Kohei Nakamura

Letrozole as second-line hormonal treatment for recurrent low-grade endometrial stromal sarcoma: A case report and review of the literature

Low-grade endometrial stromal sarcoma (LGESS) is a rare malignancy. The tumor is reportedly responsive to hormonal therapy, most commonly with medroxyprogesterone acetate (MPA), but the effectiveness of aromatase inhibitors for recurrent LGESS remains unclear. The present study reports a case of stage IC LGESS presenting with abnormal uterine bleeding, and also provides a review of the literature. Following a total abdominal hysterectomy and bilateral salpingo-oophorectomy, MPA therapy was initiated; treatment was successful, but discontinued 19 months later due to disruptive side effects. A further 2 months later, the patient presented with recurrent disease and received chemotherapy. MPA treatment was restarted with a partial response. A second recurrence, 4 years later, presented with lung and para-aortic lymph node metastases. The patient responded to treatment with the aromatase inhibitor letrozole. The patient has since exhibited stable disease and remained free of symptoms for 7 years. This case suggests that aromatase-inhibitor treatment may be effective for recurrent LGESS as a second-line treatment.

Microsatellite instability is a biomarker for immune checkpoint inhibitors in endometrial cancer

In recent years, it has become evident that tumor cells have immune escape mechanisms, and immune checkpoint inhibitor therapy (anti-PD-1/PD-L1 antibody) has shown benefit in various cancers. In endometrial tumors with microsatellite-instability (MSI), somatic mutations have the potential to encode ‘’non-self’’ immunogenic antigens, and lymphocytes have been shown to infiltrate the tumor. Therefore, immune checkpoint inhibitor therapy might be effective in endometrial cancers with MSI. Expression of mismatch repair (MMR) proteins (MLH1, PMS2, MSH2, and MSH6), the presence of tumor-infiltrating lymphocytes (CD8+), and PD-1/PD-L1 expression were assessed by immunohistochemistry in 149 patients with endometrial cancer. We examined whether tumors with MSI had an enhanced immune microenvironment and whether MSI could be a predictor of the therapeutic effect of PD-1/PD-L1 immunotherapy in endometrial cancer. Loss of MMR protein expression was identified in 42 (28.2%) of 149 patients (MSI group) with endometrial cancer. There was no significant relationship between MSI status and age (p = 0.193), histological grade (p = 0.097), FIGO stage (p = 0.508), pelvic lymph node metastasis (p = 0.139), or depth of myometrial invasion (p = 0.494). However, the presence of tumor-infiltrating lymphocytes (CD8+) and PD-L1/PD-1 expression were significantly higher in the MSI group compared to the microsatellite-stable group (p = 0.002, p = 0.001, and p = 0.008, respectively). These results suggest that immune checkpoint inhibitors (anti-PD-1/PD-L1 antibody) could be effective in endometrial cancers with MSI. The presence of MSI may be a biomarker for good response to PD-1/PD-L1 immunotherapy in endometrial cancer.

Loss of autophagy-related protein Beclin 1 may define poor prognosis in ovarian clear cell carcinomas

The aim of the present study was to clarify the role of autophagy in cisplatin (CDDP) sensitivity in OCCCs and the role of Beclin 1 in OCCC progression. Autophagy was measured using: i) western blot analysis of LC3 and p62 and ii) microscopic observation of GFP-LC3 puncta. Autophagy was suppressed using chloroquine and Beclin 1 siRNA. Surgical specimens were examined for Beclin 1 protein expression by immunohistochemistry. The correlations between the loss of Beclin 1 expression and clinicopathological characteristics, prognosis and chemosensitivity were investigated. Inhibition of autophagy by chloroquine or Beclin 1 siRNA did not enhance the sensitivity of the ES2 and TOV-21G OCCC cell lines to CDDP. Loss of Beclin 1 expression was observed in 38.3% (23/60) of the analyzed tumors. There was no significant correlation between loss of Beclin 1 expression and FIGO stage, CA125 levels, patient age, status of endometriosis, Ki-67 labeling index, chemotherapy regimen or status of residual tumor. However, negative expression of Beclin 1 was associated with a shorter progression-free survival in comparison to positive Beclin 1 expression in OCCC who received cytoreductive surgery, followed by a standard platinum-based chemotherapy regimen (P=0.027, log-rank test). Beclin 1-negative tumors were no more resistant to primary adjuvant chemotherapy than were Beclin 1-positive tumors (50.0 vs. 66.7%, P=0.937). Beclin 1 knockdown using siRNA increased cell growth but not cell migration and invasion in ES2 and TOV-21G OCCC cell lines. Autophagy defects caused by loss of Beclin 1 are not related to chemoresistance and metastasis, but may be associated with malignant phenotype and poor prognosis of OCCC.

