A practical approach to the clinical diagnosis of Ewing's sarcoma/primitive neuroectodermal tumour and other small round cell tumours sharing EWS rearrangement using new fluorescence in situ hybridisation probes for EWSR1 on formalin fixed, paraffin wax embedded tissue

Abstract

Background: Over 90% of Ewing's sarcoma/primitive neuroectodermal tumour (ES/PNET) cases have the t(11;22) chromosomal rearrangement, which is also found in other small round cell tumours, including desmoplastic small round cell tumour (DSRCT) and clear cell sarcoma (CCS). Although this rearrangement can be analysed by fluorescence in situ hybridisation (FISH) using routinely formalin fixed, paraffin wax embedded (FFPE) tissues when fresh or frozen tissues are not available, a sensitive and convenient detection method is needed for routine clinical diagnosis.

Aims: To investigate the usefulness of newly developed probes for detecting EWS rearrangement resulting from chromosomal translocations using FISH and FFPE tissue in the clinical diagnosis of ES/PNET, DSRCT, and CCS.

Methods: Sixteen ES/PNETs, six DSRCTs, and six CCSs were studied. Three poorly differentiated synovial sarcomas, three alveolar rhabdomyosarcomas, and three neuroblastomas served as negative controls. Interphase FISH analysis was performed on FFPE tissue sections with a commercially available EWSR1 (22q12) dual colour, breakapart rearrangement probe.

Results: One fused signal and one split signal of orange and green, demonstrating rearrangement of the EWS gene, was detected in 14 of 16 ES/PNETs, all six DRSCTs, and five of six CCSs, but not in the negative controls.

Conclusions: Interphase FISH using this newly developed probe is sensitive and specific for detecting the EWS gene on FFPE tissues and is of value in the routine clinical diagnosis of ES/PNET, DSRCT, and CCS.

Figure 1

(A) Low power view of a Ewing’s sarcoma and primitive neuroectodermal tumour of the pancreas, composed of a monotonous proliferation of small round cells, with an undifferentiated appearance (haematoxylin and eosin; original magnification, ×100). (B) Low power view of an intra-abdominal desmoplastic small round cell tumour. Compact nests consisting of small cells separated by abundant desmoplastic stroma, with an undifferentiated appearance, can be seen (haematoxylin and eosin; original magnification, ×100). (C) A clear cell sarcoma showing nested epithelioid cells with clear cytoplasm and prominent nucleoli. The nests of tumour cells were separated by fibrous bands (haematoxylin and eosin; original magnification, ×100).

Figure 2

Results of hybridisation of the EWSR1 dual colour breakapart probes. Tumour cells of (A) a Ewing’s sarcoma and primitive neuroectodermal tumour, (B) a desmoplastic small round cell tumour, and (C) a clear cell sarcoma show one fusion, one orange, and one green signal pattern, indicative of a rearrangement of one copy of the EWSR1 region. (D) Tumour cells of an alveolar rhabdomyosarcoma show two fusion signal patterns, reflecting the two intact copies of EWSR1.