A characterization of NR0B1 in the pathogenesis of Ewing's sarcoma

Ewing's sarcoma is a devastating pediatric malignancy. Current treatments are highly toxic, invasive, and not sufficiently targeted or efficacious. It is hoped that knowledge of the molecular mechanisms required for disease development and maintenance will assist in the generation of more directed and successful therapeutic options for Ewing's sarcoma patients.;Using a loss-of-function approach targeting EWS/FLI in multiple patient-derived Ewing's sarcoma cells coupled with microarray analysis, we identified genes dysregulated by the fusion protein. Notably, NR0B1 was the most consistently up-regulated target of EWS/FLI. NR0B1 is a unique orphan member of the nuclear hormone receptor superfamily that functions throughout the steroidogenic axis. NR0B1 is expressed in Ewing's sarcoma patient tumor samples and cell lines, and is required for the transformed phenotype. Therefore, NR0B1 is a critical EWS/FLI target that has relevance to the bona fide disease.;NR0B1 was identified as being directly regulated by EWS/FLI via a GGAA microsatellite in the NR0B1 promoter. This element was enriched in EWS/FLI up-regulated genes, but not down-regulated targets. These data suggest the EWS/FLI down-regulated gene signature may be modulated by EWS/FLI effector proteins. NR0B1 is a strong candidate to mediate this repression downstream of EWS/FLI.;Microarray and whole genome localization studies were performed and NR0B1 was found to function both as a co-activator and a co-repressor. Furthermore, NR0B1 and EWS/FLI were found to co-occupy a subset of genomic loci. A direct interaction that appears to have transcriptional and transforming consequences was then identified between the two proteins.;A breakthrough occurred with the observation that the majority of Ewing's tumors harbored a reciprocal translocation, t(11;22)(q24;q12). This chromosomal abnormality results in the synthesis of a fusion protein, EWS/FLI, which functions as an oncogenic aberrant transcription factor. Identifying and understanding the genes influenced by EWS/FLI may provide molecular insight into the transformed phenotype of Ewing's sarcoma.;Taken together this work has identified NR0B1 as a critical direct target of EWS/FLI, whose protein product contributes to transformation by interacting with EWS/FLI and functioning as a co-factor for transcription. NR0B1 may then serve as a novel therapeutic target for the treatment of patients with Ewing's sarcoma.