Neuroblastic Tumors Research Articles

Abstract 3274: Neuroblastoma express a novel EGFR extracellular mutation, EGFRΔ768, which possesses distinct biological and biochemical properties

Abstract Background: Epidermal Growth Factor Receptor (EGFR) is a biologic target for cancer therapy. Clinical response to EGFR inhibitors is variable. Recently, EGFR mutations were found to predict response to anti-EGFR therapy. Since a few children with Neuroblastoma (NB) benefited from EGFR inhibitors in clinical trials, we screened for EGFR mutations in primary NB. Interestingly, we identified the expression of both EGFRvIII and a novel mutation, EGFRΔ768, in these tumors. In this study, we characterized the EGFRΔ768 mutation by comparing its biological and biochemical effect with that of wild type (WT)-EGFR and EGFRvIII. Hypothesis: EGFRΔ768 expression confers an aggressive cancer phenotype in NB cells Materials and Methods: We obtained 62 Snap Frozen primary neuroblastic tumors from the Columbus Bipathology Repository Center and 80 NB RNA from the Children Oncology Group (COG). Primary NB were prepared for RT-PCR and sequencing analyses using EGFR exon 1 & 8 primers. Five NB cell lines were obtained and confirmed by STR profiling. The EGFRΔ768 and EGFRvIII deletions were subcloned into a WT-EGFR expression vector with a C-terminal GFP tag. Next, stable 3T3 and NB clones expressing WT and mutant EGFR were generated. The clones were visualized under fluorescence microscopy. Invasion and XTT assays were performed with the stable clones. Responses to Erlotinib from 0.01 - 1 ng/ul and Etoposide from 0.5 - 2 uM were compared among the clones by XTT and western analyses. EGFR phosphorylation and downstream signaling pathways were examined by Western Blot and phosphokinase array. Results: We previously reported that 32% (20/62) of the Columbus neuroblastic tumors expressed EGFR extracellular deletion mutants, EGFRΔ768 (an in-frame deletion from nucleotides 102 of exon 2 to 869 of exon 7) and EGFRvIII (Δ801). In the 80 COG NB RNA, 7 expressed EGFRΔ768, 7 expressed EGFRvIII and 1 expressed both. EGFRΔ768 expression was also detected in the NB cell line, BE2M17. This novel mutant localized to the cell surface and enhanced the invasion and proliferation of NB cells in vitro when compared to control GFP+ cells. Its expression also confers sensitivity to Erlotinib, but the biochemical inhibition of Erlotinib on EGFRΔ768 is significantly less effective than that on activated WT-EGFR from 0.01 - 0.1 ng/ul. EGFRΔ768+ cells were also significantly more resistant to Etoposide than the GFP and WT-EGFR+ cells. Although both EGFRΔ768 and EGFRvIII autophosphorylated without ligand, the cellular proliferation rate and baseline autophosphorylation of EGFRΔ768+ cells were significantly higher than those of EGFRvIII+ cells. Finally, we found that EGFRΔ768 did not activate downstream signaling pathways of WT-EGFR such as MAPK or STAT. It has a different tyrosine phosphorylation profile than WT-EGFR. Conclusion: EGFRΔ768 confers aggressive cancer cell behaviors and has distinct biological as well as biochemical properties. Citation Format: James Keller, Kristen A. VanHeyst, Anjaruwee S. Nimnual, Mathew Varghese, Michael J. Hayman, Edward L. Chan. Neuroblastoma express a novel EGFR extracellular mutation, EGFRΔ768, which possesses distinct biological and biochemical properties. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3274. doi:10.1158/1538-7445.AM2015-3274

GIANT GANGLIONEUROMA IN PEDIATRIC PATIENT

Ganglioneuroma is a rare, benign neuroblastic tumor, arising mainly from the central or peripheral autonomic nervous system, especially the sympathetic system. We present the case of a 7 year old male patient, B.S., with a giant asymptomatic ganglioneuroma located in the retroperitoneum.

