Overexpression of p53 protein in primary Ewing's sarcoma of bone: relationship to tumour stage, response and prognosis.
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Biopsy tissues of 52 patients with Ewing's sarcoma of bone treated between 1983 and 1993 were examined immuno-histochemically to determine the significance of p53 protein in diagnosis and prognosis of Ewing's sarcoma. Mean age at diagnosis was 17 years (range 6-36) and minimum follow-up was 30 months. The tumours were located in the extremities and central bones in 35 and 17 patients respectively. Metastases were present in seven patients at diagnosis. Treatment consisted of chemotherapy, surgery and/or radiotherapy in all the patients. Overexpression of p53 protein was demonstrated in seven patients (14%). There was no relationship between expression of p53 and site of tumours. Patients who overexpressed p53 protein appeared to have more advanced diseases at diagnosis and poorer response to chemotherapy than those without p53 overexpression. The 5-year relapse-free survival and overall survival in patients without metastases at the time of diagnosis were 66% and 71%, respectively, in p53 protein-negative patients compared with 20% relapse-free and overall survival in those with p53 protein overexpression (P= 0.01). The poorer prognosis in p53 protein-positive patients was independent of site, local treatment or necrosis of the tumours (P < 0.05). Over-expression of p53 protein is an independent poor prognostic factor in Ewing's sarcoma of bone. (+info)
Measurement of DNA cross-linking in patients on ifosfamide therapy using the single cell gel electrophoresis (comet) assay.
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The single cell gel electrophoresis comet assay has become established as a sensitive technique for measuring DNA strand breaks. The technique has been modified to allow the sensitive detection and quantitation of DNA interstrand cross-linking at the single cell level. Cells are irradiated immediately before analysis to deliver a fixed level of random strand breakage. After embedding of cells in agarose and lysis, the presence of cross-links retards the electrophoretic mobility of the alkaline denatured cellular DNA. Cross-links are, therefore, quantitated as the decrease in the comet tail moment compared with irradiated controls. Using this method, a linear response of cross-linking versus dose of chlorambucil over a wide dose range was demonstrated in human lymphocytes after drug treatment ex vivo. The method was also sensitive enough to determine cross-linking in clinical samples after chemotherapy. For example, crosslinking was observed in the lymphocytes of patients receiving ifosfamide (3 g/m2/day) as a continuous infusion for 3-5 days or as a 3-h infusion daily for 3 days. Cross-links were detected in all patients within 3 h, with no evidence of DNA single strand break formation. In patients receiving continuous infusion, a plateau of cross-linking was reached by 24 h. In the patients receiving ifosfamide over 3 h, a clear decrease in the peak level of cross-linking was observed before subsequent infusions. (+info)
Differential transactivation by alternative EWS-FLI1 fusion proteins correlates with clinical heterogeneity in Ewing's sarcoma.
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The t(11;22)(q24;q12) translocation is present in up to 95% of cases of Ewing's sarcoma and results in the formation of an EWS-FLI1 fusion gene which encodes a chimeric transcription factor. The proximate role of EWS-FLI1 in the pathogenesis of Ewing's sarcoma is thought to involve the activation of as yet largely unknown target genes. Many alternative forms of EWS-FLI1 exist because of variations in the locations of the EWS and FLI1 genomic breakpoints. The most common form, designated "type 1," consists of the first seven exons of EWS joined to exons 6-9 of FLI1 and accounts for approximately 60% of cases. The "type 2" EWS-FLI1 fusion also includes FLI1 exon 5 and is present in another 25%. We and others have observed previously that the type 1 fusion is associated with a significantly better prognosis than the other fusion types. Because EWS-FLI1 is an aberrant transcription factor, we investigated whether these differences in clinical behavior may be correlated to functional differences by comparing transactivation by the type 1 EWS-FLI1 with other types in both heterologous cells (HeLa, NIH3T3) and homologous cells (Ewing's sarcoma cell lines). In a panel of seven Ewing's sarcoma cell lines, we found transactivation of a transiently transfected FLI1-responsive reporter construct to be significantly lower in cell lines with the type 1 fusion than in cell lines with the type 2 fusion (P = 0.003). Cotransfection of the same reporter construct with each of a series of seven EWS-FLI1 expression constructs (corresponding to the two major fusion types and five less common types) also showed that type 1 EWS-FLI1 was a significantly weaker transactivator than the type 2 product in both HeLa and NIH3T3 cells (P = 0.003, and P = 0.033, respectively). Electromobility shift assays showed equivalent binding of the type 1 and type 2 EWS-FLI1 to the consensus FLI1-responsive binding site, indicating that differences in transactivation were not due simply to differences in DNA binding affinity. The finding that the type 1 EWS-FLI1 fusion, associated with less aggressive clinical behavior, encodes a less active chimeric transcription factor may provide the basis for a molecular explanation of clinical heterogeneity in Ewing's sarcoma. (+info)
Tumour volume as a predictor of necrosis after chemotherapy in Ewing's sarcoma.
