The soft tissue sarcomas (STS) are a diverse collection of malignant tumours of the connective tissues arising from the primitive mesoderm and ectoderm. While the primary treatment of most is surgery, chemotherapy can be offered to patients presenting with locally advanced or metastatic disease although sarcomas are resistant to the majority of anticancer drugs. The reasons for this are not fully understood but it is thought that p53 abnormalities and mdm2 overexpression may be involved. Samples from twenty eight adult patients with soft tissue sarcomas have been analysed for p53 mutations in exons 4 to 9 both by denaturing high performance liquid chromatography (dHPLC) and by direct automated sequencing. By sequencing we found mutations in 7/28 patients, giving a mutation rate of 25%. 4/6 were point mutations in exons 5, 7 and 8 and the remaining three were deletions in exons 4, 7 and 8. Six of these samples gave abnormalities in dHPLC analysis with a concordance rate of 97.5% between the sequencing and dHPLC data. Thirty nine and forty samples have been assessed by immunohistochemistry for p53 and mdm2 expression respectively. Do7 antibody which recognises the N terminus of p53 and F4-14 which recognises the carboxy-terminus of mdm2 were used. Immunohistochemistry was scored semiquantitatively by two independent observers and the results scored accordingly: low (< 20%), intermediate (20–80%) and high (> 80%). The initial results showed that 23/40 (58%) of patients were high staining for mdm2 in contrast to only 15/39 (38%) of patients for p53. All patients with deletions in p53 had intermediate staining for mdm2. 2/3 of these had intermediate staining for p53 and 1/3 had high staining for p53. One patient with a point mutation had high staining for both p53 and mdm2 but the other two have yet to be analysed by immunohistochemistry. These results confirm the overexpression of mdm2 in STS. Future experiments are planned using fluorescent in situ hydridisation (FISH) to determine whether MDM2 amplification is one of the mechanisms involved in mdm2 overexpression

Epithelioid hemangioendothelioma (EHE) is a rare vaso-formative tumor of variable biological behavior that has been considered a tumor of borderline malignancy and low-grade angiosarcoma. The majority of cases are associated with low mortality, but some metastasize and cause patient death. Its principal sites of occurrence are soft tissues, liver, lung, and bone. EHE develops as a solitary, painful mass in superficial or deep soft tissue of the extremities and it generally arises from a vessel. Cytogenetic findings are very limited and comprises three reports on totally 4 cases, describing translocations between chromosomes 1 and 3 in two cases, chromosomes 7 and 22 in one case and chromosome 10 and 14 in the last case. We characterized immunohistochemically 5 cases of this tumour type and currently we are performing Real-Time PCR assays to analyze the expression of two genes (MDM2 and CDK4) known to be involved in pathogenesis of tumours. Three out 5 patients presented epithelioid hemangioendothelioma of the bone while two affected soft tissues. All the samples showed positivity for CD34 and CD31; 4 samples out 5 were also positive for FLI1. We tested Factor VIII immunostaining on 3 of these cases which resulted positive; EMA was positive on 3 samples out 5. Cytocheratins (AE1/AE3, CAM 5.2 and CK7) were always negative except in one case which showed CAM 5.2 positivity. Our preliminary results by Real-Time PCR analysis performed on 5 cases suggest that MDM2 and CDK4 have a different expression in epithelioid hemangioendotheliomas compared to normal tissue. Our study shows that use of molecular techniques such as Real-Time PCR could complement histopathological diagnosis in order to identify a marker of biologic behaviour of this enigmatic tumour type

Objectives

Osteoporosis is a chronic disease. The aim of this study was to identify key genes in osteoporosis.