Hirofumi Nakanishi

Immunohistochemical Detection of bcl-2 and p53 Proteins and Apoptosis in Soft Tissue Sarcoma: Their Correlations with Prognosis

Information on prognostic factors is essential to establish appropriate therapeutic modalities for soft tissue sarcoma (STS). To evaluate the biological nature and prognostic factors of STS, p53 and bcl-2 expression was immunohistochemically studied on paraffin-embedded sections from 70 patients with STS in the extremities and trunk. In addition, the degree of apoptosis was examined by in situ end-labeling. Histologic diagnoses in these cases were malignant fibrous histiocytoma in 29 cases, liposarcoma in 11, synovial sarcoma in 11, leiomyosarcoma in 5, malignant neurogenic tumor in 5, and others in 9. Tumor cells in 31 of 70 cases (44%) showed positive nuclear staining for p53 protein. There was no correlation between p53 expression and tumor size, histologic grade, argyrophilic nucleolar organizer region (AgNOR) count, cellularity and extent of neerosis. Expression of p53 did not correlate with survival of patients. Tumor cells in 24 of 56 cases (43%) were positive for bcl-2 protein expression. The frequency of bcl-2 expression in the tumor cells showed a direct proportion to tumor size (> or = 10 vs. < 10 cm) but inverse proportion to AgNOR counts and cellularity. The 5-year survival rate in patients with bcl-2-positive tumors (87%) was more favorable than in those with bcl-2-negative tumors (53%; p < 0.05). The frequency of apoptosis in low-grade STS was significantly higher than that in the intermediate and high-grade STS (p < 0.001). Extent of necrosis, a well-known prognostic indicator in STS, was not correlated with the frequency of apoptosis. Multivariate analysis showed that cellularity, bcl-2 and AgNOR counts were independent prognostic factors in patients with STS. The current study revealed that STS with a higher expression of bcl-2 had lower proliferative activity and larger size than those without. Immunohistochemical detection of bcl-2 is useful for predicting prognosis in patients with STS.

Mutations of p53 tumor-suppressor gene in angiosarcoma

Transgenic mice deficient for the p53 gene were reported to frequently develop angiosarcoma (AS), suggesting that alterations in the gene are associated with tumorigenesis of AS. However, little is known about genetic changes, including p53 gene alterations, in human AS because of its rarity. We analyzed p53 mutations on paraffin‐embedded specimens from 33 patients with AS by polymerase chain reaction–single‐strand conformation polymorphism (PCR‐SSCP) followed by direct sequencing. Age of patients ranged from 18 to 91 (median 70) years, with a male to female ratio of 1.5:1. Sites of tumor were the head in 13 patients, the trunk in 4, the extremities in 4, the heart in 4, bones in 2 and others in 6. PCR‐SSCP revealed aberrant mobility shifts of bands in 17 cases: 11 in exon 5, 5 in exon 7 and 4 in exon 8. Direct sequencing on these 17 cases revealed a total of 20 mutations. The frequency of p53 mutations was different by site of tumors: 7 of 13 in head, all 4 in extremities, 2 of 4 in heart and none of 4 in trunk. Our findings suggest that occurrence of p53 mutation is a major pathway for development of human AS.Int. J. Cancer 71: 952‐955, 1997.

Cystic synovial sarcomas: imaging features with clinical and histopathologic correlation.

ObjectiveTo characterize the radiological and clinicopathologic features of cystic synovial sarcoma.Design and patientsSeven patients with primary cystic synovial sarcoma were evaluated. Computed tomography (CT) and magnetic resonance (MR) imaging were undertaken at the first presentation. The diagnosis of synovial sarcoma was made on the basis of histological examinations followed by molecular analysis. Radiological and clinicopathologic findings were reviewed.ResultsCT showed well-defined soft tissue mass without cortical bone erosion and invasion. Calcification was seen at the periphery of the mass in three cases. T2-weighted MR images showed multilocular inhomogeneous intensity mass in all cases, five of which showed fluid-fluid levels. On gross appearance, old and/or fresh hematomas were detected in six cases. In the one remaining case, microscopic hemorrhage in the cystic lumen was proven. Four cases had poorly differentiated areas. In five cases prominent hemangiopericytomatous vasculature was observed. Histologic grade was intermediate in one tumor and high in six. One case had a history of misdiagnosis for tarsal tunnel syndrome, one for lymphadenopathy, two for sciatica and two for hematoma.ConclusionAll cystic synovial sarcomas demonstrated multilocularity with well-circumscribed walls and internal septae. Synovial sarcoma should be taken into consideration in patients with deeply situated multicystic mass with triple signal intensity on T2-weighted MR imaging.

