Purpose miR-409-3p/-5p is a microRNA expressed by embryonic stem cells and its role in cancer biology and metastasis is unknown. Our pilot studies demonstrated elevated miR-409-3p/-5p expression in human prostate cancer bone metastatic cell lines, therefore we defined the biological impact of manipulation of miR-409-3p/-5p in prostate cancer progression and correlated the levels of its expression with clinical human prostate cancer bone metastatic specimens. Experimental Design miRNA profiling of prostate cancer bone metastatic EMT cell line model was performed. Gleason score human tissue array was probed for validation of specific miRNAs. Additionally, genetic manipulation of miR-409-3p/-5p was performed to determine its role in tumor growth, epithelial to mesenchymal transition (EMT) and bone metastasis in mouse models. Results Elevated expression of miR-409-3p/-5p was observed in bone metastatic prostate cancer cell lines and human prostate cancer tissues with higher Gleason scores. Elevated miR-409-3p expression levels correlated with prostate cancer patient progression free survival. Orthotopic delivery of miR-409-3p/-5p in the murine prostate gland induced tumors where the tumors expressed, EMT and stemness markers. Intracardiac inoculation (to mimic systemic dissemination) of miR-409-5p inhibitor treated bone metastatic ARCaPM prostate cancer cells in mice, led to decreased bone metastasis and increased survival compared to control vehicle-treated cells. Conclusion miR-409-3p/-5p plays an important role in prostate cancer biology by facilitating tumor growth, EMT and bone metastasis. This finding bear’s particular translational importance since miR-409-3p/-5p appears to be an attractive biomarker and/or possibly a therapeutic target to treat bones metastatic prostate cancer.

We report here syntenic loci in humans and mice incorporating gene clusters coding for secreted proteins each comprising 10 cysteine residues. These conform to three-fingered protein/Ly-6/urokinase-type plasminogen activator receptor (uPAR) domains that shape three-fingered proteins (TFPs). The founding gene is PATE, expressed primarily in prostate and less in testis. We have identified additional human PATElike genes (PATE-M, PATE-DJ, and PATE-B) that co-localize with the PATE locus, code for novel secreted PATE-like proteins, and show selective expression in prostate and/or testis. Anti-PATE-B-specific antibodies demonstrated the presence of PATE-B in the region

Hepsin, a cell surface protease, is widely reported to be overexpressed in more than 90% of human prostate tumors. Hepsin expression correlates with tumor progression, making it a significant marker and target for prostate cancer. Recently, it was reported that in a prostate cancer mouse model, hepsin up-regulation in tumor tissue promotes progression and metastasis. The underlying mechanisms, however, remain largely uncharacterized. Hepsin transgenic mice displayed reduced laminin-332 (Ln-332) expression in prostate tumors. This is an intriguing cue, since proteolytic processing of extracellular matrix macromolecules, such as Ln-332, is believed to be involved in cancer progression, and Ln-332 expression is lost during human prostate cancer progression. In this study, we provide the first direct evidence that hepsin cleaves Ln-332. Cleavage is specific, since it is both inhibited in a dose-dependent manner by a hepsin inhibitor (Kunitz domain-1) and does not occur when catalytically inactive hepsin is used. By Western blotting and mass spectrometry, we determined that hepsin cleaves the ␤3 chain of Ln-332. N-terminal sequencing identified the cleavage site at ␤3 Arg 245 , in a sequence context (SQLR 245 2LQGSCFC) conserved among species and in remarkable agreement with reported consensus target sequences for hepsin activity. In vitro cell migration assays showed that hepsin-cleaved Ln-332 enhanced motility of DU145 prostate cancer cells, which was inhibited by Kunitz domain-1. Further, hepsin-overexpressing LNCaP prostate cancer cells also exhibited increased migration on Ln-332. Direct cleavage of Ln-332 may be one mechanism by which hepsin promotes prostate tumor progression and metastasis, possibly by up-regulating prostate cancer cell motility.Prostate cancer is the second leading cause of cancer death in men in the United States; according to the American Cancer Society, 186,320 new prostate cancer cases and 28,660 deaths from prostate cancer are projected to occur in 2008 (1). This high rate of mortality is largely due to metastasis of the primary tumor (2). For metastasis to occur, primary tumor cells must breach the basement membrane (BM) 3 by degrading extracellular matrix (ECM) molecules to initiate the invasion process (3). Escaped tumor cells interact with neighboring ECM molecules to promote this activity. This interaction sometimes promotes remodeling of the ECM to create a more conducive environment for tumor cell migration and invasion and thereby aids in cancer progression (4). The remodeling of certain ECM molecules has been reported to occur as a direct result of protease processing, which results in increased tumor cell migration and invasion (5-7).Laminin-332 (Ln-332; previously known as laminin-5), an ECM molecule, is an important component of BM (8). It is a trimeric glycoprotein consisting of disulfide-bonded subunits: ␣3, ␤3, and ␥2 polypeptide chains (9). The importance of Ln-332 in BM assembly was established by the discovery of the occurrence of a lethal skin blistering disorder, jun...

