Next we compared the metastatic potential of PR9692 E9, PR9692 E9 mock and PR9692 E9 caRhoA cells. Lungs of animals injected www.selleckchem.com/products/Tipifarnib(R115777).html with PR9692 E9, PR9692 E9 mock or PR9692 E9 caRhoA cells were inspected for the presence of metastases 28, 35 and 45 days post injection. We found that the activation of RhoA signaling in PR9692 E9 caRhoA cells led to a restoration of metastatic potential. The formation of metastases, includ ing all three metastatic categories, was detected in about 55% of animals injected with PR9692 E9 caRhoA cells, while no metastasis was detected in animals injected with PR9692 E9 or PR9692 E9 mock cells. Taken together, these re sults suggest that the activation of Rho ROCK MLC sig naling through the expression of constitutive active RhoA is sufficient to rescue the invasive and metastatic capabil ity of non metastatic PR9692 E9 cells.

Discussion Amoeboid invasiveness in vitro was for the first time de scribed in 2003 and has been studied extensively since then. However, its relevancy for invasiveness and metastasis in vivo is still not clear. Sabeh et al. suggested that the amoeboid invasiveness Inhibitors,Modulators,Libraries of tumor cells observed in vivo can only occur under specific con ditions and may not be an effective and widespread alternative to protease dependent tumor cell migration. Yet, several studies have provided evidence for the plausibility of amoeboid invasion in vivo and Rho ROCK dependent metastasis. Initially, indirect supportive evidence for possible involvement Inhibitors,Modulators,Libraries of amoeboid invasiveness in the process of in vivo metastasis came from clinical studies with ROCK kinase inhibitors.

The ROCK kinase inhibitor fasudil was shown to reduce the dissemination Inhibitors,Modulators,Libraries of cancer in the peritoneal cavity, blood borne metasta sis to the lung, and prevent the establishment of breast tumors in the mammary fat pad. Similarly, Y 27632 was shown to inhibit the tumor growth and intrahepatic metastasis of hepatocellular Inhibitors,Modulators,Libraries carcinoma. In none of these studies, however, were cells with a defined protease independent amoeboid Inhibitors,Modulators,Libraries invasiveness used. At least two independent studies have associated signaling changes leading to microtubule destabilization with the rounded morphology associated with increased invasion and metastatic potential. Hager et al. observed that DIAPH3 silencing in human carcinoma cells resulted in microtubule destabilization, a rounded morphology, and enhanced MYPT1 phosphorylation associated with increased invasive capability and metastatic potential in mice.

Belleti et al. found that stathmin stimulated cell motility through http://www.selleckchem.com/products/Paclitaxel(Taxol).html the extracellular matrix in vitro and increased the metastatic potential of sarcoma cells in vivo. Accordingly, a less phosphorylable stathmin point mutant impaired ECM induced microtubule stabilization and conferred a higher invasive potential, inducing a rounded cell shape coupled with amoeboid like motility in three dimensional matrices.