Shinya Yamawaki

ErbB2 expression is correlated with increased survival of patients with osteosarcoma

Elevated ErbB2 expression and gene amplification have been shown to be associated with poor prognosis in many cancers. Recently, it has been demonstrated that overexpression of ErbB2 protein in osteosarcoma is associated with the presence of pulmonary metastasis and decreased survival. By contrast, a previous study showed that the expression of ErbB2 declines in individual osteosarcomas as they become metastatic. In the current study, the authors determined the relation between ErbB2 status and outcome in a large number of selected patients with high‐grade osteosarcoma.

Diagnosis of Synovial Sarcoma with the Reverse Transcriptase-polymerase Chain Reaction: Analyses of 84 Soft Tissue and Bone Tumors

The chimeric transcript SYT-SSX is generated as a result of reciprocal translocation t(X;18), which is the primary cytogenetic abnormality found in, and appears to be specific for, synovial sarcoma. We performed a reverse transcriptase-polymerase chain reaction (RT-PCR) for SYT-SSX transcripts in a series of 84 tumors (61 soft tissue tumors and 23 bone tumors), including a variety of histologic types, to assess its usefulness in molecular diagnosis. Ten synovial sarcomas, three tumors initially unclassified, and one malignant peripheral nerve sheath tumor contained the chimeric transcripts. A review of the original slides and additional examination showed that a diagnosis of synovial sarcoma was appropriate for these cases. Additionally, in situ hybridization with an SSX1 probe indicated that the chimeric transcripts exist not only in the cells of special components but also in cells showing a variety of histologic patterns. Therefore, RT-PCR can be considered a useful molecular biological technique that can provide objective evidence for diagnosis of synovial sarcoma. Northern blot analysis with an SSX1 probe also detected chimeric SYT-SSX transcripts in the synovial sarcoma cases. The additional smaller bands, however, were also detected in six peripheral primitive neuroectodermal tumors (pPNETs) and one embryonal rhabdomyosarcoma. In five of these pPNETs, other bands ranging in size from 2.0 to 2.2 kb were also found, and it seems possible that these bands might represent novel karyotypic aberrations and/or splicing variants of SSX.

Increased pre-therapeutic serum vascular endothelial growth factor in patients with early clinical relapse of osteosarcoma

To investigate the clinical significance of circulating angiogenic factors, especially in association with early relapse of osteosarcoma, we quantified pre-therapeutic levels of vascular endothelial growth factor, basic fibroblast growth factor and placenta growth factor in the sera of 16 patients with osteosarcoma using an enzyme-linked immunosorbent assay. After a 1-year follow-up, the serum level of angiogenic factors was analysed with respect to microvessel density of the biopsy specimen and clinical disease relapse. The serum vascular endothelial growth factor levels were positively correlated with the microvessel density with statistical significance (P=0.004; Spearman rank correlation) and also significantly higher in seven patients who developed pulmonary metastasis than the remaining nine patients without detectable disease relapse (P=0.0009; The Mann–Whitney U-test). In contrast, the serum levels of basic fibroblast growth factor or placenta growth factor failed to show significant correlation with the microvessel density or relapse of the disease. Although there was no significant correlation between serum vascular endothelial growth factor levels and the tumour volume, the serum vascular endothelial growth factor levels were significantly higher in patients with a vascular endothelial growth factor-positive tumour than those with a vascular endothelial growth factor-negative tumour. These findings suggest that the pre-therapeutic serum vascular endothelial growth factor level reflects the angiogenic property of primary tumour and may have a predictive value on early disease relapse of osteosarcoma.

Multiinstitutional phase II study of neoadjuvant chemotherapy for osteosarcoma (NECO study) in Japan: NECO-93J and NECO-95J

BackgroundOsteosarcoma is the most frequent primary malignant bone tumor. In Europe and the United States, its prognosis has been greatly improved by the use of multimodal treatment, including preoperative and postoperative chemotherapy as well as surgery. In Japan, however, only a few clinical studies on osteosarcoma have been carried out.MethodsTo evaluate the efficacy of neoadjuvant chemotherapy on nonmetastatic, operable osteosarcoma arising in the extremities, a prospective multiinstitutional phase II trial, the Neoadjuvant Chemotherapy for Osteosarcoma (NECO) study, was conducted. Preoperative chemotherapy included high-dose methotrexate (HD-MTX), cisplatin (CDDP), and adriamycin (ADR). If the induction therapy was assessed as not effective, high-dose ifosfamide (IFO) was added to the chemotherapy regimen. A total of 124 patients were enrolled in this trial, and ultimately 113 patients were eligible.ResultsThe 5-year overall survival (OAS) and event-free survival (EFS) rates in the NECO study were 77.9% and 65.5%, respectively. A good histological response to the induction chemotherapy resulted in favorable OAS (78.7%). The patients assessed as poor histological responders with progressive disease after the induction chemotherapy exhibited comparable outcomes (OAS 89.5%, EFS 68.2%). There were no significant differences between the OAS and EFS rates of the patients in terms of response to preoperative chemotherapy.ConclusionsWe analyzed the results of the intensive neoadjuvant chemotherapy and the effects of adding IFO on patients with osteosarcoma in Japan. The results suggest efficacy of the high-dose IFO addition to the standard three-drug chemotherapy regimen. However, a randomized clinical study is needed to establish the true impact of IFO on patients with osteosarcoma.