Identification of factors that mediate visceral and bone metastatic spread and subsequent bone remodeling events is highly relevant to successful therapeutic intervention in advanced human prostate cancer (PCa). TBX2, a T-box family transcription factor that negatively regulates cell cycle inhibitor p21, plays critical roles during embryonic development, and recent studies have highlighted its role in cancer. Here we report that TBX2 is overexpressed in human PCa specimens and bone metastases from xenograft mouse models of human PCa. Blocking endogenous TBX2 expression in PC3 and ARCaPM PCa cell models using a dominant negative construct resulted in decreased tumor cell proliferation, colony formation, and invasion in vitro. Blocking endogenous TBX2 in human PCa mouse xenografts decreased invasion and abrogation of bone and soft tissue metastasis. Furthermore, blocking endogenous TBX2 in PCa cells dramatically reduced bone colonizing capability through reduced tumor cell growth and bone remodeling in an intra-tibial mouse model. TBX2 acted in trans by promoting transcription of the canonical WNT (WNT3A) promoter. Genetically rescuing WNT3A levels in PCa cells with endogenously blocked TBX2 partially restored the TBX2-induced PCa metastatic capability in mice. Conversely, WNT3A neutralizing antibodies or WNT antagonist SFRP-2 blocked TBX2 induced invasion. Our findings highlight TBX2 as a novel therapeutic target upstream of WNT3A, where WNT3A antagonists could be novel agents for the treatment of metastasis and for skeletal complications in PCa patients.

Proinflammatory cytokines, especially tumor necrosis factor alpha (TNFalpha), is a pleiotropic mediator of a diverse array of physiologic and neurologic functions and is upregulated during various inflammatory and neurodegenerative diseases. A common survival response during such situations is the increased expression of the hormone insulin-like growth factor 1 (IGF-1). Although it was thought previously that the mechanisms of TNFalpha and IGF-1 action were unrelated, it has been shown that low doses of TNFalpha can inhibit the survival effects of IGF-1 in mouse cerebellar granule neurons. We used a neuronal cell line SH-SY5Y, which underwent apoptosis in response to TNFalpha and this process could be reversed substantially by IGF-1. Crosstalk between signaling pathways of these two factors was found at various points downstream of their signal transduction. To determine the mechanisms of IGF-1-mediated rescue, we looked at the MAP kinases, which are known to be involved in IGF-1 as well as TNFalpha signaling. The c-Jun N-terminal kinase pathway, which is known normally to promote cell death, was found to actually promote survival of TNFalpha-mediated cell death. Inhibiting the c-Jun survival pathway completely reversed the rescue mediated by IGF-1. In addition, the Akt pathway played an equally important role in this rescue.