The CXCR4/CXCR7/CXCL12 Axis Is Involved in a Secondary but Complex Control of Neuroblastoma Metastatic Cell Homing.

Neuroblastoma (NB) is one of the most deadly solid tumors of the young child, for which new efficient and targeted therapies are strongly needed. The CXCR4/CXCR7/CXCL12 chemokine axis has been involved in the progression and organ-specific dissemination of various cancers. In NB, CXCR4 expression was shown to be associated to highly aggressive undifferentiated tumors, while CXCR7 expression was detected in more differentiated and mature neuroblastic tumors. As investigated in vivo, using an orthotopic model of tumor cell implantation of chemokine receptor-overexpressing NB cells (IGR-NB8), the CXCR4/CXCR7/CXCL12 axis was shown to regulate NB primary and secondary growth, although without any apparent influence on organ selective metastasis. In the present study, we addressed the selective role of CXCR4 and CXCR7 receptors in the homing phase of metastatic dissemination using an intravenous model of tumor cell implantation. Tail vein injection into NOD-scid-gamma mice of transduced IGR-NB8 cells overexpressing CXCR4, CXCR7, or both receptors revealed that all transduced cell variants preferentially invaded the adrenal gland and typical NB metastatic target organs, such as the liver and the bone marrow. However, CXCR4 expression favored NB cell dissemination to the liver and the lungs, while CXCR7 was able to strongly promote NB cell homing to the adrenal gland and the liver. Finally, coexpression of CXCR4 and CXCR7 receptors significantly and selectively increased NB dissemination toward the bone marrow. In conclusion, CXCR4 and CXCR7 receptors may be involved in a complex and organ-dependent control of NB growth and selective homing, making these receptors and their inhibitors potential new therapeutic targets.

Primary cervical ganglioneuroblastoma

ObjectivesGanglioneuroblastomas represent a histological subgroup of the rare neuroblastic tumours with intermediate malignant potential arising from neural crest progenitor cells of sympathetic nerves. Diagnosis can often be difficult based on imaging alone. We describe 4 cases of children presenting with a solitary neck mass with histology ultimately revealing ganglioneuroblastoma. MethodsA retrospective case note review was carried out of all patients with cervical ganglioneuroblastoma seen at Great Ormond Street Hospital, UK. ResultsMean age at presentation was 5 years. Based on imaging, the initial diagnoses for three of the cases were: lymphatic malformation, carotid body tumour, paraganglioma, respectively, whilst the remaining case had an immediate incisional biopsy revealing the correct diagnosis. All cases were managed by surgical excision with no evidence of recurrence after a median follow up of 6 years. ConclusionOtolaryngologists should be aware of ganglioneuroblastoma when establishing the differential diagnosis of a child presenting with a neck mass. Biopsy is recommended as the gold standard investigation to avoid an incorrect diagnosis.

Opsoclonus Myoclonus Syndrome: A Case Report

Introduction: Opsoclonus myoclonus syndrome is defined through the following criteria: opsoclonus, myoclonus, and ataxia, often behavioral and sleep problems are associated. Mostly, it is paraneoplastic associated with different kind of cancers, in children mostly with neuroblastic tumors. In discussion is an autoimmune origin.

Image-defined risk factor assessment of neurogenic tumors after neoadjuvant chemotherapy is useful for predicting intra-operative risk factors and the completeness of resection.

