Tumor-expressed FVII-dependent expression of prometastatic genes. (A) Based on microarray analysis of MDA^pcDNA and MDA^FVII cells (n = 4), upstream pathways were predicted using Ingenuity Pathway Analysis. (B) Activity of the canonical TGF-β pathway in MDA^pcDNA and MDA^FVII cells as measured using a construct containing a SMAD3/4-controlled luciferase insert (CAGA-Luc; n = 3). (C) Pathways and processes as predicted with Enrichr. (D) Activity of the Wnt pathway in MDA^pcDNA and MDA^FVII cells as measured using a construct containing a β-catenin–controlled luciferase insert (Bat-Luc; n = 3). (E) Expression of Wnt pathway constituents Axin2 and LEF1 as measured using quantitative polymerase chain reaction (qPCR) in wild-type MDA-MB-231 cells (n = 3). (F) Expression of Wnt pathway constituents as measured using qPCR in MDA-MB-453 after shRNA approaches or FVII antibody blockade (n = 5). (G) Expression of Wnt pathway constituents in MDA-MB-468 after FVII antibody blockade (n = 5). (H) Western blot analysis of Slug, Sox9, and vimentin in MDA^pcDNA and MDA^FVII cells. GAPDH was used as a loading control. (I) mRNA expression of EMT factors and Sox9 in MDA^pcDNA and MDA^FVII, as determined using qPCR (n = 4). (J) Expression of Slug and Sox9 in MDA-MB-453 after shRNA or antibody treatment (n = 5). (K) Expression of Snail and Sox9 in MDA-MB-468 after FVII antibody treatment (n = 5). All graphs show the mean and SD. Statistically significant differences were tested using t tests. Con, control; TNF, tumor necrosis factor; IL-1, interleukin 1; VEGF, vascular endothelial growth factor.