Pharmacological inhibition of the LIF/LIFR autocrine loop reveals vulnerability of ovarian cancer cells to ferroptosis.

Of all gynecologic cancers, epithelial-ovarian cancer (OCa) stands out with the highest mortality rates. Despite all efforts, 90% of individuals who receive standard surgical and cytotoxic therapy experience disease recurrence. The precise mechanism by which leukemia inhibitory factor (LIF) and its receptor (LIFR) contribute to the progression of OCa remains unknown. Analysis of cancer databases revealed that elevated expression of LIF or LIFR was associated with poor progression-free survival of OCa patients and a predictor of poor response to chemotherapy. Using multiple primary and established OCa cell lines or tissues that represent five subtypes of epithelial-OCa, we demonstrated that LIF/LIFR autocrine signaling is active in OCa. Moreover, treatment with LIFR inhibitor, EC359 significantly reduced OCa cell viability and cell survival with an IC50 ranging from 5-50 nM. Furthermore, EC359 diminished the stemness of OCa cells. Mechanistic studies using RNA-seq and rescue experiments unveiled that EC359 primarily induced ferroptosis by suppressing the glutathione antioxidant defense system. Using multiple in vitro, ex vivo and in vivo models including cell-based xenografts, patient-derived explants, organoids, and xenograft tumors, we demonstrated that EC359 dramatically reduced the growth and progression of OCa. Additionally, EC359 therapy considerably improved tumor immunogenicity by robust CD45+ leukocyte tumor infiltration and polarizing tumor-associated macrophages (TAMs) toward M1 phenotype while showing no impact on normal T-, B-, and other immune cells. Collectively, our findings indicate that the LIF/LIFR autocrine loop plays an essential role in OCa progression and that EC359 could be a promising therapeutic agent for OCa.

Novel LIPA-Targeted Therapy for Treating Ovarian Cancer.

Ovarian cancer (OCa) is the most lethal form of gynecologic cancer, and the tumor heterogeneities at the molecular, cellular, and tissue levels fuel tumor resistance to standard therapies and pose a substantial clinical challenge. Here, we tested the hypothesis that the heightened basal endoplasmic reticulum stress (ERS) observed in OCa represents an exploitable vulnerability and may overcome tumor heterogeneity. Our recent studies identified LIPA as a novel target to induce ERS in cancer cells using the small molecule ERX-41. However, the role of LIPA and theutility of ERX-41 to treat OCa remain unknown. Expression analysis using the TNMplot web tool, TCGA data sets, and immunohistochemistry analysis using a tumor tissue array showed that LIPA is highly expressed in OCa tissues, compared to normal tissues. ERX-41 treatment significantly reduced the cell viability and colony formation ability and promoted the apoptosis of OCa cells. Mechanistic studies revealed a robust and consistent induction of ERS markers, including CHOP, elF2α, PERK, and ATF4, upon ERX-41 treatment. In xenograft and PDX studies, ERX-41 treatment resulted in a significant reduction in tumor growth. Collectively, our results suggest that ERX-41 is a novel therapeutic agent that targets the LIPA with a unique mechanism of ERS induction, which could be exploited to treat heterogeneity in OCa.

PELP1 inhibition by SMIP34 reduces endometrial cancer progression via attenuation of ribosomal biogenesis.

Endometrial carcinoma (ECa) is the fourth most common cancer among women. The oncogene PELP1 is frequently overexpressed in a variety of cancers, including ECa. We recently generated SMIP34, a small-molecule inhibitor of PELP1 that suppresses PELP1 oncogenic signaling. In this study, we assessed the effectiveness of SMIP34 in treating ECa. Treatment of established and primary patient-derived ECa cells with SMIP34 resulted in a significant reduction of cell viability, colony formation ability, and induction of apoptosis. RNA-seq analyses showed that SMIP34-regulated genes were negatively correlated with ribosome biogenesis and eukaryotic translation pathways. Mechanistic studies showed that the Rix complex, which is essential for ribosomal biogenesis, is disrupted upon SMIP34 binding to PELP1. Biochemical assays confirmed that SMIP34 reduced ribosomal biogenesis and new protein synthesis. Further, SMIP34 enhanced the efficacy of mTOR inhibitors in reducing viability of ECa cells. SMIP34 is also effective in reducing cell viability in ECa organoids in vitro and explants ex vivo. Importantly, SMIP34 treatment resulted in a significant reduction of the growth of ECa xenografts. Collectively, these findings underscore the potential of SMIP34 in treating ECa.

Pharmacological inhibition of KDM1A/LSD1 enhances estrogen receptor beta-mediated tumor suppression in ovarian cancer.

