Yasunori Hashiguchi

Identification of Overexpression and Amplification of ABCF2 in Clear Cell Ovarian Adenocarcinomas by cDNA Microarray Analyses

Purpose: Patients with ovarian clear cell adenocarcinoma generally have a poor response to combination chemotherapy and have overall poorer prognosis than patients with other histologic types of ovarian cancer. Genetic changes in this group of cancer have not been thoroughly explored. Identification of these changes may provide us new therapeutic targets to treat this disease. Experimental Design: Genomic and expression array analyses were applied on 30 clear cell ovarian cancer cases and 19 serous cases using a 10,816-element cDNA microarray platform. Further validation and clinical correlation studies were done on differentially expressed genes that are related to chemoresistance. Results: Based on array analyses, 12 genes showed a significant increase in DNA and mRNA copy number and 5 genes showed a significant decrease in DNA and RNA copy number in clear cell tumors compared with those in the serous type. One of the genes was ABCF2, which belongs to the ATP-binding cassette gene superfamily and has been shown to amplify in other tumor types. Validation studies were done using real-time quantitative PCR and immunohistochemistry. The results showed significantly higher ABCF2 DNA and mRNA copy number and protein levels in clear cell cases compared with those in serous cases. Furthermore, in 20 clear cell cases with chemoresponse data available, ABCF2 cytoplasmic staining was significantly higher in nonresponders than that in the responders (60.0% versus 28.5%; P = 0.0002). Conclusions: These data suggest that ABCF2 protein may be a prognostic marker for ovarian clear cell ovarian adenocarcinoma.

Cyclin E Amplification and Overexpression in Clear Cell Adenocarcinoma of the Ovary

Objective: The purpose of this study is to compare DNA, mRNA and protein levels of the cyclin E between clear cell (CC) and serous (SC) ovarian carcinomas, and evaluate the relationship between cyclin E and p53 status. Method: We examined the DNA, mRNA and protein levels of cyclin E and the protein level of p53 in 44 CCs and 39 SCs using microdissected tissues. Results: Relative cyclin E mRNA expression was significantly higher in CC (3.62, 95% CI, 2.24–4.99) than in SC (1.75, 95% CI, 1.05–2.45; p = 0.0098). The percentage of positive nuclear staining of cyclin E was significantly higher in CC (48.3, 95% CI, 40.4–56.1) than SC (25.3, 95% CI, 17.4–33.3; p = 0.0001). The mRNA and protein expression of cyclin E was significantly correlated (r = 0.66, p < 0.0001). However, the correlation between relative DNA copy number and relative mRNA expression was not significant (r = –0.063; p = 0.66). Percentage of positive nuclear staining of cyclin E was significantly higher in p53 positive cases (51.8, 95% CI, 40.0–63.5) than p53 negative cases (36.2, 95% CI, 28.2–44.2; p = 0.028). Conclusions: Cyclin E expression is significantly higher in CC than in SC. Cyclin E expression is significantly related with p53 positivity.

Alteration of G2 Cell Cycle Regulators Occurs during Carcinogenesis of the Endometrium

Objectives: In this study, we examined the alteration of the G2 pathway in endometrial hyperplasia (EH) and endometrioid-type endometrial cancer (EC), and analyzed the relationship between the G2 pathway status and the p53 pathway status. Methods: A total of 103 cases (proliferative phase of the endometrium: 20, EH: 22, and endometrioid-type EC: 61 (I: 39, II: 5, III: 15, recurrence: 2)) were included in this study. We examined the ATM, chk2, CDC25C, cdc2, and cyclin B1 protein expression by immunohistochemistry. In 55 cases (EH: 15; EC: 40), we analyzed chk2 mutations by RT-PCR-SSCP. Results: There were no chk2 mutations in endometrial disease. Elevated or reduced expression rates of ATM, chk2, CDC25C, cdc2 and cyclin B1 were 4.5% (1/22), 0%, 0%, 0% and 4.5% (1/22) in EH and 3.3% (2/61), 4.9% (3/61), 13.1% (8/61), 9.8% (6/61) and 9.8% (6/61) in EC. Alteration of the G2 pathway was higher in EC (32.8%; 20/61) than in EH (9.1%; 2/22; p = 0.047). The G2 pathway was significantly higher in the altered p53 pathway group (48.4%; 15/31) than in the normal p53 pathway group (16.7%; 5/30) in EC (p = 0.0134). The altered p53 pathway tended to be related with the cdc2/cyclin B1 status (p = 0.0529). Conclusions: Alteration of the G2 pathway is thought to occur during carcinogenesis of the endometrium.

Comparison of osteopontin expression in endometrioid endometrial cancer and ovarian endometrioid cancer

This study compared the DNA, RNA, and protein levels of osteopontin (OPN) in endometrioid endometrial cancer (EEC) and ovarian endometrioid cancer (OEC). In total, 63 cancer cases (EEC: 33, OEC: 30) were included. Of these, 47 (EEC: 26, OEC: 21) were examined by quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and 48 (EEC: 25, OEC: 23) were examined by quantitative PCR. OPN expression was detected in 15 (50.0%) of 30 EECs and in 14 (50.0%) of 28 OECs. There was no significant difference in the percentage of positive cytoplasmic OPN staining between the EECs and OECs (12.8 vs 10.4; p=0.6811). The correlation between relative mRNA and protein expression levels was significant in both the EECs and OECs; however, the correlation between relative DNA and mRNA levels was not significant. There was no significant difference in OPN expression between the EECs and OECs.

Malignant rhabdoid tumour of the uterine cervix

A 36 year old nulliparous woman presented with abnormal vaginal bleeding for the previous month. She had a 7cm solid mass in the uterine cervix. The tumour was ®rm, oval and pigmented. Computed tomography and magnetic resonance imaging showed a solid mass and no evidence of metastasis. A cervical smear showed polygonal malignant cells with pigmentation (Fig. 1). The presumptive diagnosis was malignant melanoma. She underwent a total abdominal hysterectomy, but no evidence of metastatic disease was found. Following surgery, she had post-operative chemotherapy consisting of carboplatin, etoposide and ifosfamide: etoposide 100mg/m per day for three days at weeks 0, 3, 9, 12, 18, and 21. carboplatin 400mg/m at weeks 0, 3, 9, 12, 18, and 21, and ifosfamide 1.5g/m per day for ®ve days at weeks 6, 15, and 24. The woman was well and without disease 38 months after her operation. The tumour was ®rm, oval and pigmented and arose from posterior lip of cervix (Fig. 2). The majority of the tumour was composed of sheets of round or polygonal cells containing single, spherical eosinophilic cytoplasmic inclusions that sometimes compressed the nucleus. The nucleus was prominent and vesicular (Fig. 3). Masson-Fontana and haemosiderin staining revealed the pigment as haemosiderin (Fig. 4). Immunohistochemistry demonstrated that the eosinophilic cytoplasmic inclusions reacted strongly with anti-vimentin and were negative for cytokeratin, desmin, myoglobin, CD34, EMA, S100-protein and HMB45. Electron microscopy showed intermediate ®laments in the cytoplasm; however, typical paranuclear aggregates of intermediate ®laments arranged in concentric whorls were not present (Fig. 5). No junctional complexes, basal lamina, microvilli, melanosomes, premelanosomes or myo®lament arrays were identi®ed. The ®ndings were consistent in a malignant rhabdoid tumour.

Chemotherapy-induced neutropenia and febrile neutropenia in patients with gynecologic malignancy

Chemotherapy-induced neutropenia is a common complication in cancer treatment. In this study, we investigated chemotherapy-induced neutropenia that was recently detected in all patients with gynecologic malignancy. Between January 2009 and December 2011, we examined cases of chemotherapy-induced neutropenia reported in our hospital. We analyzed the incidence and clinical features of chemotherapy-induced neutropenia and febrile neutropenia in patients with gynecologic malignancy. During the study period, we administered over 1614 infusions (29 regimens) to 291 patients. The median age of the patients was 60 years (range 24–84 years). Chemotherapy-induced neutropenia occurred in 147 (50.5%) patients over 378 (23.4%) chemotherapy cycles. Febrile neutropenia occurred in 20 (6.9%) patients over 25 (1.5%) cycles. The mean duration of neutropenia and fever was 3.6 days (range 1–12 days) and 3.4 days (range 1–9 days), respectively. The source of fever was unexplained by examination or cultures in 14 (56.0%) cycles. There were two cases of neutropenia-related death. Chemotherapy-induced neutropenia was associated with older age (over 70 years) (P<0.0001), less than five previous chemotherapy cycles (P=0.02), disseminated disease (P=0.03), platinum-based regimens (P<0.0001), taxane-containing regimens (P<0.0001), and combined therapy (P<0.0001). Febrile neutropenia was associated with poor performance status (P<0.0001), no previous chemotherapy (P<0.05), disseminated disease (P<0.0001), and distant metastatic disease (P=0.03). Neither chemotherapy-induced neutropenia nor febrile neutropenia was associated with bone marrow metastases or previous radiotherapy. By identifying risk factors for febrile neutropenia, such as performance status, no previous chemotherapy, disseminated disease, and distant metastatic disease, the safe management of chemotherapy-induced neutropenia may be possible in patients with gynecologic malignancy.

Minimal deviation mucinous adenocarcinoma of the uterine cervix that proved difficult to differentiate from endometrial cancer: A case report

Minimal deviation adenocarcinoma (MDA), also known as adenoma malignum of the uterine cervix, accounts for only ~1% of uterine cervical adenocarcinomas. Adenoma malignum of the uterine cervix was initially described by Gusserow in 1870. Using magnetic resonance imaging (MRI), MDA appears as multilocular lesions with solid components that extend from the endocervical glands to the deep cervical stroma. Cytological evaluation and biopsies have low detection rates, therefore, it is difficult to diagnose MDA accurately prior to treatment. The current study describes a rare case of MDA that was difficult to differentiate from endometrial adenocarcinoma of the corpus uteri preoperatively, as the endometrial biopsy results suggested a well-differentiated endometrioid adenocarcinoma and MRI did not show typical images for MDA. A total abdominal hysterectomy with bilateral salpingo-oophorectomy was performed under the diagnosis of endometrial cancer, and the mass was subsequently diagnosed as MDA of the uterine cervix by pathological examination of the hysterectomy specimen. Postoperatively, although two types of adjuvant chemotherapy were performed, the remaining tumor continued to grow, causing obstruction of the bilateral ureters and leading to bilateral hydronephrosis. The patient is currently alive with the disease 10 months following the surgery.

A retrospective clinical analysis of 5 cases of vaginal melanoma

Vaginal melanoma is a rare tumor, accounting for <1% of all melanomas and ~1-5% of all vaginal malignant tumors. The prognosis of vaginal melanoma is extremely poor, as it is often resistant to chemotherapy and radiotherapy, and metastases may develop in the early stages of the disease. The present study investigated 5 patients with vaginal melanoma treated at the Department of Gynecology of Osaka City University Hospital (Osaka, Japan) between October, 2000 and April, 2014. All the cases presented with abnormal genital bleeding as the main complaint. Notably, in 3 of the 5 cases the tumors appeared as non-pigmented polyps. Local resection was performed as the primary treatment in all 5 cases. After surgery, dermal injection of interferon β (feron maintenance therapy) was performed in 3 cases, and dacarbazine, nimustine, vincristine and interferon β (DAVFeron therapy) was administered in 1 case as adjuvant therapy. All 5 cases recurred within 1 year. The site of recurrence varied, and included the vaginal wall, liver, brain and lung. The median overall survival was 419 days and the median progression-free survival 177 days. In this cohort, all the cases presented with abnormal genital bleeding as the main complaint. Therefore, malignant melanoma of the vagina must be considered along with other gynecological malignancies in patients with abnormal genital bleeding. In this study, over half of the cases had a non-pigmented polypoid lesion of the vagina. Therefore, malignant melanoma of the vagina must be considered when a polypoid lesion is identified on the vaginal wall.

