More than 80% of KPC mice, but only 30% of FKPC mice, developed abdominal distension due to hemorrhagic ascites ( Figure 6C). On average, KPC mice harbored 1.57 mL ascitic fluid, and FKPC mice showed almost none ( Figure 6C). Metastasis was dramatically reduced in FKPC mice ( Figure 6B and Supplementary Table 4). Around 95% of KPC mice and only 55% of FKPC mice had local metastasis to intestinal mesentery ( Figure 6B, D, and E). Forty-four percent of KPC mice, but only 13% of FKPC mice, developed diaphragm metastasis ( Figure 6B). Similar to local metastasis, 52% of KPC mice and only 13% of FKPC mice showed

distant liver metastasis ( Figure 6B click here and F). Both mesenteric and liver metastases of KPC mice were positive for fascin and p53 ( Figure 6D and F). KPC mice had shorter survival overall than FKPC with liver metastases ( Figure 6E). We conclude that loss of fascin significantly reduces ascites and metastasis to mesentery, diaphragm, and liver. To further investigate the mechanism by which fascin promotes metastasis, we first

examined the actin dynamics of PDAC cells (105768) from the FKPC mice compared with the same cell line rescued with GFP-fascin. GFP-fascin concentrated in filopodia (Figure 7A and FDA approved Drug Library Video 1). Fascin rescue cells showed dynamic filopodia assembly and turnover ( Supplementary Figure 8A and B). Filopodia were significantly less frequent, shorter, and shorter-lived in fascin-deficient cells than fascin-rescued cells ( Supplementary Figure 8B). Lamellipodial dynamics were greater in fascin-rescued cells ( Supplementary Figure 8C and Video 2). Expression of fascin significantly enhanced protrusion frequency, distance protruded, and protrusion rate, and decreased protrusion persistence ( Supplementary Figure 8C). Fascin-rescued PDAC cells migrated faster PJ34 HCl than fascin-deficient cells ( Supplementary Figure 8D and Video 3). Fascin-expression status did not affect growth in 2D or 3D ( Supplementary Figure 8E), similar to PDAC in vivo. In addition, fascin-rescued cells behaved similarly to fascin-deficient cells during anoikis ( Supplementary Figure 8E). Fascin expression increases PDAC cell migration

via lamellipodial and filopodial dynamics, but does not affect growth and survival. Formation of mesenteric and diaphragm metastases involves transmigration of cancer cells through the mesothelial cell (MC) layer.27 and 28 We tested a potential role for fascin in mesothelial transmigration by plating PDAC cells (105768) on top of a monolayer of human Met5a MCs. PDAC cells opened MC junctions and intercalated themselves between MCs (Supplementary Figure 9A). GFP-fascin localized intensively to the filopodia at the leading edge of transmigrating PDAC cells ( Figure 7A and Video 4). About 75% of fascin-rescued PDAC cells, but only 35% of fascin-deficient cells, intercalated by 10 hours ( Figure 7B, Supplementary Figure 9B, and Video 5).