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.

Aims: Angiogenesis is a multistep phenomenon, critical for tumor growth and prognosis in many solid neoplasms. Microvessels density (MVD) is a method of assessing angiogenesis and adversely affects DFS and OS in breast, lung and colorectal cancer. Few data are available in STS in which stage and grading are until now fundamental.

Methods: Our perspective study determined the level of MVD in a series of STS and correlated these results with DFS and OS, comparing its prognostic value with grading and stage. MVD was determined with CD 31 immunostains in formalin fixed, paraffin embedded tissues. Intratumoral MVD was assessed by light microscopic analysis. Hot spots defined the positive areas. The study included 45 patients, 35 with localized and 10 with metastatic disease at diagnosis. All tumors were located in upper or lower extremities. Histology were: 13 liposarcoma,11 MFH, 5 leiomiosarcoma, 5 PNST, 3 rabdo, 3 synovialsarcoma, 3 undifferentiated and 2 fibrosarcoma. Following Coindre classification 23 pts had low grade and 22 high grade STS.

Results: median follow up is 23 months (2–84). At present 20 pts (44.4%) are alive and DF, 11 (24.4%) alive with disease, 14 (31.1%) dead. Median survival is 75 months. Median MVD of all specimens is 62 microvessels/mm² (7–161). 32 pts (71.1%) have low MVD (group A) and 13 pts (29.9%) high MVD (group B). Mean survival is 62.7 mo in group A (median 75) and 36 mo in group B (median not reached) (p 0.01); median DFS respectively 24 mo and 15 mo (p 0.01). There is also a significant association with histological grade and survival: 75 mo in low grade and 34 mo in high grade tumors (p 0.05) and presence of metastasis at diagnosis (median survival: M+23 mo, M−75 mo). Unfortunately no relationship between angiogenesis and grading is found.

Conclusions: Our study confirms the prognostic importance of grade and staging in pts with STS. Moreover the role of MVD in prognosis is well defined and should be used as a routine marker in STS histological diagnosis.

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.