Vulvar neuroendocrine carcinoma that is independent of merkel cell polyomavirus and human papillomavirus suggests endometrial cancer recurrence: a case report.

BACKGROUND: Vulvar neuroendocrine carcinomas with small cell morphology need an appropriate differential diagnosis with respect to primary Merkel cell carcinomas, primary small cell neuroendocrine carcinomas, and secondary/metastatic carcinomas. Herein, we report a woman with a history of endometrial carcinoma led to neuroendocrine vulvar carcinoma. CASE PRESENTATION: An 82-y-old woman with right vulvar swelling was transferred to our hospital. Computed tomography scan showed a 75 mm irregular mass in her right vulva. Three years ago, she had been diagnosed with endometrial endometrioid carcinoma stage IA and had undergone surgery. Vulvar biopsy revealed neuroendocrine carcinomas with small cell morphology. Immunohistochemical staining showed that the vulvar tumor was positive for CD56 and chromogranin A, but negative for Merkel cell polyomavirus and cytokeratin 20. Incidentally, her endometrial carcinoma was also positive for CD56 and chromogranin A. Human papillomavirus DNA typing analysis of vulvar tumor was negative. Hence, the vulvar tumor seemed to be a recurrence of the endometrial cancer rather than a primary vulvar neuroendocrine carcinoma. The patient died of the disease within a month. CONCLUSION: We report a case of vulvar neuroendocrine carcinoma that is independent of Merkel cell polyomavirus and human papillomavirus, thereby suggesting a recurrence of endometrial cancer. Immunohistochemical and virological analyses helped in the differential diagnosis of the neuroendocrine carcinoma.

Pembrolizumab monotherapy in Japanese patients with advanced ovarian cancer: Subgroup analysis from the KEYNOTE-100.

Interim results from the two-cohort, phase 2 KEYNOTE-100 study (NCT02674061) of 376 patients with previously treated advanced recurrent ovarian cancer (ROC) showed that pembrolizumab monotherapy was associated with an objective response rate (ORR) of 8.0% (95% CI, 5.4-11.2). We present outcomes for the Japanese patients (n = 21) enrolled in KEYNOTE-100. Patients with epithelial ROC had received either 1-3 prior chemotherapy lines and had platinum-free interval or treatment-free interval (PFI; TFI) of 3-12 months (cohort A) or 4-6 prior chemotherapy lines and had PFI/TFI of ≥3 months (cohort B). All patients received pembrolizumab 200 mg every 3 weeks as monotherapy for 2 years or until progression, death, unacceptable toxicity or consent withdrawal. Primary objectives were ORR per RECIST v1.1 for each cohort and higher programmed death ligand-1 (PD-L1) tumor expression. The relationship between PD-L1 expression (measured as combined positive score [CPS]) and ORR was assessed. Twenty-one Japanese patients (cohort A, n = 19; cohort B, n = 2) were treated. The median (range) age was 57 (37-78) years; 19 (90.5%) patients had ECOG status of 0 and 16 (76.2%) patients had stage III-IV disease. ORR was 19.0% (95% CI, 5.4-41.9) and seemed to increase with increasing PD-L1 expression. A total of 13 (61.9%) patients had treatment-related adverse events (TRAE), and 5 (23.8%) had grade 3-4 TRAE. There were no treatment-related deaths in this subpopulation. Pembrolizumab monotherapy was associated with antitumor activity in Japanese patients with ROC, with no new safety signals identified in this subpopulation. The data suggested a trend toward higher PD-L1 expression among some patients with higher ORR.

hsa-miR-100-5p, an overexpressed miRNA in human ovarian endometriotic stromal cells, promotes invasion through attenuation of SMARCD1 expression.

