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High intratumoral levels of urokinase‐type plasminogen activator (uPA)‐plasminogen activator inhibitor‐1 (PAI‐1) heteromers predict impaired survival and treatment response in early breast cancer. The pathogenetic role of this protein complex remains obscure. Here, we demonstrate that heteromerization of uPA and PAI‐1 multiplies the potential of the single proteins to attract pro‐tumorigenic neutrophils. To this end, tumor‐released uPA‐PAI‐1 utilizes very low‐density lipoprotein receptor and mitogen‐activated protein kinases to initiate a pro‐inflammatory program in perivascular macrophages. This enforces neutrophil trafficking to cancerous lesions and skews these immune cells toward a pro‐tumorigenic phenotype, thus supporting tumor growth and metastasis. Blockade of uPA‐PAI‐1 heteromerization by a novel small‐molecule inhibitor interfered with these events and effectively prevented tumor progression. Our findings identify a therapeutically targetable, hitherto unknown interplay between hemostasis and innate immunity that drives breast cancer progression. As a personalized immunotherapeutic strategy, blockade of uPA‐PAI‐1 heteromerization might be particularly beneficial for patients with highly aggressive uPA‐PAI‐1high tumors. Synopsis: Uhl et al report that heteromerization of the serine protease urokinase‐type plasminogen activator (uPA) and the serpin plasminogen activator inhibitor‐1 (PAI‐1) enforces the trafficking of pro‐tumorigenic neutrophils to malignant lesions in highly aggressive subtypes of breast cancer. High levels of uPA‐PAI‐1 heteromers in human breast cancer were related to extensive neutrophil infiltration of the malignant tumors and a poor clinical outcome.uPA‐PAI‐1 heteromers induced the release of the cytokine TNF in perivascular macrophages via VLDLr, in turn activating microvascular endothelial cells.Endothelially‐deposited uPA‐PAI‐1 heteromers subsequently supported intravascular adhesion and extravasation of circulating neutrophils by promoting β2 integrin clustering on their cell surface.uPA‐PAI‐1‐primed neutrophils unfolded pro‐tumorigenic properties, potently stimulating breast cancer cell proliferation by the release of neutrophil elastase.Pharmacological blockade of uPA‐PAI‐1 heteromerization by a novel small molecule inhibitor prevented neutrophil infiltration and tumor progression in experimental breast cancer. [ABSTRACT FROM AUTHOR]

ABSTRACT Although carbon-based nanomaterials (CBNs) have been shown to exert prothrombotic effects in microvessels, it is poorly understood whether CBNs also have the potential to interfere with the process of leukocyte-endothelial cell interactions and whether the shape of CBNs plays a role in these processes. Thus, the aim of this study was to compare the acute effects of two differently shaped CBNs, fiber-shaped single-walled carbon nanotubes (SWCNT) and spherical ultrafine carbon black (CB), on thrombus formation as well as on leukocyte-endothelial cell interactions and leukocyte transmigration in the murine microcirculation upon systemic administration in vivo. Systemic administration of both SWCNT and CB accelerated arteriolar thrombus formation at a dose of 1 mg kg-1 body weight, whereas SWCNT exerted a prothrombotic effect also at a lower dose (0.1 mg kg-1 body weight). In vitro, both CBNs induced P-selectin expression on human platelets and formation of platelet-granulocyte complexes. In contrast, injection of fiber-shaped SWCNT or of spherical CB did not induce leukocyte-endothelial cell interactions or leukocyte transmigration. In vitro, both CBNs slightly increased the expression of activation markers on human monocytes and granulocytes. These findings suggest that systemic administration of CBNs accelerates arteriolar thrombus formation independently of the CBNs' shape, but does not induce leukocyte-endothelial cell interactions or leukocyte transmigration. Copyright © 2014 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Background and Purpose: Roscovitine, a cyclin-dependent kinase (CDK) inhibitor that induces tumour cell death, is under evaluation as an anti-cancer drug. By triggering leukocyte apoptosis, roscovitine can also enhance the resolution of inflammation. Beyond death-inducing properties, we tested whether roscovitine affects leukocyte-endothelial cell interaction, a vital step in the onset of inflammation.Experimental Approach: Leukocyte-endothelial cell interactions were evaluated in venules of mouse cremaster muscle, using intravital microscopy. In primary human endothelial cells, we studied the influence of roscovitine on adhesion molecules and on the nuclear factor-κB (NF-κB) pathway. A cellular kinome array, in vitro CDK profiling and RNAi methods were used to identify targets of roscovitine.Key Results: In vivo, roscovitine attenuated the tumour necrosis factor-α (TNF-α)-induced leukocyte adherence to and transmigration through, the endothelium. In vitro, roscovitine strongly inhibited TNF-α-evoked expression of endothelial adhesion molecules (E-selectin, intercellular cell adhesion molecule, vascular cell adhesion molecule). Roscovitine blocked NF-κB-dependent gene transcription, but not the NF-κB activation cascade [inhibitor of κB (IκB) kinase activity, IκB-α degradation, p65 translocation]. Using a cellular kinome array and an in vitro CDK panel, we found that roscovitine inhibited protein kinase A, ribosomal S6 kinase and CDKs 2, 5, 7 and 9. Experiments using kinase inhibitors and siRNA showed that the decreased endothelial activation was due solely to blockade of CDK5 and CDK9 by roscovitine.Conclusions and Implications: Our study highlights a novel mode of action for roscovitine, preventing endothelial activation and leukocyte-endothelial cell interaction by inhibition of CDK5 and 9. This might expand its usage as a promising anti-inflammatory compound. [ABSTRACT FROM AUTHOR]