Abstract
Hepatocyte growth factor activator inhibitor type 1 (HAI-1) is a transmembrane protease inhibitor that regulates the activities of membrane-bound and extracellular serine proteases. HAI-1 has two Kunitz-type inhibitor domains with the N-terminal Kunitz domain (KD1) responsible for inhibiting known target proteases. Previously, we reported that knockdown of HAI-1 in the human pancreatic carcinoma cell line SUIT-2 resulted in epithelial to mesenchymal transition. To evaluate the role of HAI-1 in metastasis, we examined the metastatic capability of SUIT-2 cells that did or did not stably express HAI-1 short-hairpin RNA in an experimental pulmonary metastasis assay using nude mice. The extent of pulmonary metastasis was verified by histological examination and direct measurement of human cytokeratin 19 mRNA levels. One week after injecting SUIT-2 cells into mouse tail veins, apparent metastatic colonization was observed in 36% (4/11) of mice injected with HAI-1-knockdown SUIT-2, whereas none (0/11) of the control mice were positive for metastasis. After 2 weeks the metastasis positive ratios were 80% (4/5) and 40% (2/5), and after 4 weeks the ratios were 82% (9/11) and 45% (5/11) for HAI-1-knockdown and control SUIT-2 cells, respectively. Thus, loss of HAI-1 promoted pulmonary metastasis. Co-injection of recombinant KD1 abolished metastasis produced by HAI-1-knockdown SUIT-2 cells after 1 week. Moreover, recombinant KD1 restored E-cadherin levels in HAI-1 knockdown SUIT-2 cells and reduced their invasiveness in vitro. These data indicate that HAI-1 regulates pulmonary metastasis of SUIT-2, and KD1 may have therapeutic application for inhibiting metastatic cancer cell spreading.