Patients with neuroblastoma are now stratified at diagnosis according to the presence and number of image-defined risk factors (IDRFs). We examined the added value of IDRF assessment after neoadjuvant chemotherapy for predicting surgical resection. From 2009-2012, 39 out of 91 patients operated on in our institution for neuroblastic tumors received neoadjuvant chemotherapy based on ongoing SIOPEN protocols or treatment guidelines. IDRFs were assessed both at diagnosis and preoperatively on CT and/or MRI. Median age at diagnosis was 30 months [range 2-191]. The tumor locations were adrenal (n = 20), paravertebral (n = 13) and perivascular (n = 6). INRGSS stages were L2 (n = 13), M (n = 25) and Ms (n = 1). Eleven tumors (28%) were MYCN-amplified. Chemotherapy reduced the number of IDRFs in 54% of patients overall (21/39): 61.5% (16/26) of M and Ms patients, and 38.5% (5/13) of non metastatic patients (P < 0.001). The number of IDRFs lost after chemotherapy was proportional to the degree of tumor shrinkage (P = 0.002), independent of the primary tumor location (P = 0.73), although the number was higher in patients with left versus right adrenal locations (P = 0.004). Patients with neuroblastoma on post-surgical histology lost more IDRFs (median: 1[0-9]) than patients with ganglioneuroblastoma (median: 0[0-4]) (P < 0.001). The completeness of resection was related only to the number of preoperative IDRFs (P = 0.028). IDRF assessment after neoadjuvant chemotherapy is useful for predicting completeness of resection of neurogenic tumors. A larger international study is needed to confirm these results and to explore a possible correlation between preoperative IDRF status and survival.

Cyclin D1 in human neuroblastic tumors recapitulates its developmental expression: An immunohistochemical study

The protein cyclin D1 (CD1), which belongs to a family of proteins functioning as regulators of CDKs (cyclin-dependent kinases) throughout the cell cycle, has been immunohistochemically detected in a wide variety of human malignant tumors. The aim of the present study was to investigate immunohistochemically the expression and distribution of CD1 in the developing human peripheral sympathetic nervous system (PSNS) and in childhood peripheral neuroblastic tumors (neuroblastomas, ganglioneuroblastomas, and ganglioneuromas). The above mentioned fetal and neoplastic tissues represent an in vivo model in which undifferentiated neuroblastic cells undergo ganglion cell differentiation. During development, a strong nuclear expression of CD1 was restricted to neuroblasts, disappearing progressively from the maturing ganglion cells with increasing gestational age. In neoplastic tissues, CD1 immunoreactivity was restricted to neuroblastic cell component of all neuroblastomas and ganglioneuroblastomas, whereas it was absent or only focally detectable in maturing/mature ganglion cell component of differentiating neuroblastomas, ganglioneuroblastomas, and ganglioneuromas. We conclude that CD1 is a reliable marker, which can be used routinely to stain neuroblastic cells in both developing and neoplastic tissues. Furthermore, our results indicate that CD1 expression in childhood peripheral neuroblastic tumors recapitulates the changes during normal development of PSNS, as previously reported for Bcl-2 oncoprotein, c-ErbB2, insulin-like growth factor 2, β-2-microglobulin, and cathepsin D. This is consistent with the current view that childhood peripheral neuroblastic tumors exhibit gene expression profiles mirroring those occurring during PSNS development.

Variations in apparent diffusion coefficient values following chemotherapy in pediatric neuroblastoma.

In children the assessment of solid tumors' response to chemotherapy is based primarily on size reduction, which can be unreliable and a late marker, in the presence of necrosis. We aimed to establish whether apparent diffusion coefficient (ADC) values of childhood neuroblastomas show proportional changes in relation to chemotherapy response. We evaluated 15 pediatric patients with abdominopelvic neuroblastomas, who had undergone MRI before and after chemotherapy. Two radiologists retrospectively analyzed all images by drawing a round uniform region-of-interest in the solid/contrast-enhancing portion of the lesions in consensus. The ADC values from pre- and postchemotherapy images were compared. Postchemotherapy ADC values were significantly higher than those obtained before treatment (P < 0.05, for minimum, maximum, and median ADC values). Our results support diffusion-weighted MRI as a promising noninvasive biomarker of therapeutic responses. To the best of our knowledge, this is the first report to compare diffusion- weighted imaging findings before and after chemotherapy in childhood neuroblastic tumors.

