Multiple splice variants of EWSR1-ETS fusion transcripts co-existing in the Ewing sarcoma family of tumors

Abstract

The Ewing sarcoma family of tumors (EFT) is characterized by fusions of the EWSR1 gene on chromosome 22q12 with either one of the genes encoding members of the ETS family of transcription factors, in the majority of cases FLI1 or ERG. Many alternative EWSR1-ETS gene fusions have been encountered, due to variations in the locations of the EWSR1 and ETS genomic breakpoints. The resulting heterogeneity in EWSR1-ETS fusion transcripts may further be increased by the occurrence of multiple splice variants within the same tumor. Here we present a retrospective study designed to detect all of the EWSR1-FLI1 and EWSR1-ERG fusion transcripts in a series of 23 fresh frozen EFT tissues. RT-PCR and nested fluorescent multiplex PCR were used to amplify EWSR1-FLI1 and EWSR1-ERG transcripts from EFT tissues. Fusion transcripts were identified by laser-induced fluorescent capillary electrophoresis and confirmed by sequence analysis. Nine different EWSR1-FLI1 fusion transcripts and one EWSR1-ERG fusion transcript were identified in 21 out of 23 fresh frozen EFT tissue samples. In five cases multiple fusion transcripts were found to coexist in the same tumor sample. We additionally reviewed previous reports on twelve cases with multiple EWSR1-ETS fusion transcripts. Alternative splicing may frequently affect the process of EFT-associated fusion gene transcription and, as such, may significantly contribute to the pathogenic role of EFT-associated chromosome translocations. In a considerable number of cases this may result in multiple splice variants within the same tumor.