BACKGROUND: A number of microRNAs are aberrantly expressed in endometriosis and are involved in its pathogenesis. Our previous study demonstrated that has-miR-100-5p expression is enhanced in human endometriotic cyst stromal cells (ECSCs). The present study aimed to elucidate the roles of has-miR-100-5p in the pathogenesis of endometriosis. METHODS: Normal endometrial stromal cells (NESCs) were isolated from normal eutopic endometrium without endometriosis. Using hsa-miR-100-5p-transfected NESCs, we evaluated the effect of hsa-miR-100-5p on the invasiveness of these cells by Transwell invasion assay and in-vitro wound repair assay. We also investigated the downstream signal pathways of hsa-miR-100-5p by microarray analysis and Ingenuity pathways analysis. RESULTS: hsa-miR-100-5p transfection enhanced the invasion and motility of NESCs. After hsa-miR-100-5p transfection, mRNA expression of SWItch/sucrose non-fermentable-related matrix-associated actin-dependent regulator of chromatin subfamily D member 1 (SMARCD1) was significantly attenuated. Whereas, the expression of matrix metallopeptidase 1 (MMP1) mRNA and active MMP1 protein levels was upregulated. CONCLUSION: We found that SMARCD1/MMP-1 is a downstream pathway of hsa-miR-100-5p. hsa-miR-100-5p transfection enhanced the motility of NESCs by inhibiting SMARCD1 expression and MMP1 activation. These findings suggest that enhanced hsa-miR-100-5p expression in endometriosis is involved in promoting the acquisition of endometriosis-specific characteristics during endometriosis development. Our present findings on the roles of hsa-miR-100-5p may thus contribute to understand the epigenetic mechanisms involved in the pathogenesis of endometriosis.

ATM expression is attenuated by promoter hypermethylation in human ovarian endometriotic stromal cells.

A number of genes involved in the pathogenesis of endometriosis are silenced by the hypermethylation of their promoter regions. We assessed the effect and mechanism of the DNA demethylating agent 5-aza-2'-deoxycytidine (5-aza-dC) (10 µM) on the cell cycle in human endometriotic cyst stromal cells (ECSCs) and normal endometrial stromal cells (NESCs) by flow cytometry. The DNA methylation status of G2/M checkpoint regulators were investigated by methylation-specific PCR. The expression of ataxia telangiectasia mutated (ATM) and the effect of 5-aza-dC on its expression were also evaluated by quantitative RT-PCR and western blotting analysis. 5-aza-dC treatment resulted in the cell cycle arrest of ECSCs at the G2/M phase. In contrast, 5-aza-dC did not affect the cell cycle of NESCs. The promoter region of the ATM gene was hypermethylated in ECSCs, but not in NESCs. ATM mRNA expression was attenuated in ECSCs compared to that in NESCs. Further, 5-aza-dC was found to restore ATM expression in ECSCs by its promoter demethylation. Our findings indicate that ATM promoter hypermethylation occurs in endometriosis, and that ATM silencing is involved in tumorigenesis during this disease; moreover, selective DNA demethylating agents and molecular target drugs against ATM silencing are promising for the treatment of endometriosis.

Ruptured external iliac artery pseudoaneurysm following surgery for ovarian cancer: A case report and literature review.

Ruptured pseudoaneurysm following pelvic surgery is a rare and fatal complication. Because of its rarity, existing evidence is limited to a small case series. A 60-year-old woman underwent staging laparotomy, including pelvic and para-aortic lymphadenectomy, for ovarian cancer. On the 11th day, the patient developed a sudden lumbar pain and loss of consciousness, which resulted in a state of shock. She was diagnosed as having a pelvic abscess and ruptured external iliac artery pseudoaneurysm. We performed ligation of the external iliac artery to restrain hemorrhage and femoro-femoral artery bypass to prevent infection, which saved the patient's life. Our study had two findings. First, this is the first reported case of ruptured external iliac artery pseudoaneurysm following surgery for ovarian cancer. Second, treatment of ruptured pseudoaneurysm requires rapid hemostasis, prevention of infection, and revascularization.

Expression of angiogenic factors in sclerosing stromal tumours of the ovary.

Sclerosing stromal tumours (SSTs) are rare benign tumours and are generally observed in individuals in their teens to 20 s. Many cases are suspected as malignant tumours pre-operatively, owing to their high vascularity on diagnostic imaging. Herein, we aimed to evaluate the expressions of various angiogenic factors in SSTs. The expressions of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and hepatocyte growth factor (HGF) were examined by immunohistochemical staining. Upon immunohistochemistry, overexpressions of VEGF, bFGF, and HGF in the ovarian stroma were observed in SSTs. In the normal ovary, the expressions were strong around the vessels and weak in the stroma of the ovary, while a Sertoli-Leydig cell tumour showed weak staining with VEGF, locally strong staining with bFGF and negative staining with HGF. This is the first report about angiogenic factors such as bFGF, and HGF in SST. Impact statement What is already known on this subject? Sclerosing stromal tumour (SST) is a rare benign tumour with high vascularity. The high vascularity of SSTs are reported to be related to the overexpression of vascular endothelial growth factor (VEGF). But there is no study on the expressions of other angiogenic factors in SST. What do the results this study add? In the immunohistochemical analysis, VEGF, bFGF and HGF were found to be widely stained in SSTs. This results suggest the possibility that the high vascularity seen on diagnostic imaging, and the many small vessels observed microscopically may be related to the expressions of VEGF, bFGF and HGF in SSTs. What are the implications of these findings for clinical practice and/or further research? Our results suggest the possibility that the combined expression pattern of VEGF, bFGF, and HGF may be used as marker of SSTs.

Arcyriaflavin a, a cyclin D1-cyclin-dependent kinase4 inhibitor, induces apoptosis and inhibits proliferation of human endometriotic stromal cells: a potential therapeutic agent in endometriosis.

BACKGROUND: We previously showed that microRNA-503 (miR-503) transfection into endometriotic cyst stromal cells (ECSCs) induced cell cycle arrest at the G0/G1 phase by suppressing cyclin D1. This finding prompted us to evaluate the potential therapeutic effects of cyclin D1 inhibitors in endometriotic cells. This study aimed to determine whether arcyriaflavin A, a representative inhibitor of cyclin D1-cyclin-dependent kinase 4 (CDK4), is beneficial in the treatment of endometriosis. METHODS: ECSCs were isolated from the ovarian endometriotic tissues of 32 women. The effects of arcyriaflavin A on cell viability and proliferation, vascular endothelial growth factor A expression, apoptosis, and cell cycle progression were evaluated using a modified methylthiazoletetrazolium assay, enzyme-linked immunosorbent assay (ELISA), Caspase-Glo® 3/7 assay, and flow cytometry. RESULTS: Arcyriaflavin A significantly inhibited cell viability, proliferation, and angiogenesis of ECSCs as assessed using the 5-bromo-2-deoxyuridine (BrdU) and methylthiazoletetrazolium bromide (MTT) assays, and vascular endothelial growth factor (VEGF) ELISA. Arcyriaflavin A induced apoptosis as shown in the Caspase-Glo® 3/7 assay and cell death detection ELISA whilethe cell cycle was arrested at the G0/G1 phase. CONCLUSION: The findings indicate that cyclin D1-CDK4 inhibitors may be promising candidates for the treatment of endometriosis. This is the first study to demonstrate the potential usefulness of arcyriaflavin A as a therapeutic agent for endometriosis. Further studies of the effects of cyclin D1-CDK4 inhibitors on endometriosis may provide useful information on pathogenesis and treatment.

Current status of tertiary debulking surgery and prognosis after secondary debulking surgery for recurrent Müllerian epithelial cancer in Japan: a retrospective analysis of 164 patients (KCOG-G1402).

BACKGROUND: This study aimed to evaluate the current status of secondary debulking surgery (SDS) and tertiary debulking surgery (TDS; performed for recurrence after SDS) and to assess the overall survival after recurrence of Müllerian epithelial cancer in Japan. We also evaluated the data of patients who underwent a fourth debulking surgery (i.e., quaternary debulking surgery (QDS)). METHODS: We conducted a retrospective study of 164 patients with recurrent Müllerian epithelial cancers (i.e., ovarian, tubal, and peritoneal cancers). The SDS was performed between January 2000 and September 2014 in 20 Japanese hospitals. Clinicopathological data were collected and analyzed. RESULTS: Of the 164 patients, 66 patients did not have a recurrence or died after SDS. Ninety-eight patients had a recurrence after SDS. Forty-three of the 98 patients underwent TDS; 55 of the 98 patients did not undergo TDS and were classified into the non-TDS group. The overall survival (OS) after SDS was significantly better in the TDS group than in the non-TDS group. The median OS after SDS was 123 and 42 months in the TDS group and non-TDS group, respectively. Of the 43 patients who received TDS, 11 patients were further treated with QDS. The median OS after SDS was 123 months for patients who underwent QDS. CONCLUSIONS: This multicenter study on the prognosis of post-SDS is apparently the first report on QDS in Japan. Patients undergoing TDS have a good prognosis, compared to patients in the non-TDS group. Novel drugs are being evaluated; however, debulking surgery remains a necessary treatment for recurrence.

miR-503, a microRNA epigenetically repressed in endometriosis, induces apoptosis and cell-cycle arrest and inhibits cell proliferation, angiogenesis, and contractility of human ovarian endometriotic stromal cells.

STUDY QUESTION: Is the micro-RNA (miRNA) miR-503, downregulated in endometriotic cyst stromal cells (ECSCs) and does this affect the cell cycle, cell proliferation, angiogenesis and contractility of these cells? SUMMARY ANSWER: miR-503 expression is downregulated in ECSCs by DNA hypermethylation and this contributes to their proliferation, resistance to apoptosis, extracellular matrix (ECM) contractility and angiogenesis through effects on cyclin D1, B-cell lymphoma/leukemia (Bcl)-2, Ras homology A  and vascular endothelial growth factor A (VEGF-A). WHAT IS KNOWN ALREADY: A variety of miRNAs are demonstrated to involve in the pathogenesis of endometriosis. miR-503 is a miRNA with tumor-suppressor functions, whose expression is suppressed in ECSCs. STUDY DESIGN, SIZE, DURATION: We isolated ECSCs and normal endometrial stromal cells (NESCs) from ovarian endometriotic tissues (n = 32) and eutopic endometrial tissues without endometriosis (n = 8), respectively. PARTICIPANTS/MATERIALS, SETTING, METHODS: We investigated the functions of miR-503 by using miR-503-transfected ECSCs and the DNA methylation status of miR-503 gene in ECSCs and NESCs by combined bisulfite restriction analysis. MAIN RESULTS AND THE ROLE OF CHANCE: In ECSCs, miR-503 is downregulated by the DNA hypermethylation of its gene. The transfection of miR-503 into ECSCs resulted in the inhibition of cell proliferation and induction of cell-cycle arrest at G0/G1 phase through the suppression of cyclin D1, the induction of apoptosis through Bcl-2 suppression, the inhibition of VEGF-A production and the attenuation of ECM contractility via the suppression of Rho/Rho-associated coiled-coil-forming protein kinase-pathways. LARGE SCALE DATA: NA. LIMITATIONS, REASONS FOR CAUTION: The present experiments were carried out only with the stromal component of endometriosis and eutopic endometrium. The experiments with the eutopic endometrial stromal cells from women with endometriosis are not performed. WIDER IMPLICATIONS OF THE FINDINGS: Our findings indicate that epigenetically repressed miR-503 in ECSCs is involved in the acquisition of endometriosis-specific cellular functions. STUDY FUNDING/COMPETING INTERESTS: This work was supported in part by Grants-in-Aid for Scientific Research from the Japan Society for the Promotion of Science (no. 13237327 to K.N., no. 26861335 to K.K. and no. 23592407 to H.N.) and the Kanzawa Medical Research Foundation (to K.K.). There are no conflicts of interest to declare.

Efficacy and safety of olanzapine combined with aprepitant, palonosetron, and dexamethasone for preventing nausea and vomiting induced by cisplatin-based chemotherapy in gynecological cancer: KCOG-G1301 phase II trial.

PURPOSE: Olanzapine is effective in chemotherapy-induced nausea and vomiting (CINV). In patients receiving highly emetogenic chemotherapy (HEC), its efficacy was reported as rescue therapy for breakthrough emesis refractory to triplet therapy (palonosetron, aprepitant, and dexamethasone). However, its preventive effects with triplet therapy for CINV are unknown. This study aimed to investigate efficacy and safety of preventive use of olanzapine with triplet therapy for CINV of HEC. METHODS: This study is a prospective multicenter study conducted by Kansai Clinical Oncology Group. Forty chemo-naïve gynecological cancer patients receiving HEC with cisplatin (≥50 mg/m(2)) were enrolled. Oral olanzapine (5 mg) was administered with triplet therapy a day prior to cisplatin administration and on days 1-5. The primary endpoint was complete response (no vomiting and no rescue) rate for the overall phase (0-120 h post-chemotherapy). Secondary endpoints were complete response rate for acute phase (0-24 h post-chemotherapy) and delayed phase (24-120 h post-chemotherapy) and complete control (no vomiting, no rescue, and no significant nausea) rate and total control (no vomiting, no rescue, and no nausea) rate for each phase. These endpoints were evaluated during the first cycle of chemotherapy. RESULTS: Complete response rates for acute, delayed, and overall phases were 97.5, 95.0, and 92.5 %, respectively. Complete control rates were 92.5, 87.5, and 82.5 %, respectively. Total control rates were 87.5, 67.5, and 67.5 %, respectively. There were no grade 3 or 4 adverse events. CONCLUSIONS: Preventive use of olanzapine combined with triplet therapy gives better results than those from previously reported studies of triplet therapy.

Wisteria floribunda agglutinin-binding glycan expression is decreased in endometriomata.

BACKGROUND: Glycosylation is one of the most common post-translational modifications of eukaryotic proteins and is known to undergo dynamic changes in a wide range of biological processes. To date, however, the glycan expression profiles in endometriosis are largely unknown. The objective of the study was to identify the panel of glycans that were aberrantly expressed in endometriosis, a hormone-dependent disease. METHODS: The glycan expression profiles in primary cultured human endometriotic cyst stromal cells (ECSCs) and normal endometrial stromal cells (NESCs) were determined by lectin microarray analysis. Distribution of Wisteria floribunda agglutinin (WFA)-binding glycans in ovarian endometriotic cysts and eutopic proliferative phase endometrium were assessed by lectin histochemistry. The expressions of N-acetylgalactosaminyl transferases that synthesize WFA-binding glycans were evaluated in ECSCs and NESCs. RESULTS: We found that the levels of WFA-binding glycans were decreased in ECSCs. Lectin histochemistry revealed that WFA-binding glycans were decreased only in the stromal components of the ovarian endometriotic cysts, but not in the epithelial components, compared to the eutopic proliferative phase endometrium. The expressions of N-acetylgalactosaminyl transferases that synthesize WFA-binding glycans were downregulated in ECSCs. CONCLUSIONS: Utilizing lectin microarray analysis and lectin histochemistry, we found that WFA-binding glycans were decreased in endometriosis. The synthetic enzymes of WFA-binding glycans were significantly downregulated in ECSCs. It is suggested that reduced expression of N-glycans with WFA-binding properties on ECSCs is a novel characteristics of endometriosis.

Retrospective analysis of outcomes of secondary debulking surgery for recurrent epithelial ovarian cancer with favorable prognostic factors.

AIM: Ovarian cancer is the second most common gynecological malignancy, yet it has the highest case-fatality ratio of all gynecologic malignancies. Surgery followed by combination platinum-taxane chemotherapy is the standard approach to the management of primary epithelial ovarian cancer. However, standard treatment of patients with recurrent ovarian cancer remains poorly defined. Secondary cytoreductive surgery (SDS) at the time of relapse has been proposed as a means of improving the prognosis of recurrent ovarian cancer patients with a treatment-free interval of at least 6 months. METHODS: In the present study, we retrospectively collected 16 patients with recurrent epithelial ovarian cancer who might benefit most from SDS and evaluated the impact of SDS on the outcomes for this highly select patient group. RESULTS: We found that SDS led to excellent outcomes, with a 73.1% 8-year overall survival rate after initial treatment, a 67.9% 5-year overall survival rate after prior SDS, and a 31.3% 5-year progression-free survival rate after prior SDS. Although the findings were not significant, these results suggest that repeated SDS might improve outcomes for this patient group. CONCLUSION: The present study may provide a platform for discussion of the impact of aggressive or repeated SDS on the survival of patients with recurrent epithelial ovarian cancer and favorable prognostic factors. Further multi-institutional studies with larger number of patients are mandatory to confirm the present findings.