Endoglin (CD 105) as a potential prognostic factor in neuroblastoma.

Endoglin (CD105) is a cytokine that modulates angiogenesis by regulating different cellular functions, including endothelial proliferation, differentiation, migration and formation of microvessels. CD105 is expressed strongly in the tumor vasculature, and intratumoral microvessel density (IMVD), as determined by the use of antibodies to CD105, it has been found to be an important prognostic indicator for outcome in various malignances. This study aims to determine if the clinical outcome of children with neuroblastoma is correlated with IMVD, as determined by CD105 staining and other prognostic factors. Tumor tissue specimens from 38 patients with peripheral neuroblastic tumors who underwent surgical resection or biopsy of their primary tumor without any preoperative therapy were retrospectively reviewed. IMVD was identified immunohistochemically using monoclonal antibodies against CD105. Prognostic factors, such as the MYCN oncogene, disease stage, histopathology and age, were correlated with outcome. Among 38 examined specimens, the median IMVD value was 23.2 (15.1-28.4). The IMVD identified by CD105 was significantly higher in patients with unfavorable histology, metastatic disease, MYCN amplification and COG high risk group. ROC analysis was used to find significant IMVD level regarding EFS. The cut-off >18 was selected according to the greatest sensitivity (100%) and specificity (68.42%). The multivariate Cox proportional hazards analysis demonstrated that MYCN amplification and IMVD were significant prognostic factors in predicting EFS (hazard ratio for MYCN amplification: 3.61; 95% CI: 1.20-10.90; P = 0.023 and for IMVD: 1.05; 95% CI: 1.00-1.09; P = 0.037). IMVD determined by CD105 appeared to be an independent prognostic factor for neuroblastoma.

Prognostic significance of MYCN amplification in children neuroblastic tumors

To summarize the clinicopathologic features of neuroblastic tumors (NT), and to explore the prognostic significance of MYCN amplification in NT. The clinicopathologic data of 267 NT were reviewed. MYCN gene amplification was detected by fluorescence in situ hybridization (FISH) in 119 cases and the relationship with pathological characteristics and prognostic significance were analyzed. The study included 267 cases of children NT from patients aged from 1 day to 13 years (median 27 months). The male to female ratio was 1.43. There were 38 cases (14.2%), 43 cases (16.1%), 71 cases (26.6%), and 115 cases (43.1%) of INSS stages I, II, III and IV respectively.Favorable histology group had 157 cases (59.9%); unfavorable histology group had 110 cases (40.1%).Of the 119 NT cases with MYCN FISH performed, 18 cases (15.1%) showed amplification and the signal ratio of MYCN to CEP2 was 4.08-43.29. One hundred and one cases of non-amplified MYCN included MYCN gain in 79 cases (66.3%) and MYCN negative in 22 cases (18.5%). MYCN expression showed significant difference (P = 0.000) between ages, gender, NT type and MKI, but not INPC and clinical stage (P > 0.05).Of the 18 cases with MYCN amplification, 3 were undifferentiated, and 15 poorly differentiated; 17 had high MKI and one moderate MKI. All 18 cases were in unfavorable histology group; the overall survival rate was 3/18, with an average survival time of (17.9 ± 2.4) months.Of the 101 MYCN non-amplification cases, the overall survival rate was 68.3% (69/101), with an average survival time of (29.8 ± 1.3) months. Survival analysis showed the cases with MYCN amplification had worse prognosis (P < 0.05). NT were commonly diagnosed in early ages and easily to metastasize. Most of cases with favorable histology. The cases of MYCN amplification showed unfavorable histology, and the majority cases with high MKI; The patients with MYCN gene amplification had poor prognosis.