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We studied the CT and MR scans, and the histology of 50 patients with primary Ewing's sarcoma of bone to determine the association between the change in tumour volume and necrosis after chemotherapy, and to ascertain their influence on prognosis. The mean age of the patients was 17 years. The limbs were involved in 40 and the axial bones in ten. The volume of the tumour at diagnosis varied from 31 to 1790 ml. There was a significant relationship between necrosis and the measured change in volume of the tumour after chemotherapy. Progression of the tumour despite chemotherapy was seen only in patients with necrosis of grades 4 to 6. Necrosis significantly influenced survival (p < 0.05), but the effect of change in volume was less significant. Change in volume of the tumour is a good predictor of necrosis induced by chemotherapy. Necrosis is a strong prognostic factor in Ewing's sarcoma. (+info)
Tyrosine kinase Pyk2 mediates G-protein-coupled receptor regulation of the Ewing sarcoma RNA-binding protein EWS.
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Ewing family tumors result from the effects of chromosomal translocations that fuse the Ewing sarcoma (EWS) gene to various genes encoding transcription factors. The resulting chimeric EWS fusion proteins are transcriptional activators with transforming potential that have received much study. By contrast, the cellular function of somatic EWS remains obscure. EWS belongs to a family of RNA-binding proteins thought to play role in RNA synthesis or processing. Here, we show that EWS interacts with Pyk2, a protein tyrosine kinase implicated in a variety of signal transduction processes. G-protein-coupled receptor signaling and other stimuli of Pyk2 kinase activity significantly block the interaction between EWS and Pyk2. Furthermore, as assessed by sucrose gradient centrifugation, EWS partitions with dense ribosome-containing fractions in a manner that is enhanced by signaling from the G-protein-coupled m1 muscarinic acetylcholine receptor (mAChR). We conclude that extranuclear EWS is a previously unrecognized target of G-protein-coupled receptor regulation. (+info)
Management of cancer in pregnancy: a case of Ewing's sarcoma of the pelvis in the third trimester.
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Ewing's sarcoma of the pelvic bones was diagnosed in a 21-year childbearing woman, raising major medical and ethical problems. The diagnostic and therapeutic approaches during the sixth month of gestation were tailored in order to cure the patient and avoid unnecessary toxicity to the fetus. Ancillary tests included ultrasound and MRI studies of the pelvis. Ifosfamide and adriamycin, premedicated by granisetron, were administered during gestation, and were found to be safe. Cesarean section was the preferred way of delivery since the tumor involved the pelvic bones. The outcome was a disease-free patient and a small healthy baby who is now two years of age. (+info)
The EWS/TEC fusion protein encoded by the t(9;22) chromosomal translocation in human chondrosarcomas is a highly potent transcriptional activator.
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The EWS/TEC gene fusion generated by the t(9;22) chromosomal translocation found in extraskeletal myxoid chondrosarcomas encodes a fusion protein containing the amino-terminal domain of the EWS protein fused to the whole coding sequence of the orphan nuclear receptor TEC. We have compared the DNA-binding and transcriptional activation properties of various TEC isoforms and the corresponding EWS/TEC fusion proteins. Band-shift experiments show that the full-length TEC receptor can efficiently bind the NGFI-B Response Element (NBRE), whereas an isoform lacking the entire carboxyl-terminal domain of the receptor binds much less efficiently the NBRE. Addition of the amino-terminal domain of EWS to either isoforms does not alter significantly their DNA-binding properties to the NBRE. Co-transfection experiments of COS cells and human chondrocytes indicate that whereas TEC moderately activates transcription from a NBRE-containing promoter, the corresponding EWS/TEC fusion protein is a highly potent transcriptional activator of the same promoter, being approximately 270-fold more active than the native receptor. EWS/TEC may thus exert its oncogenic potential in chrondrosarcomas by activating the transcription of target genes involved in cell proliferation. (+info)
Brain metastases in musculoskeletal sarcomas.
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BACKGROUND: In musculoskeletal sarcomas, brain metastases are rare, but severely affect quality of life. METHODS: All patients with musculoskeletal sarcomas who were treated at our institutions from 1975 to 1997 were reviewed for examples of brain metastasis. RESULTS: Of 480 sarcoma patients, 179 had distant metastases, including 20 patients with brain metastases (4.2%). Alveolar soft part sarcoma (3/4), extraskeletal Ewing's sarcoma (2/8), rhabdomyosarcoma (2/13) and bone Ewing's sarcoma (2/18) tended to metastasize to the brain. All 20 patients had distant or local relapses and 16 of the 20 patients had pulmonary metastases. Three patients underwent surgical treatment and two of them survived over 1 year. Mean survival after diagnosis of brain metastasis was 5.1 months. CONCLUSIONS: Patients with alveolar soft part sarcoma, Ewing's sarcoma, rhabdomyosarcoma and pulmonary metastases have a high risk of brain metastasis. (+info)