Comparison of cyclin expression between endometrioid-type endometrial cancer (EEC) and endometrioid ovarian cancer (EOC)

5013 Background: The biological behavior of EEC and EOC differs despite their histologic similarity. While EEC is usually diagnosed at an early stage and has a good prognosis, EOC is generally advanced at diagnosis and shows a poor prognosis. Cyclins are reported to regulate the cell cycle and influence the prognosis of several cancers. We compared the pattern of cyclin expression between EEC and EOC, and evaluated the relationship among cyclin expression, estrogen receptor (ER) status, and the Ki67 labeling index (Ki67-LI). METHODS There were 36 EECs and 37 EOCs. Immunohistochemistry (IHC) was used to detect cyclin A, B1, D1, E and F (CNA, CNB1, CND1, CNE, CNF), Ki67, and ER. The percentage of positive nuclear staining for cyclins was calculated. Quantitative PCR was performed for CNA, CND1 and CNE. RESULTS (1) When EEC and EOC were compared, CNA expression was higher in EEC. In contrast, CND1 and CNE expression were higher in EOC. The Ki67-LI was also higher in EOC than EEC. Conversely, ER positivity was higher in EEC than EOC (80.6% vs 35.1%, p=0.0007). (2) EEC: Only CNA expression was significantly correlated with the Ki67-LI (r=0.55, p=0.0004). ER status was not related to cyclin expression. Protein and mRNA levels were significantly correlated for CNA and CNE. (3) EOC: Expression of CNA and CND1 was weakly correlated with the Ki67-LI (r=0.54, p=0.001; r=0.38, p=0.04). CNA expression was higher in ER-negative tumors than ER-positive tumors (23.7 vs 9.6, p=0.027). CND1 expression was also higher in ER-negative than ER-positive tumors (12.1 vs 0, p=0.065). CND1, and CNE protein and mRNA levels were significantly correlated. CONCLUSIONS The pattern of cyclin expression differed between EEC and EOC. [Figure: see text] No significant financial relationships to disclose.

Age‑related differences in prognosis and prognostic factors among patients with epithelial ovarian cancer

Approximately 40% of all patients with ovarian cancer in Japan are aged ≥65 years. The aim of the present study was to evaluate the differences in prognosis and prognostic factors between elderly and younger patients with epithelial ovarian cancer. A total of 114 patients with International Federation of Gynecology and Obstetrics (FIGO) stage I-IV ovarian cancer who were initiated on primary treatment at the Osaka City University Hospital (Osaka, Japan) were included in this study. Patient characteristics, treatment outcome and prognosis were compared between elderly (aged ≥65 years) and younger patients, and the prognostic factors associated with overall survival were evaluated by univariate and multivariate analyses. The most common histological type in younger patients was clear cell carcinoma (33.8%) vs. serous carcinoma in elderly patients (44.1%), with a significant difference in the distribution of histological type (P=0.006). Complete resection was achieved in 56.2% of younger patients compared with 32.4% of elderly patients (P=0.03). The rates of standard primary treatment were comparable (56.7% of younger vs. 50.0% of elderly patients). Overall and disease-free survival did not differ significantly between the two groups. Multivariate analyses identified FIGO stage and standard primary therapy as prognostic factors in younger patients and performance status in elderly patients. Age was not an independent significant prognostic factor among patients with ovarian cancer. Therefore, performance status, rather than age, should be considered when selecting the optimal treatment for elderly patients based on objective assessment.