Identification and Pharmacological Inactivation of the MYCN Gene Network as a Therapeutic Strategy for Neuroblastic Tumor Cells

The MYC family of transcription factors consists of three well characterized members, c-MYC, L-MYC, and MYCN, deregulated in the majority of human cancers. In neuronal tumors such as neuroblastoma, MYCN is frequently activated by gene amplification, and reducing its expression by RNA interference has been shown to promote growth arrest and apoptosis of tumor cells. From a clinical perspective, RNA interference is not yet a viable option, and small molecule inhibitors of transcription factors are difficult to develop. We therefore planned to identify, at the global level, the genes interacting functionally with MYCN required to promote fitness of tumor cells facing oncogenic stress. To find genes whose inactivation is synthetically lethal to MYCN, we implemented a genome-wide approach in which we carried out a drop-out shRNA screen using a whole genome library that was delivered into isogenic neuroblastoma cell lines expressing or not expressing MYCN. After the screen, we selected for in-depth analysis four shRNAs targeting AHCY, BLM, PKMYT1, and CKS1B. These genes were chosen because they are directly regulated by MYC proteins, associated with poor prognosis of neuroblastoma patients, and inhibited by small molecule compounds. Mechanistically, we found that BLM and PKMYT1 are required to limit oncogenic stress and promote stabilization of the MYCN protein. Cocktails of small molecule inhibitors of CKS1B, AHCY, BLM, and PKMYT1 profoundly affected the growth of all neuroblastoma cell lines but selectively caused death of MYCN-amplified cells. Our findings suggest that drugging the MYCN network is a promising avenue for the treatment of high risk, neuroblastic cancers.

Significance of Cytological Findings of Neuroblastomas: Rosette Arrangement and Neuropil Structure

Objectives: Improve treatment outcomes and clarify the biological characteristics of neuroblastoma, development of an international histological classification started several years ago. Aiming at the establishment of a cytology criteria corresponding to the new histological classification, we investigated a criteria comparing lesions related to neuroblastoma on referring to the morphological indices of neuroblastoma reported in the international classification. Methods: Several tumor specimens were investigated: 37 cases of neuroblastoma (undifferentiated type: 3, poorly differentiated type: 34), 3 cases of ganglioneuroblastoma (mixed type: 2, nodular type: 1), and one case of ganglioneuroma. Stamp cytology samples were prepared from cut surfaces of the tumors and then stained to the Papanicolaou method. Results: In neuroblastoma of the undifferentiated type, tumor cells contained a small oval nucleus with a high N/C ratio, showing a bare nucleus, and the nucleolus was distinct: no rosette formation or neutrophil was observed. In the poorly differentiated type, tumor cells showed a round-oval bare nucleus were scattered: rosette arrangement was observed in the background neuroblasts containing a bare nucleus. In ganglioneuroblastoma, immature neuroblasts showed a round-oval nucleus and large ganglion-like cells possessed a distinct nucleolus similar to poorly differentiated-type epithelial adenocarcinoma. Conclusion: In neuroblastoma, neutrophils were stained light green, and a partial Homer-Wright-type rosette arrangement was observed in the background. In the poorly differentiated type, tumor cells were generally large compared to those observed in the undifferentiated type. In ganglioneuroblastoma, cytological diagnosis can be relatively easily made when differentiated mature ganglion like cells are observed. In the case of surgery, a histological diagnosis of nervous system tumors is often performed using frozen sections, however tissue is usually damaged during freezing. Thus, cytology is more advantageous for diagnosis. The diagnostic accuracy can be improved utilizing the cytological characteristics of neuroblastic tumors.

Influence of segmental chromosome abnormalities on survival in children over the age of 12 months with unresectable localised peripheral neuroblastic tumours without MYCN amplification.

