Diagnostic value of the molecular genetic detection of the t(11;22) translocation in Ewing's tumours.

One consistent feature of the Ewing's tumour family is the presence of a balanced translocation involving band q12 and band q24 of chromosome 22 and chromosome 11. Recent cloning of the chromosome breakpoint regions of t(11;22)(q24;q12) Ewing's sarcoma translocation has revealed that the breakpoints were localized within the Ewing's sarcoma gene (EWS gene) on chromosome 22 and the Fli-1 gene on chromosome 11. Molecular genetic techniques can thus be applied to the detection of the t(11;22) translocation in Ewing's tumours. By reverse transcription and polymerase chain reaction technique (RT-PCR) 11 Ewing's tumour derived cell lines, 12 primary Ewing's tumours, and 11 tumours after treatment were analysed for the occurence of the t(11;22) translocation. Furthermore, blood and bone marrow samples from 5 patients were available for RT-PCR. In 78% of the cell lines and 91% of the primary Ewing's tumours the t(11;22) translocation was detectable by RT-PCR. In bone marrow samples from a Ewing's sarcoma patient presenting in relapse tumour cells were detected by molecular genetic analysis. Our results indicate that molecular genetic detection of the t(11;22) translocation is valuable in the differential diagnosis of small round cell tumours and will provide important information for the staging and prognosis of Ewing's tumour.

Rapid detection of loss of heterozygosity of chromosome 17p by polymerase chain reaction-based variable number of tandem repeat analysis and detection of single-strand conformation polymorphism of intragenic p53 polymorphisms.

Intragenic restriction site polymorphisms in amino acid residue 72 in exon 4 and a Mspl polymorphism in intron 6 of the p53 tumour suppressor gene can both serve as polymorphic markers. Probe YNZ22 (D17S5) is a highly polymorphic, variable number of tandem repeat (VNTR) marker which maps to chromosome 17p13.1 where the p53 gene is located. Locus specific amplification by polymerase chain reaction (PCR) technique and subsequent non-isotopic single-strand conformation polymorphism analysis of the PCR fragments was used for the detection of loss heterozygosity (LOH) of 17p including the p53 gene locus. In combination with a PCR-based method for the analysis of the VNTR locus D17S5 using unique sequences flanking the polymorphic region of YNZ22 we investigated tumour DNA and corresponding constitutional DNA from 69 patients, including 39 patients with gastric cancer, 21 patients with osteosarcomas and 9 patients with Ewing's sarcomas. Using all three methods, 49/69 (71%) patients were informative for LOH, which revealed allelic loss in 5/39 (12.8%) gastric cancers, 1/9 (11.1%) Ewing's sarcoma, and 4/20 (20%) osteosarcomas.

[Detection of EWS-/FLI-1 gene fusion transcripts by RT-PCR as a tool in the diagnosis of tumors of the Ewing sarcoma group]. / Nachweis von EWS/FLI-1-Genfusionstranskripten mit RT-PCR als diagnostische Hilfe bei Tumoren der Ewing-Sarkom-Gruppe.

Recent cloning of the chromosome breakpoint regions of the reciprocal chromosomal t(11;22) (q24;q12) has revealed that the breakpoints were localized within the EWS gene (Ewings sarcoma gene) on chromosome 22 and the FLI-1 gene on chromosome 11. Thus, molecular genetic techniques were applicable for the detection of this genetic aberration, which occurs as a consistent feature of the Ewings tumor family. By reverse transcription and polymerase chain reaction technique (RT-PCR) in 78% of Ewings sarcoma derived cell lines, and in 91% of primary Ewings tumor tissue t(11;22) specific EWS/FLI-1 fusion transcripts were detected. Furthermore, in bone marrow samples from an Ewings sarcoma patient contaminating tumor cells could be shown by RT-PCR. Our results indicate that molecular genetic detection of the t(11;22) translocation opens a new modality for the differential diagnosis and the staging of Ewings tumor patients.

[Molecular genetic detection of t(11;22)(q24;12) translocation in Ewing sarcoma and malignant peripheral neuroectodermal tumors]. / Molekulargenetischer Nachweis der t(11;22)(q24;q12)-Translokation in Ewing-Sarkomen und malignen peripheren neuroektodermalen Tumoren (MPNT).

Ewing's sarcomas and malignant peripheral neuroectodermal tumors (MPNTs) show very little evidence of differentiation and lack characteristic morphological features at the light-microscopic level. These malignancies have always presented a significant differential diagnostic challenge to the pathologist. Electron microscopy, immunohistochemical staining for neural antigens such as neuron-specific enolase (NSE), Leu 7, synaptophysin and, more recently, the detection of Mic-2 gene expression have been included in the routine histopathological diagnostic procedure. However, the expression of these antigens is not restricted to this entity. Thus, further modalities are required to prove diagnostic reliability. One consistent feature of the Ewing's sarcoma family is the presence of the reciprocal chromosomal t(11;22)(q24;q12) translocation. Recent cloning of the t(11;22) break point has led to the identification of the genes involved in this translocation. This provides the possibility of molecular genetic detection of the t(11;22) translocation in Ewing's sarcomas and MPNTs. We have established a method using reverse transcription and the polymerase chain reaction (RT-PCR) for the detection of the specific gene fusion transcript caused by the 11;22 translocation. The validity of our approach was proved by analyzing Ewing's tumor cell lines and tissue material obtained from primary biopsies and tumor resections. Molecular genetic detection of the 11;22 translocation by RT-PCR analysis should perhaps be included in the diagnostic work-up of suspected Ewing's sarcoma and MPNT.