Malignant granular cell tumor of the radial nerve. An autopsy observation with electron microscopic and tissue culture studies

Malignant granular cell tumor closely associated with the radial nerve in the left upper arm, was found in a 34‐year old man. The tumor increased rapidly in size, was locally invasive and showed recurrence and metastases to lymph nodes and the lungs. The tumor was composed solely of granular cells and its histological features were very similar to those of benign granular cell tumors. However, considerable variation of cellular size and shape, hyperchromatic plump nuclei, and some disordered cellular arrangement were thought to be significant indications of malignancy. Ultrastructural study of the tumor revealed many lysosome‐like dense granules in the cytoplasm. The presence of a few axon‐like cytoplasmic processes between granular cells and the intimate anatomical association of the tumor with the radial nerve seemed to support the concept of peripheral nerve origin of the malignant granular cell tumor.

Reconstruction and limb salvage after resection for malignant bone tumour of the proximal humerus: A SLING PROCEDURE USING A FREE VASCULARISED FIBULAR GRAFT

We assessed the intermediate functional results of eight patients after wide resection of the proximal humerus for malignant bone tumour. We used a free vascularised fibular graft as a functional spacer and a sling procedure to preserve passive scapulohumeral movement. Scapulohumeral arthrodesis was not carried out. Five patients had osteosarcoma, two achondrosarcoma and one a malignant fibrous histiocytoma of the bone. The mean duration of follow-up was 70 months (median, 76) for the seven patients who were still alive at the time of the latest follow-up. One patient died from the disease 12 months after surgery. There were no local recurrences. The functional results were described and graded quantitatively according to the rating system of the Musculoskeletal Tumour Society. Our results were satisfactory with regard to pain, emotional acceptance and manual dexterity. Function and lifting ability were unsatisfactory in two patients. One patient had delayed union between host and graft, but this united after six months without further surgery. Radiographs of the shoulder showed absorption or collapse of the head of the fibula in four of the eight patients and a fracture in another. No functional problems related to absorption or fracture of the head of the fibula were noted. There was no infection or subluxation of the head. We conclude that this is a reasonably effective technique of limb salvage after resection of the proximal humerus.

Loss of ErbB2 Expression in Pulmonary Metastatic Lesions in Osteosarcoma

Objectives: The c-erbB2 protooncogene is located on human chromosome 17 and encodes a 185-kilodalton transmembrane glycoprotein (ErbB2). Elevated ErbB2 expression or gene amplification has been shown to be associated with a poor prognosis in many cancers. Recently, it has been demonstrated that overexpression of the ErbB2 protein in osteosarcoma is associated with the presence of pulmonary metastasis and decreased survival. To further investigate the role of ErbB2 overexpression in pulmonary metastasis of osteosarcoma, its expression in the primary and metastatic lesions of the same osteosarcoma patients was compared. Methods: We compared the expression levels of ErbB2 receptor protein between the biopsy samples and pulmonary metastatic lesions in each of 19 patients with osteosarcoma who had not presented with metastasis at diagnosis. All archival materials from patients were retrieved and stained with monoclonal antibody CB11 to detect ErbB2 protein. Results: The rate of overexpression was significantly lower in the pulmonary metastatic tumors than in the biopsy samples (11 versus 42%; p = 0.03). Among 8 patients who had shown increased levels of ErbB2 in the biopsy samples, 7 exhibited complete absence of ErbB2 in the pulmonary metastatic lesions. Overall loss of ErbB2 expression was noted in 14 of 19 patients as the initial tumor became metastatic. Conclusions: It is suggested that overexpression of ErbB2 decreases within individual osteosarcomas as they become metastatic. Overexpression of ErbB2 may not play an important role in the development of pulmonary metastases of osteosarcoma. Further data are needed before ErbB2 can be used in making clinical decisions for osteosarcoma patients.