Evaluation of mucosal healing of ulcerative colitis by a quantitative fecal immunochemical test.

OBJECTIVES: Accumulating evidence has underlined the importance of mucosal healing as a treatment goal for ulcerative colitis (UC). Quantitative fecal immunochemical tests (FITs), which can rapidly quantify fecal blood with automated equipment, have been used recently to screen for colorectal neoplasia. The aim of this study is to determine whether an FIT can evaluate mucosal healing in UC. METHODS: Feces collected from UC patients who underwent colonoscopy were examined by FITs, and results were compared with colonoscopic findings. Mucosal status was assessed using the Mayo endoscopic subscore classification. Maximum score for the colorectum in each patient was recorded. RESULTS: Evaluated were FIT results in conjunction with 310 colonoscopies that were performed in 152 UC patients. A large majority of patients with a Mayo 0 endoscopic score had negative FIT (<100 ng/ml) results (92%), and the proportion of negative FIT results decreased with increases in the Mayo score (Mayo 1: 47%, Mayo 2: 13%, Mayo 3: 12%, P<0.0001, Cochran-Armitage trend test). When the negative FIT was defined as <100 ng/ml, the sensitivity and specificity of a negative FIT for mucosal healing (Mayo 0) were 0.92 and 0.71, respectively. When mucosal healing was defined as Mayo 0 or 1, those were 0.60 and 0.87, respectively. In addition, a positive FIT (≥100 ng/ml) predicted mucosal inflammation (Mayo 2 or 3) with sensitivity 0.87 and specificity 0.60, respectively. CONCLUSIONS: The FIT can effectively and noninvasively evaluate mucosal healing in UC. This easy, rapid method can help evaluate and control disease activity of UC.

Colorectal endoscopic submucosal dissection for elderly patients at least 80 years of age.

BACKGROUND: Endoscopic submucosal dissection (ESD) has been used recently for successful en bloc resection of even large lesions, although no consensus appears in medical literature concerning its application to elderly patients. This prospective cohort study aimed to evaluate the efficacy and safety of colorectal ESD for patients 80 years of age or older. METHODS: Colorectal ESD procedure findings were compared with clinical outcomes, including associated complications and mortalities, for two age groups totaling 196 consecutive patients with 202 colorectal lesions. Of the 196 patients, 31 patients (16%) were 80 years of age or older (group E), and 165 patients (84%) were younger than 80 years (group Y). RESULTS: The median ages were 82 years in group E and 68 years in group Y. The frequency of chronic concomitant diseases was significantly higher in group E (65%) than in group Y (27%) (p = 0.003). No significant pressure decrease or need for oxygenation was observed in either group. In addition, groups E and Y did not differ significantly in terms of mean lesion sizes (40.9 vs. 39.7 mm) en bloc resection rates (84% vs. 93%), curative rates (78% vs. 84%), median procedure times (65 vs. 70 min), or associated complications (no perforation or delayed bleeding cases [0%] vs. 5 perforations [3%]) The median postprocedure hospitalization period was 3 days in both groups. Except for 10 cases requiring subsequent lymph node dissection surgery, follow-up colonoscopy examinations showed no recurrences or ESD-related mortalities in either group. CONCLUSION: Colorectal ESD is a safe and effective treatment for elderly patients (age ≥ 80 years) despite a significantly higher frequency of chronic concomitant diseases than among younger patients.

Cadmium chloride induces the expression of metallothionein mRNA by endometrial stromal cells and amnion-derived (WISH) cells.

