Nanodiamond as a vector for siRNA delivery to Ewing sarcoma cells

We investigated the ability of diamond nanoparticles (nanodiamonds, NDs) to deliver small interfering RNA (siRNA) in Ewing sarcoma cells, in the perspective of in vivo anti-cancer nucleic acid drug delivery. siRNA was adsorbed onto NDs previously coated with cationic polymer. Cell uptake of NDs has been demonstrated by taking advantage of NDs intrinsic fluorescence coming from embedded color center defects. Cell toxicity of these coated NDs was shown to be low. Consistent with the internalization efficacy, we have shown a specific inhibition of EWS/Fli-1 gene expression at the mRNA and protein level by the ND vectorized siRNA in a serum containing medium

ESF-EMBO symposium "molecular biology and innovative therapies in sarcomas of childhood and adolescence" Sept 29–Oct 4, Polonia Castle Pultusk, Poland

Rhabdomyosarcoma (RMS) and Ewing sarcoma (ES) are among the most common pediatric sarcomas (Arndt et al., 2012). Despite sarcomas representing a highly heterogeneous group of tumors, ES and alveolar RMS (ARMS) typically share one common genetic characteristic, namely a specific chromosomal translocation (Helman and Meltzer, 2003; Lessnick and Ladanyi, 2012). These translocations generate fusion proteins, which are composed of two transcription factors (TF). Typically, one TF is a developmentally regulated factor that is essential for proper specification of a given lineage and provides the DNA-binding domain, while the partner TF contributes a transactivation domain that drives aberrant expression of target genes. Based on these common genetic characteristics, the first ESF-EMBO research conference entitled “Molecular Biology and Innovative Therapies in Sarcomas of Childhood and Adolescence” with special focus on RMS and ES was held at the Polonia Castle in Pultusk, Poland. The conference gathered 70 participants from more than 15 countries and several continents representing most research groups that are active in this field

In silico and in vitro drug screening identifies new therapeutic approaches for Ewing sarcoma.

The long-term overall survival of Ewing sarcoma (EWS) patients remains poor; less than 30% of patients with metastatic or recurrent disease survive despite aggressive combinations of chemotherapy, radiation and surgery. To identify new therapeutic options, we employed a multi-pronged approach using in silico predictions of drug activity via an integrated bioinformatics approach in parallel with an in vitro screen of FDA-approved drugs. Twenty-seven drugs and forty-six drugs were identified, respectively, to have anti-proliferative effects for EWS, including several classes of drugs in both screening approaches. Among these drugs, 30 were extensively validated as mono-therapeutic agents and 9 in 14 various combinations in vitro. Two drugs, auranofin, a thioredoxin reductase inhibitor, and ganetespib, an HSP90 inhibitor, were predicted to have anti-cancer activities in silico and were confirmed active across a panel of genetically diverse EWS cells. When given in combination, the survival rate in vivo was superior compared to auranofin or ganetespib alone. Importantly, extensive formulations, dose tolerance, and pharmacokinetics studies demonstrated that auranofin requires alternative delivery routes to achieve therapeutically effective levels of the gold compound. These combined screening approaches provide a rapid means to identify new treatment options for patients with a rare and often-fatal disease

Pre-B-cell acute lymphoblastic leukemia with bulk extramedullary disease and chromosome 22 (EWSR1) rearrangement masquerading as Ewing sarcoma

We report a 2-year-old female with a subcutaneous tumor who was initially misdiagnosed as suffering from Ewing sarcoma with a positive EWSR1 rearrangement and EWS/FLI1 transcript. After finding lymphoblasts in peripheral blood, the diagnosis of acute lymphoblastic leukemia was established. This necessitated further analysis of the subcutaneous tumor. The tissue was positive for immature B-cell markers and an immunoglobulin heavy chain gene rearrangement, which confirmed the final diagnosis of common type acute lymphoblastic leukemia with bulk extramedullary disease. The patient was treated with chemotherapy and was in remission 30 months after the diagnosis

C/EBPβ-1 promotes transformation and chemoresistance in Ewing sarcoma cells.

