but cause apoptotic changes, since the use of general caspase inhibitor, Z-VAD-FMK, did not inhibit the degree of adhesion observed in F4 cell lines. In vivo tumor formation assay carried out by inoculating F4 and mock cells subcutaneously in nude mice, the tumor size of F4 was significantly smaller than that of mock by 95% at 30 days after inoculation, revealing that fascin also plays an important role in tumor growth in vivo environment. Furthermore, F4 tumor showed a higher level of caspase3 expression, confirming the hypothesis that the loss of extracellular attachment by down-regulation of fascin may induce apoptotic changes, leading to the inhibition of tumor growth.
Conclusions: These results suggest that fascin overexpression plays an important role in tumor progression in ESCC and that the cell death caused by its downregulation was induced by the loss of cell adhesion, anoikis. These results indicate that fascin could be a potential target gene in ESCC therapy.
98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA