Mitsuaki Suzuki

Clinical characteristics of clear cell carcinoma of the ovary

A retrospective review of treatment results comparing women with clear cell carcinoma of the ovary (CCC) with a group with serous adenocarcinoma of the ovary (SAC) was conducted.

Low proliferation activity may be associated with chemoresistance in clear cell carcinoma of the ovary

OBJECTIVE To estimate whether and how the biologic behavior of clear cell carcinoma contributes to the chemoresistance mechanism. METHODS Forty‐one patients with clear cell carcinoma and 90 patients with serous adenocarcinoma, who had measurable disease after initial surgery, were examined. All patients underwent cytoreductive surgery followed by platinum‐based chemotherapy. P‐glycoprotein, multidrug resistance‐associated protein, and Ki‐67 expression were determined by immunohistochemical staining. RESULTS The 5‐year survival rate for patients with clear cell carcinoma was significantly poorer, compared with serous adenocarcinoma (20.0% versus 31.9%). Response rate to chemotherapy was 14.6% for clear cell carcinoma and 72.2% for serous adenocarcinoma. The expression of P‐glycoprotein and multidrug resistance‐associated protein did not differ between responders and nonresponders in both tumor types. The Ki‐67 labeling index (LI) in clear cell carcinoma was significantly lower than serous adenocarcinoma (18.4% versus 38.8%). The LI for responders was significantly higher than that for nonresponders in both tumor types. In clear cell carcinoma, the mean value of LI was 15.3% for nonresponders, but that for responders was 30.2%, which was similar to that for serous adenocarcinoma. When the cutoff value of LI was set at 18.4% (mean value), the 5‐year survival rate for high LI (over 18.4%) patients was significantly greater than that for low LI patients (46.3% versus 9.2%). Multivariable analysis revealed that LI and residual tumor size were the independent prognostic factors. CONCLUSION Lower proliferation of tumor may be a behavior of clear cell carcinoma of the ovary that contributes to its resistance to chemotherapy.

PTEN expression is associated with prognosis for patients with advanced endometrial carcinoma undergoing postoperative chemotherapy

The prognostic significance of PTEN expression in endometrial carcinoma has not been clear. We conducted the present study to clarify the relationship between PTEN expression and prognosis in advanced endometrial carcinoma. Of 784 patients with endometrial carcinoma who underwent primary treatment between 1985 and 2000 at 5 institutions, 98 pure endometrioid carcinomas with retroperitoneal lymph node metastasis were provided for our study. PTEN expression was determined by immunohistochemic staining. Negative or mixed PTEN staining was observed in 64 (65.3%) patients. The survival rate for PTEN‐positive patients was significantly higher than that for PTEN‐negative or ‐mixed patients. PTEN‐staining status was not associated with patient age, International Federation of Gynecology and Obstetrics (FIGO) stage, myometrial invasion or histologic grade. Of the 98 patients, 87 received radiation therapy (n = 25) or chemotherapy (n = 62) after surgery. PTEN expression did not relate to survival for patients receiving radiation therapy. In contrast, the survival rate for PTEN‐positive cases was significantly higher than that for PTEN‐negative or ‐mixed cases when patients underwent chemotherapy (62.4% vs. 11.8%). Subsequent multivariate analysis revealed that PTEN staining was an independent prognostic factor for patients undergoing chemotherapy. PTEN‐positive staining was a significant prognostic indicator of favorable survival for patients with advanced endometrial carcinoma who underwent postoperative chemotherapy.

Expression of HGF/NK4 in ovarian cancer cells suppresses intraperitoneal dissemination and extends host survival

Peritoneal dissemination is the most frequent progression pathway of ovarian cancer and is therefore a key step to improve the prognosis. NK4, a large part of the α-chain of hepatocyte growth factor, is known to inhibit cancer cell migration. To characterize the function of NK4 and investigate its potential role in gene therapy of ovarian cancer, we introduced NK4 cDNA to an ovarian cancer cell line HRA and investigated its effects both in vitro and in vivo. HRA cells were transfected with either NK4 or luciferase-expression plasmids. After selection, NK4-expressing HRA cells (HRA/NK4) and the control cells (HRA/LUC) were obtained. NK4 was detected in the culture supernatant of HRA/NK4 by Western analysis. Migration capabilities of the cells were evaluated in vitro by scratch wound healing assay. The number of migrated cells was significantly smaller in the HRA/NK4 cultures than that in the control cultures (HRA or HRA/LUC). Also, the culture supernatant of HRA/NK4 significantly suppressed migration of control cells. This suppressive effect was observed when NK4-expressing cells were mixed with control cells at the ratio of 25% or more. In the in vivo experiments, HRA transfectants were injected intraperitoneally. The number of intraperitoneal tumors of HRA/NK4 was much smaller than that of control. In mice injected with HRA/NK4, ascites formation was suppressed and the survival was significantly prolonged. These findings suggest that NK4-mediated gene therapy can improve the prognosis of ovarian cancer by suppressing peritoneal dissemination. Gene Therapy (2001) 8, 1450–1455.

Mechanisms of Cisplatin Resistance in Clear Cell Carcinoma of the Ovary

Resistance of clear cell carcinoma (CCC) of the ovary to platinum-based chemotherapy is associated with a poor prognosis. However, the mechanism underlying the resistance of CCC to platinum has not yet been understood. We conducted the present study to clarify the mechanism of cisplatin (CDDP) resistance in CCC cells. Eleven CCC and 5 serous adenocarcinoma (SAC) cell lines were used in this study. The IC50 to CDDP ranged from 1.3 to 18.0 µM for CCC cells and from 2.2 to 13.0 µM for SAC cells. There was no correlation between multidrug resistance-associated protein expression and the sensitivity to CDDP in CCC cells. In contrast, the doubling time for CCC cells was significantly longer than that for SAC cells (61.4 vs. 29.8 h). A significant reverse correlation between the S-phase fraction and the response to CDDP was observed (r = 0.647, p < 0.05). The present study suggests that the resistance of CCC to CDDP may be caused by low cell proliferation.

Erythropoietin synthesis by tumour tissues in a patient with uterine myoma and erythrocytosis

We report a patient with uterine myoma (leiomyoma) and erythrocytosis in whom erythropoietin (Epo) production in the leiomyoma tissue was identified by reverse transcription polymerase chain reaction (RT‐PCR) and enzyme‐linked immunosorbent assay (ELISA). A 48‐year‐old Japanese woman with uterine myoma showed marked erythrocytosis (haemoglobin: 20·2 g/dl, haematocrit: 61·1%, red blood cells: 6·51 × 1012/1). After hysterectomy, erythrocytosis rapidly disappeared. In the leiomyoma tissue collected from the patient, Epo mRNA expression was confirmed using RT‐PCR. Furthermore, ELISA showed that the Epo protein level was significantly increased compared with those in control tissues. It is suggested that the pathogenesis of erythrocytosis in patients with uterine myoma involves ectopic Epo production by leiomyoma tissues.

Enhanced expression of thymidylate synthase may be of prognostic importance in advanced cervical cancer.

The enhanced expression of thymidylate synthase (TS) has been associated with a poor prognosis in patients with several types of epithelial tumors. To determine the association between TS expression and the prognosis of patients with advanced cervical cancer after radiation therapy, we immunohistochemically assayed TS levels in paraffin-embedded tissue sections from 66 patients with stage IIIb cervical cancer using a polyclonal antibody to recombinant human TS. In the 30 patients with high TS expression, the cumulative 5- and 8-year survival rates were 36.8% (95% CI: 17.4–56.2) and 31.6% (95% CI: 12.4–50.7), respectively. In contrast, the 36 patients with low TS expression showed a significantly (p < 0.001) better prognosis, with cumulative 5- and 8-year survival rates of 87.2% (95% CI: 75.5–99.0) and 69.2% (95% CI: 50.7–87.7), respectively. These results suggest that TS expression may be useful in determining the prognosis of patients with advanced cervical cancer.

Screening of BRCA1 mutation using immunohistochemical staining with C-terminal and N-terminal antibodies in familial ovarian cancers

We examined the subcellular localization of BRCA1 proteins using immunohistochemical staining with C‐terminal (GLK‐2 antibody) and N‐terminal (Ab‐2 antibody) monoclonal antibodies in 44 familial ovarian cancers. Among these, 24 cases were associated with 13 independent germ‐line mutations of BRCA1, and loss of heterozygosity (LOH) at one or more BRCA1 microsatellite markers was found in all 21 informative tumors tested. With GLK‐2 antibody, cytoplasmic staining was observed in 15 of 16 tumors (93.8%) with mutation in exon 11, and BRCA1 staining was absent in 8 of 8 tumors (100%) with mutation in exons other than exon 11. When immunohistochemical staining was performed with Ab‐2 antibody, both nuclear and cytoplasmic staining were observed in 14 of 16 tumors (87.5%) with mutation in exon 11. Interestingly, nuclear staining was observed in 3 of 3 tumors (100%) with mutation downstream of exon 11, even though no staining was detected in 5 of 5 tumors (100%) with mutation upstream of exon 11. On the other hand, in familial ovarian cancers without BRCA1 mutations, nuclear staining or both nuclear and cytoplasmic staining was observed in 18 of 20 specimens (90%) and 20 of 20 specimens (100%) with GLK‐2 antibody and with Ab‐2 antibody, respectively. These results suggest that an immunohistochemical assay in combination with employing the C‐terminal and the N‐terminal antibodies appears to have potential as a reliable and useful technique for the screening of BRCA1 mutations, at least to predict the status of mutation, upstream or downstream of exon 11.

Mutation of K-RAS protooncogene and loss of heterozygosity on 6q27 in serous and mucinous ovarian carcinomas.

The genetic etiology of serous and mucinous ovarian carcinomas was investigated in 76 affected patients, focusing on the possible loss of heterozygosity (LOH) involving chromosome band 6q27 and K-RAS mutations at codon 12. The incidence of LOH in 6q27 (6q27 LOH) was 41% in 64 informative cases; 53% (20/38) and 23% (6/26) in cases of serous ovarian carcinoma and in those of mucinous ovarian carcinoma, respectively, indicating that the incidence of 6q27 LOH was significantly higher in cases of serous ovarian carcinoma (P < 0.05). The incidence of K-RAS mutations at codon 12 was 23% (15/64); 5% (2/38) and 50% (13/26) in cases of serous ovarian carcinoma and in those of mucinous ovarian carcinoma, respectively, indicating that the incidence of the K-RAS mutations was significantly higher in cases of mucinous ovarian carcinoma (P < 0.0001). Thus, K-RAS mutations at codon 12 and 6q27 LOH were suggested to be involved in the development and/or progression of mucinous ovarian carcinoma and serous ovarian carcinoma, respectively.

Loss of heterozygosity on chromosome 6q27 and p53 mutations in epithelial ovarian cancer

Loss of heterozygosity (LOH) in chromosome region 6q27 and p53 mutations were studied to attempt to clarify the genetic etiology of ovarian cancer, with particular reference to clear cell adenocarcinoma (CCG), which has a poor prognosis. 6q27 LOH in 70 epithelial oyarian cancer patients was examined using four restriction fragment length polymorphism markers located at 6q27; p53 mutations in tumor DNA were detected using polymerase chain reaction singlestrand conformation polymorphism and sequencing. 6q27 LOH was confirmed in 26 of 48 informative cases (54.2%). No differences in the incidence of 6q27 LOH were seen by histologic type; 6q27 LOH was observed in 45% (5/11) of CCCs. p53 mutations were detected in 19 of the 48 tumors (39.6%), but in only one (9%) CCC. These results suggest that a putative tumor suppressor gene involved in the onset of epithelial ovarian cancer is located at 6q27. This gene is one of the keys to clarifying the genetic etiology of epithelial ovarian cancer and particularly CCC, given the low incidence of p53 mutations in this tumor type.