Background:The prognostic impact of segmental chromosome alterations (SCAs) in children older than 1 year, diagnosed with localised unresectable neuroblastoma (NB) without MYCN amplification enrolled in the European Unresectable Neuroblastoma (EUNB) protocol is still to be clarified, while, for other group of patients, the presence of SCAs is associated with poor prognosis.Methods:To understand the role of SCAs we performed multilocus/pangenomic analysis of 98 tumour samples from patients enrolled in the EUNB protocol.Results:Age at diagnosis was categorised into two groups using 18 months as the age cutoff. Significant difference in the presence of SCAs was seen in tumours of patients between 12 and 18 months and over 18 months of age at diagnosis, respectively (P=0.04). A significant correlation (P=0.03) was observed between number of SCAs per tumour and age. Event-free (EFS) and overall survival (OS) were calculated in both age groups, according to both the presence and number of SCAs. In older patients, a poorer survival was associated with the presence of SCAs (EFS=46% vs 75%, P=0.023; OS=66.8% vs 100%, P=0.003). Moreover, OS of older patients inversely correlated with number of SCAs (P=0.002). Finally, SCAs provided additional prognostic information beyond histoprognosis, as their presence was associated with poorer OS in patients over 18 months with unfavourable International Neuroblastoma Pathology Classification (INPC) histopathology (P=0.018).Conclusions:The presence of SCAs is a negative prognostic marker that impairs outcome of patients over the age of 18 months with localised unresectable NB without MYCN amplification, especially when more than one SCA is present. Moreover, in older patients with unfavourable INPC tumour histoprognosis, the presence of SCAs significantly affects OS.

Minimally invasive surgery of neuroblastic tumors in children: Indications depend on anatomical location and image-defined risk factors.

Minimally invasive surgery (MIS) is still not a well-accepted surgical approach to remove neuroblastic tumors. We aimed to assess the indications and limits of MIS in this childhood tumor according to tumor location and image-defined risk factors (IDRFs). Between 2006 and 2012, 39 patients underwent MIS for neuroblastic tumors, using thoracoscopic (n = 20), retroperitoneoscopic (n = 1) or laparoscopic approaches (n = 18). The tumor locations were paravertebral (n = 18; thoracic n = 15, lumbar n = 3), perivascular (n = 5; abdominal n = 2; thoracic n = 3), adrenal (n = 13), pleural (n = 2) and pelvic (n = 1). Two patients were treated for relapses. According to the INRG staging system, IDRFs were absent in 20 patients and present in 19 patients. Ten patients received chemotherapy preoperatively. Mean largest diameter was 35 mm for thoracic tumors (range: 7-85 mm) and 34 mm for abdominal tumors (range: 10-75 mm). Mean follow-up was 25 months (range: 5-116 months). Resection was macroscopically incomplete (R2) for six thoracic tumors and one adrenal tumor. Conversion was necessary for three thoracic L2 tumors. Postoperative complications consisted of chylothorax in three patients with L2 paravertebral thoracic tumors, Horner's syndrome in a patient with a cervicothoracic tumor, and renal atrophy in a patient with a L2 abdominal tumor. No perioperative or postoperative complications occurred in patients with adrenal and abdominal paravertebral tumors. The overall survival rate was 98%. In carefully selected cases, MIS permits safe and efficient resection of neuroblastic tumors in children. Open surgical approach should be considered if organ or vascular control or quality of resection is jeopardized. Pediatr Blood Cancer 2015;62:257-261. © 2014 Wiley Periodicals, Inc.

