Suppression of deacetylase SIRT1 mediates tumor-suppressive NOTCH response and offers a novel treatment option in metastatic Ewing sarcoma.

The developmental receptor NOTCH plays an important role in various human cancers as a consequence of oncogenic mutations. Here we describe a novel mechanism of NOTCH-induced tumor suppression involving modulation of the deacetylase SIRT1, providing a rationale for the use of SIRT1 inhibitors to treat cancers where this mechanism is inactivated because of SIRT1 overexpression. In Ewing sarcoma cells, NOTCH signaling is abrogated by the driver oncogene EWS-FLI1. Restoration of NOTCH signaling caused growth arrest due to activation of the NOTCH effector HEY1, directly suppressing SIRT1 and thereby activating p53. This mechanism of tumor suppression was validated in Ewing sarcoma cells, B-cell tumors, and human keratinocytes where NOTCH dysregulation has been implicated pathogenically. Notably, the SIRT1/2 inhibitor Tenovin-6 killed Ewing sarcoma cells in vitro and prohibited tumor growth and spread in an established xenograft model in zebrafish. Using immunohistochemistry to analyze primary tissue specimens, we found that high SIRT1 expression was associated with Ewing sarcoma metastasis and poor prognosis. Our findings suggest a mechanistic rationale for the use of SIRT1 inhibitors being developed to treat metastatic disease in patients with Ewing sarcoma.

The first European interdisciplinary ewing sarcoma research summit.

The European Network for Cancer Research in Children and Adolescents (ENCCA) provides an interaction platform for stakeholders in research and care of children with cancer. Among ENCCA objectives is the establishment of biology-based prioritization mechanisms for the selection of innovative targets, drugs, and prognostic markers for validation in clinical trials. Specifically for sarcomas, there is a burning need for novel treatment options, since current chemotherapeutic treatment protocols have met their limits. This is most obvious for metastatic Ewing sarcoma (ES), where long term survival rates are still below 20%. Despite significant progress in our understanding of ES biology, clinical translation of promising laboratory results has not yet taken place due to fragmentation of research and lack of an institutionalized discussion forum. To fill this gap, ENCCA assembled 30 European expert scientists and five North American opinion leaders in December 2011 to exchange thoughts and discuss the state of the art in ES research and latest results from the bench, and to propose biological studies and novel promising therapeutics for the upcoming European EWING2008 and EWING2012 clinical trials.

Factors Affecting EWS-FLI1 Activity in Ewing's Sarcoma.

Ewing's sarcoma family tumors (ESFT) are characterized by specific chromosomal translocations, which give rise to EWS-ETS chimeric proteins. These aberrant transcription factors are the main pathogenic drivers of ESFT. Elucidation of the factors influencing EWS-ETS expression and/or activity will guide the development of novel therapeutic agents against this fatal disease.

An alternative transcript from the death-associated protein kinase 1 locus encoding a small protein selectively mediates membrane blebbing.

Death-associated protein kinase 1 (DAPK-1) is a multidomain protein kinase with diverse roles in autophagic, apoptotic and survival pathways. Bioinformatic screens were used to identify a small internal mRNA from the DAPK-1 locus (named s-DAPK-1). This encodes a 295 amino acid polypeptide encompassing part of the ankyrin-repeat domain, the P-loop motifs, part of the cytoskeletal binding domain of DAPK-1, and a unique C-terminal 'tail' extension not present in DAPK-1. Expression of s-DAPK-1 mRNA was detected in a panel of normal human tissues as well as primary colorectal cancers, indicating that its expression occurs in vivo. s-DAPK-1 gene transfection into cells produces two protein products: one with a denatured mass of 44 kDa, and a smaller product of 40 kDa. Double alanine mutation of the C-terminal tail extension of s-DAPK-1 (Gly296/Arg297) prevented production of the 40 kDa fragment, suggesting that the smaller product is generated by in vivo proteolytic processing. The s-DAPK-1 gene cannot substitute for full-length DAPK-1 in an mitogen-activated protein kinase kinase/extracellular signal-regulated kinase-dependent apoptotic transfection assay. However, the transfection of s-DAPK-1 was able to mimic full-length DAPK-1 in the induction of membrane blebbing. The 44 kDa protease-resistant mutant s-DAPK-1G296A/R297A had very low activity in membrane blebbing, whereas the 40 kDa s-DAPK-1Deltatail protein exhibited the highest levels of membrane blebbing. Deletion of the tail extension of s-DAPK-1 increased its half-life, shifted the equilibrium of the protein from cytoskeletal to soluble cytosolic pools, and altered green fluorescent protein-tagged s-DAPK-1 protein localization as observed by confocal microscopy. These data highlight the existence of an alternative product of the DAPK-1 locus, and suggest that proteolytic removal of the C-terminal tail of s-DAPK-1 is required to stimulate maximally its membrane-blebbing function.