Characterization of EWSR1 Break Apart FISH in Non-EWSR1 Related Samples: Avoiding the False Positive Pitfall
Navid Sadri, Paul J Zhang. University of Pennsylvania, Philadelphia, PA
Background: Hybridizing two colored DNA probes flanking break points of a specific gene can visualize a signal separation (SS) in tumor cells containing that translocation. This break-apart FISH assay is widely used on routine paraffin section for diagnosis. Currently there lacks standardization in interpreting the presence and incidence of SS. Although false positive SS are recognized to exist their characterization and incidence has not been sufficiently characterized.
Design: We performed EWSR1 break-apart probes (Vysis) on sarcomas with a non-EWSR1 translocations (synovisal sarcomas [SYS; n=3] and alveolar rhabdomyosarcomas [AR, n=4]), 3 breast carcinomas (BRCa) and 2 normal tonsils. Tumors known to have EWSR1 translocation (EWSR1+; 1 DSRCT, 3 EWS/PNET, 1 extraskeletal myxoid chondrosarcoma) were evaluated as a control. Each tumor cell was categorized to have: at least two intact alleles (IA); at least one IA with one SS; or unpaired signal(s) (UPS). Cells with no signal, only one unpaired signal, or only one intact allele were not counted. The average EWSR1 alleles per cell were determined. The signal width (SW) was calculated for SS.
Results: See table below.
|Diagnosis||IA||UPS||SS||1 SW||>2 SW||>3 SW||>4 SW|
|STS/AR (n =7)||49±18||24±14||21±5||12±3||9±4||4±3||0.7±0.6|