CCNE1 amplification is associated with aggressive potential in endometrioid endometrial carcinomas

The clinicopathological significance of amplification was investigated of the gene encoding cyclin E (CCNE1) and we assessed whether CCNE1 was a potential target in endometrioid endometrial carcinomas. CCNE1 amplification and CCNE1 or F-box and WD repeat domain-containing 7 (FBXW7) expression in endometrial endometrioid carcinoma was assessed by immunohistochemistry and fluorescence in situ hybridization. CCNE1 knockdown by small interfering RNA (siRNA) was used to assess the CCNE1 function. The results showed that CCNE1 amplification was present in 9 (8.3%) of 108 endometrial carcinomas. CCNE1 amplification was correlated with high histological grade (Grade 3; P=0.0087) and lymphovascular space invasion (P=0.0258). No significant association was observed between CCNE1 amplification and FIGO stage (P=0.851), lymph node metastasis (P=0.078), body mass index (P=0.265), deep myometrial invasion (P=0.256), menopausal status (P=0.289) or patient age (P=0.0817). CCNE1 amplification was significantly correlated with shorter progression-free and overall survival (P=0.0081 and 0.0073, respectively). CCNE1 protein expression or loss of FBXW7 expression in endometrial endometrioid carcinoma tended to be correlated with shorter progression-free and overall survival; however, this difference was not statistically significant. Multivariate analysis showed that CCNE1 amplification was an independent prognostic factor for overall survival but not for progression-free survival (P=0.0454 and 0.2175, respectively). Profound growth inhibition was observed in siRNA-transfected cancer cells with endogenous CCNE1 overexpression compared with that in cancer cells having low CCNE1 expression. CCNE1 amplification was independent of p53, HER2, MLH1 and ARID1A expression but dependent on PTEN expression in endometrial carcinomas. These findings indicated that CCNE1 amplification was critical for the survival of endometrial endometrioid carcinomas. Furthermore, the effects of CCNE1 knockdown were dependent on the CCNE1 expression status, suggesting that CCNE1-targeted therapy may be beneficial for patients with endometrial endometrioid carcinoma having CCNE1 amplification.

KRAS/BRAF Analysis in Ovarian Low-Grade Serous Carcinoma Having Synchronous All Pathological Precursor Regions

Ovarian low-grade serous carcinoma is thought to begin as a serous cystadenoma or adenofibroma that progresses in a slow stepwise fashion. Among the low-grade serous carcinomas, there is a high frequency of activating mutations in the KRAS or BRAF genes; however, it remains unclear as to how these mutations contribute to tumor progression. This is the first report to track the histopathological progression of serous adenofibroma to low-grade serous carcinoma. Each stage was individually analyzed by pathological and molecular genetic methods to determine what differences occur between the distinct stages of progression.

Efficacy of multiple microwave endometrial ablation technique for menorrhagia resulting from adenomyosis.

Conventional microwave endometrial ablation (MEA) can be insufficient to control menorrhagia resulting from adenomyosis. We compared the standard single ablation technique with multiple MEA – repeating ablation three times in the same region – in patients with adenomyosis and menorrhagia.

Affinity-purified DNA-based mutation profiles of endometriosis-related ovarian neoplasms in Japanese patients

Aim Endometriosis-related ovarian neoplasms (ERONs) have recently attracted considerable attention; however, the prevalence and patterns of ARID1A and POLE mutations in ERONs have not been studied in detail. The aim of this study was to investigate not only the carcinogenesis of ERONs, but also the prognostic significance of several gene mutations in this cohort. We used DNA purified from only tumor epithelial cells, from which fibroblasts were removed, using a specific method we called “liquid microdissection”. Methods Tissue samples from 22 ovarian carcinomas (13 endometrioid, and nine clear cell) were used. Tumor cells were isolated using a cell sorting system and DNA was purified from tumor epithelial cells. Nucleotide sequencing was conducted to analyze the mutational status of ARID1A, p53, PTEN, POLE, PIK3CA, and KRAS. Results In ERONs, the frequencies of somatic mutations in ARID1A, p53, POLE, PTEN, PIK3CA, and KRAS were 19/20 (95.0%), 7/19 (36.8%), 9/22 (40.9%), 13/19 (68.4%), 3/19 (15.8%), and 1/9 (11.1%). The frequency of ARID1A mutations was significantly higher than that reported previously. Kaplan-Meier survival analysis revealed that mutations in all genes, including POLE, were not associated with patient prognosis in our Japanese cohort. Conclusions Our results suggest that the frequency of ARID1A mutations in ERONs may be higher than that previously reported. In addition, the “liquid microdissection” method that we chose for DNA purification could be used to obtain high-quality sequencing results. The findings suggest that ARID1A mutations represent the basis of ERON carcinogenesis; other subsequent gene mutations may result in the progression of carcinogenesis.

