Cancer cells are defined by their ability to invade through the basement membrane, a critical step during metastasis. While increased secretion of proteases, which facilitates degradation of the basement membrane, and alterations in the cytoskeletal architecture of cancer cells have been previously studied, the contribution of the mechanical properties of cells in invasion is unclear. Here, we applied a magnetic tweezer system to establish that stiffness of patient tumor cells and cancer cell lines inversely correlates with migration and invasion through three-dimensional basement membranes, a correlation known as a power law. We found that cancer cells with the highest migratory and invasive potential are five times less stiff than cells with the lowest migration and invasion potential. Moreover, decreasing cell stiffness by pharmacologic inhibition of myosin II increases invasiveness, whereas increasing cell stiffness by restoring expression of the metastasis suppressor TβRIII/betaglycan decreases invasiveness. These findings are the first demonstration of the power-law relation between the stiffness and the invasiveness of cancer cells and show that mechanical phenotypes can be used to grade the metastatic potential of cell populations with the potential for single cell grading. The measurement of a mechanical phenotype, taking minutes rather than hours needed for invasion assays, is promising as a quantitative diagnostic method and as a discovery tool for therapeutics. By showing that altering stiffness predictably alters invasiveness, our results indicate that pathways regulating these mechanical phenotypes are novel targets for molecular therapy of cancer. Cancer Res; 71(15); 5075–80. ©2011 AACR.

While loss of antioxidant expression and the resultant oxidant-dependent damage to cellular macromolecules is key to tumorigenesis, it has become evident that effective oxidant scavenging is conversely necessary for successful metastatic spread. This dichotomous role of antioxidant enzymes in cancer highlights their context-dependent regulation during different stages of tumor development. A prominent example of an antioxidant enzyme with such a dichotomous role and regulation is the mitochondria-localized manganese superoxide dismutase SOD2 (MnSOD). SOD2 has both tumor suppressive and promoting functions, which are primarily related to its role as a mitochondrial superoxide scavenger and H₂O₂ regulator. However, unlike true tumor suppressor- or onco-genes, the SOD2 gene is not frequently lost, or rarely mutated or amplified in cancer. This allows SOD2 to be either repressed or activated contingent on context-dependent stimuli, leading to its dichotomous function in cancer. Here, we describe some of the mechanisms that underlie SOD2 regulation in tumor cells. While much is known about the transcriptional regulation of the SOD2 gene, including downregulation by epigenetics and activation by stress response transcription factors, further research is required to understand the post-translational modifications that regulate SOD2 activity in cancer cells. Moreover, future work examining the spatio-temporal nature of SOD2 regulation in the context of changing tumor microenvironments is necessary to allows us to better design oxidant- or antioxidant-based therapeutic strategies that target the adaptable antioxidant repertoire of tumor cells.

Loss of expression of the TGF-β superfamily coreceptor, the type III TGF-β receptor (TβRIII or betaglycan), occurs in a broad spectrum of human cancers including breast, lung, ovarian, pancreatic, prostate, and renal cell cancer. TβRIII suppresses cancer progression in vivo, at least in part, by reducing cancer cell motility. However, the mechanism by which TβRIII regulates migration is unknown. Here, we demonstrate an unexpected TGF-β signaling independent role for TβRIII in activating Cdc42, altering the actin cytoskeleton and reducing directional persistence to inhibit random migration of both cancer and normal epithelial cells. Functionally, TβRIII through its interaction with the scaffolding protein β-arrestin2, activates Cdc42 and inhibits migration. These studies identify a TGF-β independent homeostatic function for TβRIII in regulating cell migration.

Both the transforming growth factor β (TGF‐β) and integrin signalling pathways have well‐established roles in angiogenesis. However, how these pathways integrate to regulate angiogenesis is unknown. Here, we show that the extracellular matrix component, fibronectin, and its cellular receptor, α5β1 integrin, specifically increase TGF‐β1‐ and BMP‐9‐induced Smad1/5/8 phosphorylation via the TGF‐β superfamily receptors endoglin and activin‐like kinase‐1 (ALK1). Fibronectin and α5β1 integrin increase Smad1/5/8 signalling by promoting endoglin/ALK1 cell surface complex formation. In a reciprocal manner, TGF‐β1 activates α5β1 integrin and downstream signalling to focal adhesion kinase (FAK) in an endoglin‐dependent manner. α5β1 integrin and endoglin form a complex on the cell surface and co‐internalize, with their internalization regulating α5β1 integrin activation and signalling. Functionally, endoglin‐mediated fibronectin/α5β1 integrin and TGF‐β pathway crosstalk alter the responses of endothelial cells to TGF‐β1, switching TGF‐β1 from a promoter to a suppressor of migration, inhibiting TGF‐β1‐mediated apoptosis to promote capillary stability, and partially mediating developmental angiogenesis in vivo. These studies provide a novel mechanism for the regulation of TGF‐β superfamily signalling and endothelial function through crosstalk with integrin signalling pathways.

Reactive oxygen species (ROS) are acknowledged generally to be multi-faceted regulators of cellular functions that trigger various pathological states when present chronically or transiently at non-physiologically high levels. Here we focus on the physiological involvement of ROS in cellular motility, with special emphasis on endothelial cells (EC). An important source of ROS within EC is the non-phagocytic NAD(P)H oxidase, and the small GTPase Rac1 plays a central role in activating this complex. Rac1 is one of the three Rho-family molecules (Rac, Rho and Cdc42) involved in the control of the actin cytoskeleton in response to various signals. In this review we examine the evidence linking ROS production, Rac1 activation and actin organization to EC motility, considering mechanisms for direct interaction of ROS and actin and the effects of ROS on proteins that regulate the actin cytoskeleton. The accumulated evidence suggests that ROS are important regulators of the actin cytoskeletal dynamics and cellular motility, and more in-depth studies are needed to understand the underlying mechanisms.

Unresectable hepatic metastases of colon cancer respond poorly to existing therapies and are a major cause of colon cancer lethality. In this study, we evaluated the therapeutic viability of targeting the mediator kinase CDK8, an early clinical stage drug target, as a means to suppress metastasis of colon cancer. CDK8 was amplified or overexpressed in many colon cancers and CDK8 expression correlated with shorter patient survival. Knockdown or inhibition of CDK8 had little effect on colon cancer cell growth but suppressed metastatic growth of mouse and human colon cancer cells in the liver. This effect was due in part to inhibition of already established hepatic metastases, indicating therapeutic potential of CDK8 inhibitors in the metastatic setting. In contrast, knockdown or inhibition of CDK8 had no significant effect on the growth of tumors implanted subcutaneously, intrasplenically, or orthotopically in the cecum. CDK8 mediated colon cancer growth in the liver through downregulation of matrix metalloproteinase (MMP) inhibitor TIMP3 via TGFβ/SMAD-driven expression of a TIMP3-targeting microRNA, miR-181b, along with induction of Mmp3 in murine or MMP9 in human colon cancer cells via Wnt/β-catenin-driven transcription. These findings reveal a new mechanism for negative regulation of gene expression by CDK8 and a site-specific role for CDK8 in colon cancer hepatic metastasis. Our results indicate the utility of CDK8 inhibitors for the treatment of colon cancer metastases in the liver and suggest that CDK8 inhibitors may be considered in other therapeutic settings involving TGFβ/SMAD or Wnt/β-catenin pathway activation.Significance: These findings demonstrate that inhibition of the transcription-regulating kinase CDK8 exerts a site-specific tumor-suppressive effect on colon cancer growth in the liver, representing a unique therapeutic opportunity for the treatment of advanced colon cancer.