BACKGROUND: Metallothionein (MT) is known to bind to metals with high affinity. The potential for MT-1 mRNA expression in endometrial stromal cells (ESC) and amniotic cells in response to cytokines and cadmium chloride (CdCl(2)) was evaluated. METHODS: Human ESC were cultured and treated with interleukin-1α, 12-O-tetradecanoylphobol 13-acetate (TPA), forskolin, transforming growth factor-ß, and CdCl(2). Amnion-derived (WISH) cells were also cultured and treated with the same reagents. The levels of MT mRNA were evaluated by Northern blot analysis in ESC and WISH cells. RESULTS: In response to treatment with CdCl(2) (0.01-10 µM), the expression of MT mRNA markedly increased in ESC and WISH cells in a dose-dependent manner. On the other hand, the expression of MT mRNA did not increase after treatment with interleukin-1α (1 nM), TPA (10 nM), forskolin (1 µM) or transforming growth factor-ß (1 nM) in these cells. CONCLUSION: These findings indicate that MT expression in ESC and WISH cells is sensitive to the CdCl(2) concentration, which is known to be evaluated in cigarette smokers. The present results suggest that increased levels of MT may affect metal metabolism at the feto-maternal interface.

hsa-miR-199a-3p Inhibits Motility, Invasiveness, and Contractility of Ovarian Endometriotic Stromal Cells.

It is suggested that aberrantly expressed microRNAs are involved in the pathogenesis of endometriosis. Our previous study demonstrated that expression of the microRNA hsa-miR-199a-3p is attenuated in human endometriotic cyst stromal cells (ECSCs). The current study aimed to define the roles of hsa-miR-199a-3p in the development of endometriosis. ECSCs and normal endometrial stromal cells (NESCs) were isolated from ovarian endometrioma and normal endometrial tissues, respectively. We evaluated the effect of transfected hsa-miR-199a-3p on the migration, invasion, and contractility of ECSCs using Transwell migration assays, in vitro wound healing assays, Transwell invasion assays, and collagen gel contraction assays. We also examined the downstream target of hsa-miR-199a-3p with an online public database search and luciferase reporter assay. Expression of hsa-miR-199a-3p in ECSCs was significantly lower than that in NESCs, whereas the expression of p21-activated kinase 4 (PAK4) mRNA was significantly higher. Transfection of hsa-miR-199a-3p inhibited the migration, invasion, and contractility of ECSCs via inhibition of PAK4 mRNA expression. PAK4 was confirmed to be the direct target of hsa-miR-199a-3p. Transfection of PAK4 small interfering RNA and the PAK4 inhibitor PF-3758309 also inhibited ECSC migration, invasion, and contractility. These findings suggest that hsa-miR-199a-3p may act as a tumor suppressor in endometriosis development. Attenuation of hsa-miR-199a-3p expression was favorable for ECSCs to acquire the highly invasive, motile, and contractile characteristics of endometriotic cells and is involved in the development of endometriosis. Accordingly, PAK4 inhibitors may be promising for the treatment of endometriosis.

A Case of Vaginal Malignant Melanoma Completely Resected by Radical Surgery.

BACKGROUND Primary vaginal malignant melanoma is a rare and aggressive tumor with a high risk of local recurrence and distant metastasis. Although there are several available treatment options, none are considered as standard. Surgical resection is the first treatment choice because of its superior survival benefits. CASE REPORT The patient was a 56-year-old woman with a vaginal mass. At the first visit to our institution, a 20×20 mm black and flat lesion on the lower third of the posterior vaginal wall and a polypoid mass near the vaginal fornix were detected by gynecologic examination. Study of the tumor on the posterior vaginal wall suggested that it did not extend to the uterine cervix. The preoperative diagnosis was vaginal malignant melanoma FIGO stage I (cT1, cN0, cM0). The patient underwent a total vaginectomy, pelvic and inguinal lymphadenectomy, modified radical hysterectomy, and bilateral salpingo-oophorectomy. The tumor cells were arranged in sheets and nests and exhibited nuclear pleomorphism, eosinophilic cytoplasm, brisk mitotic activity, and melanin production. The overlying mucosa was ulcerated. The tumor thickness was 2.5 mm and no residual lesion was found at the surgical margin. No adjuvant therapies were performed. The patient is alive without recurrence 15 months after the initial treatment. CONCLUSIONS This is a case of vaginal malignant melanoma for which complete response was achieved by radical tumor resection, without severe adverse effects and with no observed recurrence 15 months after the surgery.

