CD40 ligand (CD40L, CD154), a transmembrane protein structurally related to the cytokine TNF-alpha, was originally identified on stimulated CD4+ T cells, and later on stimulated mast cells and basophils. Interaction of CD40L on T cells with CD40 on B cells is of paramount importance for the development and function of the humoral immune system. CD40 is not only constitutively present on B cells, but it is also found on monocytes, macrophages and endothelial cells, suggesting that CD40L has a broader function in vivo. We now report that platelets express CD40L within seconds of activation in vitro and in the process of thrombus formation in vivo. Like TNF-alpha and interleukin-1, CD40L on platelets induces endothelial cells to secrete chemokines and to express adhesion molecules, thereby generating signals for the recruitment and extravasation of leukocytes at the site of injury. Our results indicate that platelets are not only involved in haemostasis but that they also directly initiate an inflammatory response of the vessel wall.

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Acetyl-boswellic acids (acetyl-BA) are pentacyclic triterpenes derived from the gum resin of frankincense. We have previously shown that these compounds are effective cytotoxic agents, acting through a mechanism that appears to involve the inhibition of topoisomerase activity. We have now investigated the mechanism of action of acetyl-BA and show that these compounds are more potent inhibitors of human topoisomerases I and IIalpha than camptothecin, and amsacrine or etoposide, respectively. Our data demonstrate that acetyl-BA and, to a lesser extent, some other pentacyclic triterpenes, such as betulinic acid, ursolic acid, and oleanolic acid, inhibit topoisomerases I and IIalpha through a mechanism that does not involve stabilization of the cleavable complex or the intercalation of DNA. Surface plasmon resonance analysis revealed that topoisomerases I and IIalpha bind directly to an immobilized derivative of acetyl-BA. This acetyl-BA derivative interacts with human topoisomerases through high-affinity binding sites yielding K(D) values of 70.6 nM for topoisomerase I and 7.6 nM for topoisomerase IIalpha. Based on our data, we propose that acetyl-BA inhibit topoisomerases I and IIalpha through competition with DNA for binding to the enzyme. Thus, acetyl-BA are a unique class of dual catalytic inhibitors of human topoisomerases I and IIalpha.

Prospective identification of patients whose chronic myeloid leukemia (CML) will progress to blast crisis is currently not possible. PP2A is a phosphatase and tumor suppressor that regulates cell proliferation, differentiation, and survival. Cancerous inhibitor of PP2A (CIP2A) is a recently described inhibitor of PP2A in breast and gastric cancer. The aim of this study was to investigate whether CIP2A played a role in CML and whether PP2A or its inhibitor proteins CIP2A or SET could predict clinical outcome. At the time of diagnosis of CML, patients who will later progress to blast crisis have significantly higher levels of CIP2A protein (P < .0001) than patients who do not progress, suggesting that PP2A is functionally inactive. We show that the potential mechanism for disease progression is via altered phosphorylation of the oncogene c-Myc. Knockdown of CIP2A results in increased PP2A activity, decreased c-Myc levels, and a decrease in BCR-ABL1 tyrosine kinase activity. We demonstrate that CIP2A levels at diagnosis can consistently predict patients who will progress to blast crisis. The data show that CIP2A is biologically and clinically important in CML and may be a novel therapeutic target. (Blood. 2011;117(24):6660-6668) IntroductionChronic myeloid leukemia (CML) is a malignant disease of the primitive hematologic cell, characterized by inappropriate expansion of myeloid cells. Although this disease is readily controlled by imatinib, approximately one-third of patients will eventually fail treatment 1,2 ; and a significant proportion of these will progress toward blast crisis (BC), which is usually rapidly fatal. Poor response to imatinib and progression to BC have been linked to high BCR-ABL1 tyrosine kinase activity, 3 but why one patient can remain in well-controlled chronic phase for decades whereas another may rapidly progress to BC is poorly understood.The BCR-ABL1 tyrosine kinase in CML is responsible for growth and survival of the malignant cells through activation of signaling pathways, such as the mitogen-activated protein kinase cascade and the PI3K pathway. 4,5 A major cellular serine/threonine phosphatase working to down-regulate activation of these pathways is the tumor suppressor protein phosphatase 2A (PP2A). 6 In CML cells, PP2A is a key target of BCR-ABL1 signaling; this protein becomes inactivated in these cells because BCR-ABL1 stimulates prevention of its auto-dephosphorylation at tyrosine 307 . 7,8 Maintenance of pY 307 -PP2A levels in CML cells feeds back to BCR-ABL1 and facilitates increased and sustained kinase activity. Inhibition of BCR-ABL1 by imatinib results in reactivation of PP2A, inducing both suppression of growth and enhanced apoptosis of the leukemic cells. 8 However, it is unknown whether Y 307 in PP2A is a direct substrate of BCR-ABL1, and the mechanism regulating phosphorylation of PP2A at this site is not clearly defined.One proposal for the mechanism through which BCR-ABL1 regulates PP2A activity in CML cells involves expression of the PP2A inhibitor protein SET. In ...

The human protein kinome comprises 535 proteins that, with the exception of approximately 50 pseudokinases, control intracellular signaling networks by catalyzing the phosphorylation of multiple protein substrates. While a major research focus of the last 30 years has been cancer-associated Tyr and Ser/Thr kinases, over 85% of the kinome has been identified to be dysregulated in at least one disease or developmental disorder. Despite this remarkable statistic, for the majority of protein kinases and pseudokinases, there are currently no inhibitors progressing toward the clinic, and in most cases, details of their physiologic and pathologic mechanisms remain at least partially obscure. By curating and annotating data from the literature and major public databases of phosphorylation sites, kinases, and disease associations, we generate an unbiased resource that highlights areas of unmet need within the kinome. We discuss strategies and challenges associated with characterizing catalytic and noncatalytic outputs in cells, and describe successes and new frontiers that will support more comprehensive cancer-targeting and therapeutic evaluation in the future. .