CEBPB copy number gain in Ewing sarcoma was previously shown to be associated with worse clinical outcome compared to tumors with normal CEBPB copy number, although the mechanism was not characterized. We employed gene knockdown and rescue assays to explore the consequences of altered CEBPB gene expression in Ewing sarcoma cell lines. Knockdown of EWS-FLI1 expression led to a decrease in expression of all three C/EBPβ isoforms while re-expression of EWS-FLI1 rescued C/EBPβ expression. Overexpression of C/EBPβ-1, the largest of the three C/EBPβ isoforms, led to a significant increase in colony formation when cells were grown in soft agar compared to empty vector transduced cells. In addition, depletion of C/EBPβ decreased colony formation, and re-expression of either C/EBPβ-1 or C/EBPβ-2 rescued the phenotype. We identified the cancer stem cell marker ALDH1A1 as a target of C/EBPβ in Ewing sarcoma. Furthermore, increased expression of C/EBPβ led to resistance to chemotherapeutic agents. In summary, we have identified CEBPB as an oncogene in Ewing sarcoma. Overexpression of C/EBPβ-1 increases transformation, upregulates expression of the cancer stem cell marker ALDH1A1, and leads to chemoresistance

Methanol and isopropanol embryo dosage response curves for wild-type and ethanol-sensitive zebrafish

It is well established that ethanol has an array of negative effects on developing embryos, from craniofacial abnormalities to cognitive deficits and behavioral disorders. Fetal Alcohol Spectrum Disorders (FASD) describes this phenotypic spectrum caused by embryonic ethanol exposure. However, the effects of other small alcohols, such as methanol and isopropanol, have on development are poorly understood. Multiple factors can contribute to the teratogenicity of small alcohols, including timing, dosage and genetic background. Zebrafish (Danio rerio) has been shown to be a powerful model in the study ethanol teratogenesis and can serve as a model to study methanol and isopropanol teratogenicity. Here we provide evidence of the dose response to methanol and isopropanol in a wild type and an ethanol- sensitive mutant zebrafish line. We determine the lethal concentrations of methanol and isopropanol on wild type and ethanol-sensitive mutants. We also show effective dose that leads to malformations of the craniofacial skeleton, including defects to the lower jaw and palate. Our data suggest that ethanol-sensitivity may predict sensitivity to other small alcohols. Overall, our results begin to characterize the effects of methanol and isopropanol on developing embryos.Molecular Bioscience

DAX-1 Expression in Pediatric Rhabdomyosarcomas: Another Immunohistochemical Marker Useful in the Diagnosis of Translocation Positive Alveolar Rhabdomyosarcoma

OBJECTIVES: The aim of this study was to investigate the expression of DAX-1 in a series of pediatric rhabdomyosarcomas (RMS) with known translocation and compare it to Ap2\u3b2, known to be selectively expressed in ARMS. DESIGN: We revised a series of 71 alveolar rhabdomyosarcomas (ARMS), enrolled in the Italian Protocols RMS 79 and 96, and 23 embryonal rhabdomyosarcomas (ERMS) as controls. Before investigating Ap2\u3b2 and DAX-1, ARMS were reviewed and reclassified as 48 ARMS and 23 non-ARMS. RESULTS: Translocation positive ARMS showed a characteristic Ap2\u3b2/DAX-1+ staining pattern in 78% of cases, while 76% of classic ERMS were negative for both. Ap2\u3b2 alone was positive in 3.9% of RMS lacking translocation, whereas DAX-1 alone was positive in 25.4%. Conversely, 9% and 6% of translocation positive ARMS were positive only for DAX-1 or Ap2\u3b2, respectively. The 23 non-ARMS shared the same phenotype as ERMS but had a higher frequency of DAX-1 expression. CONCLUSIONS: DAX-1 is less specific than Ap2\u3b2, however it is a sensitive marker for translocation positive ARMS and can be helpful in their diagnosis if used in combination with Ap2\u3b2

Bruhat Order in the Full Symmetric sln\mathfrak{sl}_n Toda Lattice on Partial Flag Space

In our previous paper [Comm. Math. Phys. 330 (2014), 367-399] we described the asymptotic behaviour of trajectories of the full symmetric $\mathfrak{sl}_n$ Toda lattice in the case of distinct eigenvalues of the Lax matrix. It turned out that it is completely determined by the Bruhat order on the permutation group. In the present paper we extend this result to the case when some eigenvalues of the Lax matrix coincide. In that case the trajectories are described in terms of the projection to a partial flag space where the induced dynamical system verifies the same properties as before: we show that when $t\to\pm\infty$ the trajectories of the induced dynamical system converge to a finite set of points in the partial flag space indexed by the Schubert cells so that any two points of this set are connected by a trajectory if and only if the corresponding cells are adjacent. This relation can be explained in terms of the Bruhat order on multiset permutations