Chemokine expression profiles of ovarian endometriotic stromal cells in three-dimensional culture.

Endometriosis is a chronic inflammatory disease which is associated with aberrant chemokine expression. We have established a three-dimensional (3D) floating collagen gel culture of human endometriotic cyst stromal cells (ECSCs) as an in vitro model of early-stage fibrosis formation in endometriosis. We evaluated the gene expression profiles of 3D-cultured ECSCs using a gene expression microarray. We identified and confirmed with reverse transcription-polymerase chain reaction that mRNA levels of CXCL1, CXCL2, CXCL3, CXCL8, and CCL20 in 3D-cultured ECSCs were significantly higher than in 2D-cultured ECSCs. The protein levels of CXCL1, CXCL2, CXCL8, and CCL20 in the supernatant of 3D-cultured ECSCs were significantly higher than in 2D-cultured ECSCs. It has been suggested that the 3D-culture model of ECSCs is more suitable for in vitro endometriosis research than 2D-culture. This microarray data provides a new platform to identify the candidate genes involved in the pathogenesis of endometriosis which could be masked in conventional 2D-culture.

ß-catenin signaling inhibitors ICG-001 and C-82 improve fibrosis in preclinical models of endometriosis.

Endometriosis exhibits unique characteristics, such as fibrosis, resistance to apoptosis, and promotion of cell proliferation; however, its pathophysiology is not fully understood. Recurrence rates after treatment are high, and the progression risk continues until menopause; hence, more effective therapy for endometriosis is needed. CREB-binding protein (CBP)/ß-catenin signaling inhibitors have demonstrated antifibrogenetic effects in liver, lung, and skin diseases. The present study evaluated the effects of two CBP/ß-catenin signaling inhibitors, ICG-001 and C-82, on the progression of endometriosis using endometriotic cyst stromal cells from the ovary and normal endometrial stromal cells from the uterus. ICG-001 was also evaluated in a mouse model. ICG-001 and C-82 inhibited cell proliferation, fibrogenesis, and cell migration, and promoted apoptosis in vitro. ICG-001 inhibited the growth of endometriotic lesions in the mouse model. CBP/ß-catenin signaling plays an important role in the pathophysiology of endometriosis. Inhibiting the CBP/ß-catenin signal can be a therapeutic target for endometriosis.

Decidualization Differentially Regulates microRNA Expression in Eutopic and Ectopic Endometrial Stromal Cells.

Decidualization of the endometrium and endometriosis involves the morphological and biochemical reprogramming of the estrogen-primed proliferative stromal compartment under the continuing influence of progesterone. Here, we evaluated the involvement of microRNA in the decidualization processes of normal endometrial stromal cells (NESCs) and endometriotic cyst stromal cells (ECSCs). In vitro decidualization of NESCs and ECSCs was induced by long-term culture with a combination of 0.5 mmol/L of dibutyryl cyclic adenosine monophosphate and 100 nmol/L of dienogest. We investigated the effect of in vitro decidualization on the microRNA and messenger RNA (mRNA) expression profiles of the NESCs and ECSCs using global microarray techniques and an Ingenuity Pathways Analysis. Decidualization differentially enhanced the miR-30a-5p expression in the NESCs and the miR-210 expression in the ECSCs. The enhanced miR-30a-5p expression in the NESCs correlated with the increased mRNA expression of Krüppel-like factor 9 and period circadian clock 3 as well as the decreased mRNA expression of tolloid-like 1, tolloid-like 2, and paired-like homeodomain 1. The enhanced expression of miR-210 in the ECSCs correlated with the decreased mRNA expression of growth hormone receptor and thymidine kinase 1. Although there is no direct evidence, we speculate that the loss of miR-30a-5p-mediated mechanisms of decidualization and the acquisition of miR-210-mediated mechanisms of decidualization may be involved in the progesterone resistance in endometriosis. Further investigations are necessary to test this speculation.