GX15-070 (Obatoclax), a Bcl-2 family proteins inhibitor engenders apoptosis and pro-survival autophagy and increases Chemosensitivity in neuroblastoma.

BACKGROUND: Neuroblastoma (NB) is a frequent pediatric tumor associated with poor prognosis. The disregulation of Bcl-2, an anti-apoptotic protein, is crucial for the tumoral development and chemoresistance. Autophagy is also implicated in tumor cell survival and chemoresistance. The aim of our study was to demonstrate therapeutic efficiency of GX 15-070, a pan-Bcl-2 family inhibitor, used alone and in combination with conventional drugs or with hydroxychloroquine (HCQ), an autophagy inhibitor. METHODS: Five neuroblastoma cell lines were tested for the cytotoxic activity of GX 15-070 alone or in combination with cisplatin, doxorubicin, HCQ or Z-VAD-FMK a broad-spectrum caspase inhibitor. Apoptosis and autophagy levels were studied by western-blot and FACS. Orthotopic injections were performed on NOD/LtSz-scid/IL-2Rgamma null mice that were treated with either GX 15-070 alone or in combination with HCQ. RESULTS: Synergistic cytotoxicity was observed for the drug combination in all of the 5 neuroblastoma cell lines tested, including MYCN amplified lines and in cancer stem cells. GX 15-070 significantly increased apoptosis and autophagy in neuroblastoma cells as evidenced by increased levels of the autophagy marker, LC3-II. Inhibition of autophagy by HCQ, further increased the cytotoxicity of this combinatorial treatment, suggesting that autophagy induced by these agent plays a cytoprotective role. In vivo, GX 15-070 combined with HCQ significantly decreased the growth of the tumor and the number of distant metastases. CONCLUSIONS: Based on the synergistic effect of HCQ and GX 15-070 observed in this study, the combination of these two drugs may be utilized as a new therapeutic approach for neuroblastoma.

Management of skull fractures in children less than 1 year of age.

BACKGROUND: Management of skull fracture (SF) in pediatric patients varies from observation in the emergency department (ED) to floor admission. Since 2010, a protocol for admitting children with SF specifically to the trauma service was implemented at our institution. The purpose of our study was to review the management of children with SF younger than 1 year of age. METHODS: Retrospective chart review of all patients between 0 and 1year of age seen in our ED for a SF was done from 2010 to 2013. RESULTS: A total of 180 patients with a mean age of 4.5months (1day-12months) were identified. Of these, 131 patients (73%) were admitted. Mean length of stay was 1.6days. Admitted patients had more depressed (21 vs. 8%) and diastatic (43 vs. 14%) fractures. Fifty-seven children had intracranial hemorrhages (32%) but only 8 patients required non-emergent surgery for depressed fractures. Admission to the trauma service increased from none to 76% with phone follow-ups increasing from 12% to 91%. CONCLUSIONS: Instituting a protocol allowed a safer management of patients with SF. Moreover, we argue that asymptomatic infants with isolated SF can be safely discharged home after brief observation in the ED.

Genotype analysis of tumor-initiating cells expressing CD133 in neuroblastoma.

Neuroblastoma (NB) is the most common and lethal extracranial solid tumor of childhood. Despite aggressive therapy, more than half of the children with advanced NB will die of uncontrolled metastatic disease. After chemotherapy, tumor-initiating cells (TICs) could persist, cause relapses and metastasis. The aim of this study is to demonstrate the tumor-initiating properties of CD133high NB cells and to identify new specific genetic abnormalities. Isolation of the CD133high cell population from NB cell lines was followed by neurosphere formation, soft agar assays, and orthotopic injections in NOD/SCID/IL2Rγc-null mice. A differential genotyping analysis was performed with Affymetrix SNP 6.0 arrays on CD133low and CD133high populations and the frequency of the abnormalities of 36 NB tumors was determined. Our results show that CD133high NB cells possess tumor-initiating properties, as CD133high cells formed significantly more neurospheres and produced significantly more colonies in soft agar than CD133low. Injection of 500 CD133high cells was sufficient to generate primary tumors and frequent metastases in mice. Differential genotyping analysis demonstrated two common regions with gains (16p13.3 and 19p13.3) including the gene EFNA2 in the CD133high population, and two with loss of heterozygosity (16q12.1 and 21q21.3) in the CD133low population. The gain of EFNA2 correlated with increased expression of the corresponding protein. These abnormalities were found in NB samples and some were significantly correlated with CD133 expression. Our results show that CD133high NB cells have TICs properties and present different genotyping characteristics compared to CD133low cells. Our findings reveal insights into new therapeutic targets in NB TICs.