A Preliminary Report of Neoadjuvant Chemotherapy NSH-7 Study in Osteosarcoma: Preoperative Salvage Chemotherapy Based on Clinical Tumor Response and the Use of Granulocyte Colony-Stimulating Factor

Eleven patients with high-grade osteosarcoma of an extremity were treated with neoadjuvant chemotherapy with NSH-7 protocol. NSH-7 is a refinement of the T-12 Rosen protocol. Preoperative chemotherapy is initiated with a doxorubicin (ADM) and high-dose methotrexate combination. If the primary tumor progresses after the first cycle, the preoperative chemotherapy is switched to a combination of cisplatin and ADM. Postoperative adjuvant chemotherapy was selected based on histological response of the primary tumor. In addition, recombinant human granulocyte colony-stimulating factor was used to prevent leukocytopenia and to increase the dose intensity of the chemotherapy. In 1 patient, preoperative chemotherapy was switched to salvage treatment. Of the 156 courses given, there were 10 delays and 4 dose reductions. Leukocytopenia accounted for only 1 delay. All 11 patients completed the chemotherapy and 5 patients were fully able to tolerate the protocol without delay or dose reduction. Nine patients remained alive and continuously free of disease at an average follow-up of 35 months. The rate of continuous disease-free survival at 3 years was 81%, which was significantly better than that of the T-12 study of our group. These observations suggest that the NSH-7 protocol is a safe and effective treatment regimen for osteosarcoma.

Establishment of a new continuous clear cell sarcoma cell line. Morphological and cytogenetic characterization and detection of chimaeric EWS/ATF-1 transcripts.

Abstract Clear cell sarcoma (CCS), a rare tumour of deep soft tissues, often has a t(12; 22) (q13; q12) translocation that induces the formation of a hybrid EWS/ATF-1 gene. To investigate these alterations further, we established a new continuous cell line directly from a CCS taken from a 9-year-old girl. The cultures were characterized with respect to morphological, ultrastructural, immunohistochemical and karyotypical features and were tested by reverse transcription PCR (RT-PCR) for chimaeric EWS/ATF-1 transcripts. The continuous cell line, designated KAO, is tumorigenic in nude mice, and the resultant tumours resemble the primary CCS. The tumour cells and the cultured cells have melanosomes in their cytoplasm and are immunoreactive with the melanoma-specific antibody HMB45, but do not express S-100 protein. The cultured CCS cells have the t(12; 22)(q13; q12) translocation and express the hybrid EWS/ATF-1 gene. No transcripts of the hybrid gene were detected in a malignant cutaneous melanoma tested simultaneously. Although CCS and malignant melanoma are morphologically related, the present results suggest that their geneses differ at the chromosome and molecular levels. They also indicate that chromosome analysis and detection of fusion EWS/ATF-1 transcripts may be useful adjuvant tools for the diagnosis of CCS.

Histological and Molecular Evidence of Synovial Sarcoma of Bone. A Case Report

Synovial sarcoma is a clinically and morphologically well defined soft-tissue tumor that occurs predominantly in the extremities of adolescents and young adults. It tends to arise in the vicinity of large joints, especially the knee8. This tumor was designated synovial sarcoma because of its anatomical location and its histological resemblance to normal synovial tissue. However, we do not believe that there is any convincing evidence that synovial sarcoma always originates from synovial tissue. It is possible that the lesion sometimes arises from primitive mesenchymal cells rather than from preformed synovial cells; therefore, the development of synovial sarcoma is no longer thought to require the presence of preexisting synovial tissue. To the best of our knowledge, there have been no reports of synovial sarcoma arising in bone. The reciprocal translocation t(X;18)(p11.2;q11.2) was noted in all twenty-two reported cases of synovial sarcoma that have been studied cytogenetically5,18. This translocation is considered to be specific to this tumor. Cloning of the DNA adjacent to the breakpoints of the translocation t(X;18) has shown2 that this translocation results in fusion of the SYT gene at 18q11.2 to either of two genes, SSX1 or SSX2, at Xp11.2. This rearranged configuration of DNA is transcribed to messenger RNA and is called a chimeric SYT-SSX transcript. A chimeric SYT-SSX transcript does not occur normally and thus provides a specific marker for synovial sarcoma that can be detected with reverse transcriptase-polymerase chain-reaction testing. We describe the case of a patient who had synovial sarcoma of bone that was verified by the detection of a chimeric SYT-SSX transcript. A sixty-seven-year-old man had a four-month history of pain in the right wrist when he was first seen by us. Physical examination revealed moderate swelling and a restricted range of motion of the wrist (30 …