Abstract 3323: PRAF2 regulates Rab5-dependent endocytic trafficking of EGFR in neuroblastoma

Abstract Receptor tyrosine kinases, such as the epidermal growth factor receptor (EGFR), mediate extracellular signaling that activates downstream cell proliferation, differentiation, and migration. Through ligand stimulation, clathrin-mediated endocytosis regulates the EGFR signal transduction pathway. Prenylated Rab acceptor 1 domain family, membrane 2 (PRAF2) is a novel 19-kDa protein with a prenylated Rab acceptor 1 (PRA1) motif and four transmembrane-spanning domains. Similar to PRAF1 and PRAF3, PRAF2 belongs to the PRAF family of vesicle transport-associated proteins. Our lab identified PRAF2 as a candidate prognostic marker of neuroblastic tumors. Using RNA interference, we showed that PRAF2 plays an essential role in neuroblastoma tumorigenesis and metastasis. In the present study, we examined the function of PRAF2 in EGFR endocytic trafficking. We generated stable PRAF2-knockdown (SK-shPRAF2) and PRAF2-overexpressing (SK-PRAF2) neuroblastoma SK-N-SH cell lines and stimulated both cell lines with EGF (10 ng/ml) for various time periods in order to determine the role of PRAF2 in EGFR endocytic trafficking. As judged by immunofluorescence microscopy, we detected larger Rab5-positive punctuate structures in PRAF2-overexpressing SK-PRAF2 cells compared to PRAF2-knockdown SK-shPRAF2 cells or control cells, after 15 min EGF stimulation. After EGF treatment, PRAF2 clearly colocalized with Rab5-positive punctae in SK-PRAF2 cells, suggesting that PRAF2 interacts with Rab5, a critical regulator for receptor endocytosis and early endosome fusion. Moreover, using a receptor internalization assay, we observed a delay in the internalization of EGFR in EGF-treated SK-shPRAF2 cells, in contrast to SK-PRAF2 and control cells. This is in support of our immunofluorescence microscopy results in which Rab5-postive punctae distribution is observed mostly along the membrane of the EGF-treated SK-shPRAF2 cells. Notably, the level of phosphorylation of Akt (Ser473) transiently elevated in EGF-treated SK-PRAF2 cells after 15 min and diminished thereafter, suggesting that overexpression of PRAF2 accelerates the EGFR internalization and degradation. Based on our findings, we propose that PRAF2 affects EGFR endocytosis by regulating early endosome trafficking and Akt activation in neuroblastoma. Citation Format: Lisette P. Yco, Andre S. Bachmann. PRAF2 regulates Rab5-dependent endocytic trafficking of EGFR in neuroblastoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3323. doi:10.1158/1538-7445.AM2014-3323

A Cre-conditional MYCN-driven neuroblastoma mouse model as an improved tool for preclinical studies.

Neuroblastoma, a childhood cancer that originates from neural crest-derived cells, is the most common deadly solid tumor of infancy. Amplification of the MYCN oncogene, which occurs in approximately 20–25% of human neuroblastomas, is the most prominent genetic marker of high-stage disease. The availability of valid preclinical in vivo models is a prerequisite to develop novel targeted therapies. We here report on the generation of transgenic mice with Cre-conditional induction of MYCN in dopamine β-hydroxylase-expressing cells, termed LSL-MYCN;Dbh-iCre. These mice develop neuroblastic tumors with an incidence of >75%, regardless of strain background. Molecular profiling of tumors revealed upregulation of the MYCN-dependent miR-17–92 cluster as well as expression of neuroblastoma marker genes, including tyrosine hydroxylase and the neural cell adhesion molecule 1. Gene set enrichment analyses demonstrated significant correlation with MYC-associated expression patterns. Array comparative genome hybridization showed that chromosomal aberrations in LSL-MYCN;Dbh-iCre tumors were syntenic to those observed in human neuroblastomas. Treatment of a cell line established from a tumor derived from a LSL-MYCN;Dbh-iCre mouse with JQ1 or MLN8237 reduced cell viability and demonstrated oncogene addiction to MYCN. Here we report establishment of the first Cre-conditional human MYCN-driven mouse model for neuroblastoma that closely recapitulates the human disease with respect to tumor localization, histology, marker expression and genomic make up. This mouse model is a valuable tool for further functional studies and to assess the effect of targeted therapies.

