title: Integrated multimodal genetic testing of Ewing sarcoma--a single-institution experience.
abstract: Ewing sarcoma (ES) is an aggressive malignant small round cell tumor that arises in bone or soft tissue of adolescents and young adults. A characteristic molecular finding in ES is EWSR1 gene fusion with ETS (erythroblast transformation-specific) family genes including FLI1 (~90%) and ERG (>5%). Here we report our experience using integrated clinicopathologic, cytogenetic, fluorescence in situ hybridization (FISH), and reverse transcriptase polymerase chain reaction (RT-PCR) analyses of 32 pediatric patients with ES diagnosed in a single institution between 2005 and 2011. Diagnostic EWSR1 rearrangements were detected in 30 (93.8%) of 32 patients. Cytogenetics detected t(11;22) (n = 14) and t(21;22) (n = 1) in 15 (46.9%) patients. FISH detected EWSR1 rearrangements in 27 (96.4%) of 28 patients tested. RT-PCR was positive in 27 (84.4%) of 32 patients, including 24 EWSR1-FLI1 and 3 EWSR1-ERG. RT-PCR defined breakpoints and fusion partners in 7 cases with EWSR1 rearrangements detected by FISH. Sanger sequencing further delineated breakpoints in 21 (77.8%) of 27 RT-PCR positive cases. In summary, conventional cytogenetic analysis provided a global view but had a lower detection rate and longer turnaround time than other methods. FISH is a rapid method and theoretically can detect all EWSR1 rearrangements, but it cannot identify all partners and is not completely specific for ES. RT-PCR and sequencing are more sensitive and useful in identifying fusion partners and refining breakpoints; however, these methods can be compromised by poor RNA preservation and primer design. In conclusion, an integrated approach that uses all methods capable of detecting EWSR1 rearrangements has value in the workup of suspected cases of ES.
authors.affiliation: Department of Pathology and Immunology, Baylor College of Medicine, Texas Children's Hospital, Houston, TX 77030, USA.
authors.forename: Mikako
authors.initials: M
authors.lastname: Warren
authors.forename: Michael
authors.initials: M
authors.lastname: Weindel
authors.forename: Jo
authors.initials: J
authors.lastname: Ringrose
authors.forename: Clint
authors.initials: C
authors.lastname: Venable
authors.forename: Adriana
authors.initials: A
authors.lastname: Reyes
authors.forename: Keita
authors.initials: K
authors.lastname: Terashima
authors.forename: Pulivarthi
authors.initials: P
authors.lastname: Rao
authors.forename: Murali
authors.initials: M
authors.lastname: Chintagumpala
authors.forename: M John
authors.initials: MJ
authors.lastname: Hicks
authors.forename: Dolores
authors.initials: D
authors.lastname: Lopez-Terrada
authors.forename: Xin-Yan
authors.initials: XY
authors.lastname: Lu
mesh_terms: D000293:Adolescent; D001859:Bone Neoplasms / Q000235:genetics* / Q000473:pathology; D002148:Calmodulin-Binding Proteins / Q000235:genetics; D002648:Child; D002675:Child, Preschool; D002880:Chromosomes, Human, Pair 11; D002892:Chromosomes, Human, Pair 22; D004273:DNA, Neoplasm / Q000235:genetics; D005260:Female; D015321:Gene Rearrangement; D005820:Genetic Testing / Q000379:methods*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D007223:Infant; D008297:Male; D015514:Oncogene Proteins, Fusion / Q000235:genetics*; D034802:RNA-Binding Protein EWS; D016601:RNA-Binding Proteins / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012512:Sarcoma, Ewing / Q000235:genetics* / Q000556:secondary; D012983:Soft Tissue Neoplasms / Q000235:genetics* / Q000473:pathology; D015534:Trans-Activators / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics*; D000071230:Transcriptional Regulator ERG; D014178:Translocation, Genetic; D055815:Young Adult
chemical_list: D002148:Calmodulin-Binding Proteins; D004273:DNA, Neoplasm; C072312:ERG protein, human; C450460:EWS-ERG fusion protein, human; C500986:EWSR1 protein, human; D015514:Oncogene Proteins, Fusion; D034802:RNA-Binding Protein EWS; D016601:RNA-Binding Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D000071230:Transcriptional Regulator ERG
keywords: Chromosome analysis; Ewing sarcoma; FISH; Integrated testing; RT-PCR; Sequencing