Sequencing Overview of Ewing Sarcoma: A Journey across Genomic, Epigenomic and Transcriptomic Landscapes

Abstract

Ewing sarcoma is an aggressive neoplasm occurring predominantly in adolescent Caucasians. At the genome level, a pathognomonic EWSR1-ETS translocation is present. The resulting fusion protein acts as a molecular driver in the tumor development and interferes, amongst others, with endogenous transcription and splicing. The Ewing sarcoma cell shows a poorly differentiated, stem-cell like phenotype. Consequently, the cellular origin of Ewing sarcoma is still a hot discussed topic. To further characterize Ewing sarcoma and to further elucidate the role of EWSR1-ETS fusion protein multiple genome, epigenome and transcriptome level studies were performed. In this review, the data from these studies were combined into a comprehensive overview. Presently, classical morphological predictive markers are used in the clinic and the therapy is dominantly based on systemic chemotherapy in combination with surgical interventions. Using sequencing, novel predictive markers and candidates for immuno- and targeted therapy were identified which were summarized in this review.

Keywords: Ewing sarcoma; bone neoplasm; bone tumor; epigenetics; immunotherapy; micro-environment; next generation sequencing; soft tissue tumor; splicing; targeted therapy.

Figure 1

Mutation overview of reported STAG2 and TP53 in Ewing sarcoma. Overview of published mutations on STAG2 and TP53 from next generation sequencing data divided in five mutation subtypes based on data collected from 472 tumors and 54 cell lines. (A) Overview of the STAG2 mutations (B) Overview of the TP53 mutations. Amino acid sequence of the proteins is presented with different protein domains annotated in boxes and every sphere represents a reported mutation.

Figure 2

EWSR1-ETS mediated epigenetic activation and repression of gene expression. Possible mechanisms of how EWSR1-ETS acts as a transcription activator or repressor with different chromatin remodelers and is associated with different epigenetic histone modifications. (A) EWSR1-ETS activation complex binds to GGAA microsatellites. The complex attracts LSD1 in a yet unidentified activation complex and p300, which is needed for efficient transcription. The activation complex may bind to H3K4me3 and H3K27ac histone marks which, in turn, may lead to upregulation of the epigenetic modifiers SIRT1 and EZH2; (B) EWSR1-ETS repression complex binds to single GGAA elements and scavenges for p300, but, as it is insufficient to create an activating complex, it may recruit NuRD repression complex which may lead to further repressed expression. In addition, these repression sites are marked with H3K9me3 histone mark.

Figure 3

The influence of EWSR1-ETS fusion protein at the transcriptome level. EWSR1-ETS fusion protein acts as an aberrant transcription factor that influences the regulation of mRNA, lncRNA and miRNA expression levels. In addition, by binding to RHA additional transcripts can be bound and this might interfere with the stability of these transcripts. Alterations in epigenetic activity lead to up- and downregulation of a number of transcription factors and thereby interfere indirectly with gene expression. Furthermore, EWSR1-ETS fusion protein binds to the spliceosome and thereby altering splicing processes. By acting on these mediators, multiple cellular pathways are affected. The summarizing gene ontology clusters of the upregulated (green) cellular pathways are cell cycle, membrane proteins, IGF signaling and transcription and the downregulated is extracellular signaling (red). The main processes influenced by these gene ontology clusters are an increase in proliferation, pluripotency, migration and angiogenesis.