Case Report of a Patient With Speech Delay and Behavioral Disorders Associated with Neuroblastoma. Possible Role of SMARCA4

Pediatric cancers can occur within a context of predisposition syndromes, the exploration of which can in turn contribute to the detection of genes implicated in oncogenesis. For neuroblastoma, few associated malformative syndromes have been reported and these are only rarely characterized by genetic alterations identified to date. Here, we report the case of a girl with a delay in language acquisition and behavioral disorders associated with an adrenal neuroblastic tumor discovered at the age of 4 years.

Correction: Enteric Neural Crest Differentiation in Ganglioneuromas Implicates Hedgehog Signaling in Peripheral Neuroblastic Tumor Pathogenesis

Correction: Enteric Neural Crest Differentiation in Ganglioneuromas Implicates Hedgehog Signaling in Peripheral Neuroblastic Tumor Pathogenesis

Telomerase reverse transcriptase promoter mutations in tumors originating from the adrenal gland and extra-adrenal paraganglia.

Hotspot mutations in the promoter of the telomerase reverse transcriptase (TERT) gene have been recently reported in human cancers and proposed as a novel mechanism of telomerase activation. To explore TERT promoter mutations in tumors originating from the adrenal gland and extra-adrenal paraganglia, a set of 253 tumors (38 adrenocortical carcinomas (ACCs), 127 pheochromocytomas (PCCs), 18 extra-adrenal paragangliomas (ea PGLs), 37 head and neck PGLs (HN PGLs), and 33 peripheral neuroblastic tumors) was selected along with 16 human neuroblastoma (NBL) and two ACC cell lines to assess TERT promoter mutations by the Sanger sequencing method. All mutations detected were confirmed by a SNaPshot assay. Additionally, 36 gastrointestinal stromal tumors (GISTs) were added to explore an association between TERT promoter mutations and SDH deficiency. TERT promoter mutations were found in seven out of 289 tumors and in three out of 18 human cell lines; four C228T mutations in 38 ACCs (10.5%), two C228T mutations in 18 ea PGLs (11.1%), one C250T mutation in 36 GISTs (2.8%), and three C228T mutations in 16 human NBL cell lines (18.75%). No mutation was detected in PCCs, HN PGLs, neuroblastic tumors as well as ACC cell lines. TERT promoter mutations preferentially occurred in a SDH-deficient setting (P=0.01) being present in three out of 47 (6.4%) SDH-deficient tumors vs zero out of 171 (0%) SDH-intact tumors. We conclude that TERT promoter mutations occur in ACCs and ea PGLs. In addition, preliminary evidence indicates a potential association with the acquisition of TERT promoter mutations in SDH-deficient tumors.

Ventricular ganglioneuroblastoma in an adult and successful treatment with radiotherapy.

Ganglioneuroblastomas (GNBs) are rare embryonic neoplasms in the spectrum of neuroblastic tumours and 80% of cases occur in the first decade. In adults, GNBs are usually located in the retroperitoneum, mediastinum and neck but intracranial GNBs are extremely rare. A 34-year-old male applied to the Department of Neurology outpatient clinic with a two month history of headache and numbness in his legs. Detailed examinations and cranial CT were performed and showed a mass with a 5 cm diameter running from the third to the fourth ventricle. Referral to a neurosurgeon was performed for partial removal of the tumour, as the histological and immunohistochemical studies defined the diagnosis of GNB. Three months later, when the patient experienced dizziness, an MRI was performed, which showed a 4×3 cm ventricular mass, with hypointense characterisation in T1-weighted and hyperintense characteristics in T2-weighted and flair sequences. Afterwards, fractioned radiotherapy (54 Gy/30 fx) was chosen as the appropriate therapy. In the follow-up period, MRI was performed 3 months and 1 year after treatment, and revealed shrinkage of the tumour by at least 50%. Meanwhile the patient's post-irradiation course was favourable. Data following the use of radiotherapy as treatment for intracranial GNB showing favourable results has been reported.