Reconstitution of high-grade serous ovarian carcinoma from primary fallopian tube secretory epithelial cells

Fallopian tube secretory epithelial cells (FTSECs) have been suggested to be the source of high-grade serous ovarian carcinoma (HGSOC). Although several genetic alterations are known to be involved in HGSOC development, the minimal requirements remain unclear. We aimed to identify oncogenic mutations indispensable for HGSOC development in a stepwise model, using immortalized FTSECs. FTSECs were isolated from clinical samples and immortalized by overexpression of cyclin D1, CDK4R24C, and hTERT. Oncogenic mutations in the p53, c-Myc, and RAS/PI3K pathways were mimicked by lentiviral transduction. We found two distinct patterns of gene alteration essential for HGSOC development: p53/KRAS/AKT and p53/KRAS/c-Myc. Dominant-negative p53, alone or combined with oncogenic KRAS (KRASV12), constitutively active AKT (CA-AKT), and c-Myc, did not induce tumorigenesis in immortalized cells; however, overexpression of CA-AKT or c-Myc, along with dominant-negative p53 and KRASV12, conferred tumorigenic potential. Transformed FTSECs formed tumors in nude mice that were grossly, histologically, and immunohistochemically similar to human HGSOCs. Interestingly, mice harboring tumors with c-Myc amplifications displayed extensive metastases, consistent with the increased dissemination in their human counterparts. Thus, aberrant p53/KRASV12/c-Myc or p53/KRASV12/PI3K-AKT signaling was the minimum requirement for FTSEC carcinogenesis. The model based on this evidence could shed light on the early stages of HGSOC development.

Long‑term outcomes of microwave endometrial ablation for treatment of patients with menorrhagia: A retrospective cohort study

This study aimed to describe the long-term outcomes of patients with menorrhagia treated with microwave endometrial ablation (frequency, 2.45 GHz), as well as to identify factors associated with recurrence or re-surgery. This retrospective cohort study was conducted from 2007 to 2015 at Shimane University Hospital in Japan. Patients with severe menorrhagia and a desire to preserve their uterus were included in the study. Clinical factors associated with recurrence of menorrhagia or re-surgery were analyzed with a multivariable logistic regression model. Of 160 microwave endometrial ablation candidates, 100 had uterine myomas, 20 adenomyosis, 26 functional excessive menstruation, and 12 endometrial polyps. In the full cohort, age (<40) and uterine cavity length (≥10) were associated with recurrence of menorrhagia and re-surgery. Among patients with myomas, age (<48) and number of myomas (≥4) were associated with recurrence, and largest myoma size (≥5) and preoperative hemoglobin level (<9 mg/dl) were associated with re-surgery. Among subjects with adenomyosis, uterine cavity length (≥10) was associated with recurrence. Microwave endometrial ablation is thought to be a highly efficacious method to control menorrhagia caused by functional bleeding and endometrial polyps. However, microwave endometrial ablation may be less effective for patients younger than 48 years with myomas, especially those with 4 or more myomas, or with a myoma 5 cm or larger in size, and for patients with adenomyosis who have a thickened myometrium. These clinical factors may be useful predictors of success in selecting candidates for microwave endometrial ablation.

Nucleus accumbens-1/GADD45GIP1 axis mediates cisplatin resistance through cellular senescence in ovarian cancer

Nucleus accumbens-1 (NAC1), a nuclear factor belonging to the bric-a-brac-tramtrack-broad complex/pox virus and zinc finger gene family, is known to serve important roles in the proliferation and growth of tumor cells, and in chemotherapy resistance. However, the underlying molecular mechanisms through which NAC1 contributes to drug resistance remain unclear. In the present study, the role of NAC1 in drug resistance in ovarian cancer was investigated. NAC1 expression was markedly negatively associated with growth arrest and DNA-damage-inducible 45γ-interacting protein 1 (GADD45GIP1) expression in ovarian cancer. Increased NAC1 expression or decreased GADD45GIP1 expression was significantly associated with decreased progression-free survival (P=0.0041). Multivariate analysis demonstrated that NAC1/GADD45GIP1 expression was an independent prognostic factor of progression-free survival (P=0.0405). It was investigated whether cellular senescence was involved in NAC1-mediated resistance to cisplatin, a commonly used chemotherapeutic drug in the treatment of ovarian cancer. Treatment with cisplatin activated cellular senescence in ovarian cancer cell lines (SKOV3 and TOV-21G cells). Furthermore, knockdown of NAC1 by RNA interference significantly increased GADD45GIP1 expression and inhibited cisplatin-induced cellular senescence, resulting in increased cisplatin cytotoxicity in SKOV3 cells, which express increased levels of NAC1. To investigate whether the sensitizing effect of NAC1 inhibition on cisplatin-induced cytotoxicity may be attributed to the suppression of cellular senescence, the effects of NAC1 overexpression were assessed in TOV-21G cells, which do not express endogenous NAC1. Transfection with NAC1 in TOV-21G cells reduced the sensitivity of TOV-21G cells to cisplatin, indicating that suppression of cellular senescence was induced by GADD45GP1 activation. The results of the present study suggest that NAC1 is a negative regulator of cellular senescence and that NAC1-dependent suppression of senescence, mediated through GADD45GIP1, serves an important role in promoting cisplatin resistance. Therefore, the NAC1/GADD45GIP1 axis may be a potential target for the treatment of ovarian cancer, particularly in platinum-resistant cancers.