Background: Chromosomal translocation are frequently observed in leukemias and sarcomas; these translocations break specific genes in the involved chromosomes and create novel chimeric genes that encode a fusion protein. Advances in these techniques have increased knowledge of the genes involved in tumoral development; molecular techniques have enabled more precise diagnosis as well as identification of new prognostic factors.

Aims: To explore the use of Reverse-Transcriptase Poly-merase Chain Reaction (RT-PCR) assay for detecting fusion transcripts in a series of Soft Tissue Sarcomas (STS) and compare the results with histopathologic diagnosis.

Material and methods: We studied 80 biopsies performed at Orthopaedic Oncology and Reconstructive Surgery Department, CTO-CFR-M.Adelaide Hospital Turin Italy, with clinical suspect of STS. Histological diagnosis was obtained contemporary to evaluation of chimeric transcripts detected by RT-PCR. cDNA were PCR amplified using primer specific for each sarcoma. Paraffin-embedded tissue samples were not used because the poor quality of the extracted RNA may give wrong positive results. Results Histology confirmed 21 Ewing Sarcoma (ES), 14 Synovial sarcoma (SS), 7 Mixoid liposarcoma (M-LPS), 4 Extraskeletal Myxoid Chondrosarcoma (E-MCDS), 4 Dermatofibrosarcoma protuberans (DFSP), 10 Rhabdo-myosarcoma, 10 Leiomyosarcoma. Of the 21 tumors diagnosed as ES, 21 (100%) expressed EWS-FLI1 chimeric transcripts. All 14 SS were positive for SYT-SSX fusion transcripts. Of the 7 cases with diagnosis of M-LPS, six were positive for EWS-CHOP transcripts; of the four cases of E-MCS 3 were positive for EWS-CHN fusion transcripts. All 4 DFSP were positive for COL1A1-PDGFB transcripts. Expression of Myo-D1, tested in ten cases of Rhabdomyosarcoma, was positive while in ten cases of Leiomyosarcoma no expression of Myo-D1 was detected by RT-PCR Ten cases were non sarcoma and negative for molecular biology.

Conclusion These results demonstrate a strong concordance between the standard histopathological diagnosis and molecular results. These techniques could be a useful method to increase the quality of histologic diagnosis in difficult cases.

Backgroud: Leiomyosarcoma (LMS) is characterized by malignant smooth muscle cells. LMS are principally tumours of adult life, children rarely develop these tumours. The cause of LMS is not understood, however the genetic alterations are thought to be important, if not inciting, in the formation and progression of sarcoma. The silencing of tumour suppressor genes by promoter hypermethylation is a common feature among many types of malignancies; it has been proposed that DNA methylation provides an alternative pathway to gene mutation.

Aims: To evaluate the promoter methylation in LMS: two tumour-related genes (MGMT and RASSF1A) were studied.

Materials and methods: 14 LMS specimens were obtained from patients treated at Orthopaedic Oncology and Reconstructive Surgery Department, CTO-CFR-M.Adelaide Hospital, Turin. Genomic DNA was extracted from paraffin sections and frozen samples according to standard protocols. Transformation of the methylations patterns in the CpG island of RASSF1A and MGMT of DNA were determined by chemical modification. To analyzed the role of aberrant DNA in LMS, methylation status by SP-PCR was evaluated. PCR products were amplified by unmethylated (U) and methylated (M)-specific primers.

Results: The RASSF1A promoter was methylated in 4 (29%) LMS, MGMT promoter was methylated in 2 (15 %); 1 of these patients with methylation had both RASSF1A and MGMT. Eight LMS samples did not show any methylation.

Conclusions: Our data indicate that inactivation of RASSF1A is a common event in LMS. Aberrant methylation of the RASSF1A promoter region is one of the most frequent epigenetic inactivation events detected in human cancer and leads to silencing of RASSF1A; moreover inactivation of RASSF1A was usually associated to poor prognosis. According to recent reports, that demonstrated that promoter hypermethylation of the MGMT gene is not a frequent event in LMS, the present study detected MGMT in 2 (15%) tumour samples.