Purpose:Tumors of the Ewing family are characterized by chromosomal translocations that yield chimeric transcription factors, such as EWS/FLI1, which regulate the expression of specific genes that contribute to the malignant phenotype. In the present study, we show that cholecystokinin (CCK) is a new target of the EWS/FLI1oncoprotein and assess its functional role in Ewing tumor pathogenesis. Experimental Design: Relevant EWS/FLI1 targets were identified using a combination of cell systems with inducible EWS/FLI1expression, Ewing tumors and cell lines, microarrays, and RNA interference with doxycycline-inducible small hairpin RNA (shRNA) vectors. A doxycyclineinducible CCK-shRNA vector was stably transfected in A673 and SK-PN-DW Ewing cell lines to assess the role of CCK in cell proliferation and tumor growth. Results: Microarray analysis revealed that CCK was up-regulated by EWS/FLI1 in HeLa cells. CCK was overexpressed in Ewing tumors as compared with other pediatric malignancies such as rhabdomyosarcoma and neuroblastoma, with levels close to those detected in normal tissues expressing the highest levels of CCK. Furthermore, EWS/FLI1 knockdown in A673 and SK-PN-DW Ewing cells using two different doxycycline-inducible EWS/FLI1-specific shRNA vectors down-regulated CCK mRNA expression and diminished the levels of secreted CCK, showing that CCK is a EWS/FLI1 specific target gene in Ewing cells. A doxycycline-inducible CCK-specific shRNA vector successfully down-regulated CCK expression, reduced the levels of secreted CCK in Ewing cell lines, and inhibited cell growth and proliferation in vitro and in vivo. Finally, we show that Ewing cell lines and tumors express CCK receptors and that the growth inhibition produced by CCK silencing can be rescued by culturing the cells with medium containing CCK. Conclusions: Our data support the hypothesis that CCK acts as an autocrine growth factor stimulating the proliferation of Ewing cells and suggest that therapies targeting CCK could be promising in the treatment of Ewing tumors.Ewing family of tumors is a group of highly malignant tumors arising mainly in bone that most often affects children and young adults in the first two decades of life. Despite the use of multimodal therapy (chemotherapy, radiation therapy, and surgery), the long-term disease-free survival rate of Ewing tumor patients is still disappointingly low, particularly in the high-risk groups with metastasis at diagnosis or unfavorable localization (reviewed in ref. 1). The lack of new effective drugs in the treatment of Ewing tumors, together with the side effects that high-dose regimens have on young patients who have a long life expectancy, supports the need for innovative therapeutic strategies that should mainly include targeted therapies against molecules critical for the pathogenesis and progression of these tumors.The molecular hallmark of Ewing family of tumors is the presence of balanced chromosomal translocations leading to the formation of chimeric transcription factors. These aberrant...

Pediatric large B-cell lymphomas (LBCLs) share morphological and phenotypic features with adult types but have better prognosis. The higher frequency of some subtypes such as LBCL with IRF4 rearrangement (LBCL-IRF4) in children suggests that some age-related biological differences may exist. To characterize the genetic and molecular heterogeneity of these tumors, we studied 31 diffuse LBCLs (DLBCLs), not otherwise specified (NOS); 20 LBCL-IRF4 cases; and 12 cases of high-grade B-cell lymphoma (HGBCL), NOS in patients ≤25 years using an integrated approach, including targeted gene sequencing, copy-number arrays, and gene expression profiling. Each subgroup displayed different molecular profiles. LBCL-IRF4 had frequent mutations in IRF4 and NF-κB pathway genes (CARD11, CD79B, and MYD88), losses of 17p13 and gains of chromosome 7, 11q12.3-q25, whereas DLBCL, NOS was predominantly of germinal center B-cell (GCB) subtype and carried gene mutations similar to the adult counterpart (eg, SOCS1 and KMT2D), gains of 2p16/REL, and losses of 19p13/CD70. A subset of HGBCL, NOS displayed recurrent alterations of Burkitt lymphoma–related genes such as MYC, ID3, and DDX3X and homozygous deletions of 9p21/CDKN2A, whereas other cases were genetically closer to GCB DLBCL. Factors related to unfavorable outcome were age >18 years; activated B-cell (ABC) DLBCL profile, HGBCL, NOS, high genetic complexity, 1q21-q44 gains, 2p16/REL gains/amplifications, 19p13/CD70 homozygous deletions, and TP53 and MYC mutations. In conclusion, these findings further unravel the molecular heterogeneity of pediatric and young adult LBCL, improve the classification of this group of tumors, and provide new parameters for risk stratification.

Ewing sarcomas (ES) are pediatric bone tumors that arise from a driver translocation, most frequently EWS/FLI1. Current ES treatment involves DNA damaging agents, yet the basis for the sensitivity to these therapies remains unknown. Oncogene-induced replication stress (RS) is a known source of endogenous DNA damage in cancer, which is suppressed by ATR and CHK1 kinases. We here show that ES suffer from high endogenous levels of RS, rendering them particularly dependent on the ATR pathway. Accordingly, two independent ATR inhibitors show in vitro toxicity in ES cell lines as well as in vivo efficacy in ES xenografts as single agents. Expression of EWS/FLI1 or EWS/ERG oncogenic translocations sensitizes non-ES cells to ATR inhibitors. Our data shed light onto the sensitivity of ES to genotoxic agents, and identify ATR inhibitors as a potential therapy for Ewing Sarcomas.

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Dermatomyofibroma represents a rare benign fibroblastic/ myofibroblastic cutaneous tumor that mostly occurs in young adult women. It has been seldom reported in pediatric patients. In this analysis, the clinical, histopathological and immunohistochemical findings of 12 dermatomyofibromas occurring in patients up to 16 years of age are compared with those reported in adults. Six patients were male and six were female. Nine lesions were located on the neck, two on the back and one involved the chest. The usual presentation was as an asymptomatic plaque composed of bland spindled cells arranged in dermal fascicles that were oriented parallel to the epidermis. Immunohistochemically, the lesional cells expressed calponin in 11 cases, smooth muscle actin in six and muscle-specific actin in three. In contrast to prior reports from adults, dermatomyofibromas in pediatric patients do not show a female predilection. In addition, they are mostly located on the neck (56%), while in adults the most frequent location is the shoulder (35%). Dermatomyofibromas seem to stabilize after an initial period of enlargement. Punch biopsy and clinical follow up could be an alternative approach to the surgical excision in some cases of dermatomyofibroma, particularly in instances in which surgery might inflict cosmetic defects.

A number of dermatoses have been reported to appear in close temporal or spatial relationship to the intramuscular injection of vaccines. We describe two young children who had morphea consistent with the deep morphea type (morphea profunda) that appeared at the site of a previous vaccination. Trauma has been implicated as an important trigger of morphea, both in children and in adults. Furthermore, vaccines might induce an immune response, which could lead to morphea in predisposed individuals.

Pathologists should reserve the term ASC for tumors exhibiting the above mentioned appearance. In such circumstances, a metastatic origin must always be excluded.