Molecular diagnosis of Ewing sarcoma/primitive neuroectodermal tumor in routinely processed tissue: a comparison of two FISH strategies and RT-PCR in malignant round cell tumors.

Ewing sarcoma/primitive neuroectodermal tumor (EWS/PNET) is a diagnostically challenging malignant round cell tumor with signature translocations involving the EWS gene. These translocations are detectable with both reverse transcriptase-polymerase chain reaction (RT-PCR) and fluorescence in situ hybridization (FISH) in formalin-fixed paraffin-embedded tissue. However, RT-PCR is less sensitive in formalin-fixed paraffin-embedded than frozen tissue. Similarly, commercial FISH probes have recently become available, but have yet to be rigorously tested in the clinical setting. Therefore, we have compared RT-PCR with FISH using 'home brew' fusion probes for Ewing sarcoma (EWS)-FLI1 and a commercial EWS break apart probe set in 67 archival round cell tumors, including 27 EWS/PNETs. Sensitivities and specificities for both FISH assays were 91 and 100%, respectively, whereas RT-PCR had a sensitivity of 54% and a specificity of 85%. The break apart strategy was easier to interpret than probe fusion approach. We conclude that FISH is a more sensitive and reliable ancillary technique than RT-PCR for the diagnosis of EWS/PNET in formalin-fixed paraffin-embedded tissue, although the latter provides additional information regarding fusion transcript subtype and prognosis. The commercial break apart probe set is both readily available and easy to interpret, making it particularly attractive. Nonetheless, complex round cell tumors often benefit from molecular testing with multiple methods.

Cytogenetic and molecular cytogenetic evidence of recurrent 8q24.1 loss in osteochondroma.

Osteochondroma most frequently arises sporadically and as a solitary lesion, but may also arise as multiple lesions characterizing the autosomal dominant disorder hereditary multiple exostoses (HME) and the contiguous gene-deletion syndrome, Langer-Giedion syndrome (LGS). Various germline mutations of two putative tumor suppressor genes, EXT1 localized to 8q24.1 and EXT2 localized to 11p11 approximately p12, have been demonstrated in HME families. Constitutional chromosomal deletions or structural rearrangements of 8q24.1 are seen in LGS. Cytogenetic reports of sporadic and hereditary osteochondromas are few, but have revealed loss or structural rearrangements of 8q24.1 in a small number of tumors. In the current study, karyotypic evaluation of 37 osteochondroma specimens (both sporadic and hereditary lesions) revealed chromosomal anomalies of 8q24.1 in 10 specimens (27%). In an effort to determine the presence and frequency of submicroscopic deletions, molecular cytogenetic studies were performed on this same set of tumors utilizing a chromosome 8 specific centromeric probe and an 8q24.1 cosmid probe (locus D8S51, within the minimal LGS deletion region). Loss of the 8q24.1 locus was detected by fluorescence in situ hybridization in 27 of 34 (79%) osteochondroma specimens analyzed including all 10 specimens exhibiting chromosome 8 abnormalities cytogenetically. These findings indicate that a significant subset of osteochondromas harbor genetic aberrations at the EXT1 locus and suggest that loss or mutation of EXT1 plays an important role in the pathogenesis of sporadic as well as hereditary osteochondromas.