RENAL NEOPLASIAS IN PATIENTS RECEIVING DIALYSIS AND RENAL TRANSPLANTATION: CLINICO-PATHOLOGICAL FEATURES AND P53 GENE MUTATIONS

BACKGROUND In Japan, the relative risk for renal cell carcinoma (RCC) in renal transplants was about 80-fold higher than that in the general population. Depressed immune surveillance due to the use of immunosuppressive agents was considered to cause cancer. Before renal transplantation, a vast majority of patients received hemodialysis, a known causative factor for acquired cystic disease of kidney (ACDK). Because ACDK is also considered to predispose to RCC, at least two risk factors for cancer accumulate in renal transplants. METHODS In our study, clinicopathological features together with p53 gene mutations were analyzed in 218 patients with RCC: 22 received dialysis followed by renal transplantation, 39 received dialysis alone, and 157 sporadic RCC. P53 mutations were analyzed on DNA extracted from paraffin-embedded specimens with use of single strand conformation polymorphism, followed by direct sequencing. RESULTS RCC in transplants shared several clinicopathological features with those in dialysis patients, which included small size and multiplicity of tumor, relatively high frequency of presence of ACDK, and papillary type of RCC. p53 gene mutations were infrequent in RCC of any clinical setting. CONCLUSIONS Atrophic kidney at the end-stage of renal failure and under dialysis have lesions of ACDK that might predispose to RCC in dialysis and transplant patients.

Clinical implications of serum C-reactive protein levels in malignant fibrous histiocytoma.

Paraneoplastic syndromes (PNSs) associated with mesenchymal tumors are uncommon. Previous reports sporadically described inflammatory PNSs with elevated serum C‐reactive protein (CRP) levels and leukocytosis in patients with inflammatory malignant fibrous histiocytoma (MFH) of soft tissue; however, the relationship between other subtypes of MFH and PNS has not been extensively investigated. Forty‐six patients with primary MFH of soft tissues who underwent radical surgery were retrospectively analyzed. These patients were divided into 2 groups according to preoperative serum CRP level: normal (<1.0 mg/dl) and elevated (≥1.0 mg/dl). The correlation between serum CRP level and several clinicopathologic factors was analyzed. Correlation between preoperative serum CRP level and metastasis‐free and overall survival was also investigated by univariate and multivariate analyses. Elevated preoperative serum CRP levels were found in 65% of patients with a mean of 3.7 mg/dl. There were statistically significant relationships regarding tumor size, depth, histologic subtypes, grade, stage and metastatic rate among normal and elevated CRP groups (p < 0.001, p < 0.02, p < 0.005, p < 0.001, p < 0.001 and p < 0.05, respectively). When the tumor was removed, the elevated CRP level subsided into the normal range and other abnormal laboratory findings diminished in all cases. In 11/14 relapsed cases that showed elevated CRP preoperatively, the serum CRP level re‐elevated with tumor relapse. The normal CRP group showed significantly more favorable prognosis than the elevated CRP group in metastasis‐free and overall survival on univariate analysis (p < 0.02, p < 0.05, respectively). Patients with MFH frequently present with an inflammatory PNS, such as elevated serum CRP level, which can be a useful marker of disease activity and a valuable prognostic indicator.

Expression of SSX genes in human osteosarcomas

Norifumi NAKA*, Nobuhito ARAKI, Hirofumi NAKANISHI, Kazuyuki ITOH, Masayuki MANO, Shingo ISHIGURO, Diederick R.H. DE BRUIJN, Akira MYOUI, Takafumi UEDA and Hideki YOSHIKAWA Department of Orthopaedic Surgery, Osaka Medical Center for Cancer and Cardiovascular Diseases, Osaka, Japan Department of Biology, Osaka Medical Center for Cancer and Cardiovascular Diseases, Osaka, Japan Department of Pathology, Osaka Medical Center for Cancer and Cardiovascular Diseases, Osaka, Japan Department of Human Genetics, University Medical Center, Nijmegen, the Netherlands Department of Orthopaedics, Osaka University Medical School, Osaka, Japan

P-glycoprotein expression in soft-tissue sarcomas.

Multidrug resistance (MDR) is an important problem in chemotheraphy for neoplastic disease. In humans, MDR is mainly mediated by P-glycoprotein (P-gp), a product of theMDR1 gene, which acts as a transmembrane protein pump and eliminates chemotherapeutic agents from the cells. Expression of P-gp was immunohistochemically studied by using two monoclonal antibodies, JSB-1 and C-219, on paraffin-embedded sections from 55 patients with soft-tissue sarcoma. The histological diagnosis of tumors was malignant fibrous histiocytoma in 24 cases, liposarcoma in 9, synovial sarcoma in 7, malignant neurogenic tumors in 6, leiomyosarcoma in 5, others in 4. The histological grade was determined on the basis of criteria previously proposed by us. Out of 55 cases, 34 (62%) were positive for P-gp expression. There was a significant difference in P-gp expression between high-grade (90%) and intermediate and low-grade tumors (46%) (P<0.005). Tumors expressing P-gp had a less favorable prognosis than P-gp-negative tumors in the high- and intermediate-grade tumors. The current study demonstrated that the estimation of P-gp expression could be used to select appropriate therapeutic modalities.

Interleukin-6/soluble interleukin-6 receptor signaling attenuates proliferation and invasion, and induces morphological changes of a newly established pleomorphic malignant fibrous histiocytoma cell line.

Pleomorphic malignant fibrous histiocytoma (MFH) is occasionally associated with inflammatory paraneoplastic syndrome (PNS). Recently, we reported that interleukin (IL)-6, one of the candidate cytokines, which induces such systemic inflammatory reaction, may be a tumor-associated factor involved in the pathogenesis and its clinical manifestations of MFH. In the local microenvironment, tumor-induced inflammatory reaction may play a role favoring tumor progression. To clarify the biological relevance of IL-6 in MFH, we established a human MFH cell line, named MIPS-2, derived from a resected specimen of a patient presenting with PNS. In this patient, the serum IL-6 level ran parallel to the disease course: elevated serum IL-6 concentration normalized immediately after radical surgery, and re-elevation occurred on tumor recurrence. MIPS-2 presented pleomorphic appearance, severe nuclear abnormalities with prominent nucleoli, and tumorigenesis in nude mice. MIPS-2 expressed IL-6, IL-6 receptor (IL-6R), and glycoprotein 130 (gp130) but lacked the soluble form of IL-6R (sIL-6R), as determined by flow cytometry and reverse transcriptase-polymerase chain reaction analyses. Stimulation of MIPS-2 with IL-6 combined with exogenous sIL-6R induced phosphorylation of both signal transducer and activator of transcription 3 (STAT3) and mitogen-activated protein kinase (MAPK), decreased cell proliferation, attenuated invasion, and induced morphological changes. Collectively, these data suggested that the IL-6/sIL-6R signaling pathway plays a pivotal role for proliferation, invasion, and morphology of MFH via STAT3 and MAPK pathway as autocrine and/or paracrine manner, and proposed the therapeutic potential for the use of both anti-growth factor and proinflammatory cytokine-targeting strategies to combat devastating MFH.

Neoadjuvant and adjuvant chemotherapy with high-dose ifosfamide, doxorubicin, cisplatin and high-dose methotrexate in non-metastatic osteosarcoma of the extremities: a phase II trial in Japan

Abstract From 1997 to 2003, 40 patients (all <40 years of age) with non-metastatic osteosarcoma of the extremities were treated with OOS-D and definitive surgery. Two cycles of doxorubicin 90 mg/m2 plus cisplatin 120 mg/m2 and ifosfamide 15 g/m2 were given as neoadjuvant chemotherapy, and two cycles of doxorubicin/cisplatin and ifosfamide, and two cycles of high-dose methotrexate (10–12 g/m2) were given post-operatively. All patients underwent limb salvage surgeries, and 66% showed good response to neoadjuvant chemotherapy. With a median follow-up period of 117 months, 31 of the evaluable 40 patients were continuously disease-free, 7 were currently alive with no evidence of disease, and 2 died of disease. There was no local recurrence. The 5-year event-free and overall survival rates were 83 and 98%, respectively. The 10-year event-free and overall survival rates were 80 and 95%, respectively. The major form of toxicity was haematological one.

Skeletal metastasis in patients with gastric cancer

To clarify metastatic patterns, and histologic and radiologic features in skeletal metastases from gastric cancer, 48 patients were retrospectively analyzed. The mean age of the patients at the time of diagnosis of gastric cancer was 59 years. In 31 patients with a history of the radical surgery, the mean interval between surgery and diagnosis of skeletal metastasis was 14 months. The mean duration between diagnosis of skeletal metastasis and death was 60 days. Scintigraphic assessment showed that solitary osseous lesions were found in four patients, whereas the remaining 44 had multiple skeletal lesions. In 28 patients with bone-only metastases with absence of visceral metastases, a higher incidence of thoracolumbar metastases at the level nearest the stomach was found. The incidence of skeletal metastasis in each histologic type was intestinal in 19 and diffuse in 29. Radiologic examination revealed that the ratio between the presence and the absence of osteosclerosis was 1:2. Osteosclerosis was seen in three of 19 patients with intestinal type metastasis, whereas with the diffuse type 13 of 29 patients had osteosclerosis.