Ovarian cancer (OCa) is the most lethal gynecologic cancer. Emerging data indicates that estrogen receptor beta (ERß) functions as a tumor suppressor in OCa. Lysine-specific histone demethylase 1A (KDM1A) is an epigenetic modifier that acts as a coregulator for steroid hormone receptors. However, it remain unknown if KDM1A interacts with ERß and regulates its expression/functions in OCa. Analysis of TCGA data sets indicated KDM1A and ERß expression showed an inverse relationship in OCa. Knockout (KO), knockdown (KD), or inhibition of KDM1A increased ERß isoform 1 expression in established and patient-derived OCa cells. Further, KDM1A interacts with and functions as a corepressor of ERß, and its inhibition enhances ERß target gene expression via alterations of histone methylation marks at their promoters. Importantly, KDM1A-KO or -KD enhanced the efficacy of ERß agonist LY500307, and the combination of KDM1A inhibitor (KDM1Ai) NCD38 with ERß agonist synergistically reduced the cell viability, colony formation, and invasion of OCa cells. RNA-seq and DIA mass spectrometry analyses showed that KDM1A-KO resulted in enhanced ERß signaling and that genes altered by KDM1A-KO and ERß agonist were related to apoptosis, cell cycle, and EMT. Moreover, combination treatment significantly reduced the tumor growth in OCa orthotopic, syngeneic, and patient-derived xenograft models and proliferation in patient-derived explant models. Our results demonstrate that KDM1A regulates ERß expression/functions, and its inhibition improves ERß mediated tumor suppression. Overall, our findings suggest that KDM1Ai and ERß agonist combination therapy is a promising strategy for OCa.

Inhibition of LIFR Blocks Adiposity-Driven Endometrioid Endometrial Cancer Growth.

Endometrial cancer (EC) is the fourth most common cancer in women, and half of the endometrioid EC (EEC) cases are attributable to obesity. However, the underlying mechanism(s) of obesity-driven EEC remain(s) unclear. In this study, we examined whether LIF signaling plays a role in the obesity-driven progression of EEC. RNA-seq analysis of EEC cells stimulated by adipose conditioned medium (ADP-CM) showed upregulation of LIF/LIFR-mediated signaling pathways including JAK/STAT and interleukin pathways. Immunohistochemistry analysis of normal and EEC tissues collected from obese patients revealed that LIF expression is upregulated in EEC tissues compared to the normal endometrium. Treatment of both primary and established EEC cells with ADP-CM increased the expression of LIF and its receptor LIFR and enhanced proliferation of EEC cells. Treatment of EEC cells with the LIFR inhibitor EC359 abolished ADP-CM induced colony formation andcell viability and decreased growth of EEC organoids. Mechanistic studies using Western blotting, RT-qPCR and reporter assays confirmed that ADP-CM activated LIF/LIFR downstream signaling, which can be effectively attenuated by the addition of EC359. In xenograft assays, co-implantation of adipocytes significantly enhanced EEC xenograft tumor growth. Further, treatment with EC359 significantly attenuated adipocyte-induced EEC progression in vivo. Collectively, our data support the premise that LIF/LIFR signaling plays an important role in obesity-driven EEC progression and the LIFR inhibitor EC359 has the potential to suppress adipocyte-driven tumor progression.

LIF/LIFR oncogenic signaling is a novel therapeutic target in endometrial cancer.

Endometrial cancer (EC) is the fourth most common cancer in women. Advanced-stage EC has limited treatment options with a poor prognosis. There is an unmet need for the identification of actionable drivers for the development of targeted therapies in EC. Leukemia inhibitory factor receptor (LIFR) and its ligand LIF play a major role in cancer progression, metastasis, stemness, and therapy resistance. However, little is known about the functional significance of the LIF/LIFR axis in EC progression. In this study using endometrial tumor tissue arrays, we identified that expression of LIF, LIFR is upregulated in EC. Knockout of LIFR using CRISPR/Cas9 in two different EC cells resulted in a significant reduction of their cell viability and cell survival. In vivo studies demonstrated that LIFR-KO significantly reduced EC xenograft tumor growth. Treatment of established and primary patient-derived EC cells with a novel LIFR inhibitor, EC359 resulted in the reduction of cell viability with an IC50 in the range of 20-100 nM and induction of apoptosis. Further, treatment with EC359 reduced the spheroid formation of EC cancer stem cells and reduced the levels of cancer stem cell markers SOX2, OCT4, NANOG, and Axin2. Mechanistic studies demonstrated that EC359 treatment attenuated the activation of LIF-LIFR driven pathways, including STAT3 and AKT/mTOR signaling in EC cells. Importantly, EC359 treatment resulted in a significant reduction of the growth of EC patient-derived explants ex vivo, EC cell line-derived xenografts, and patient-derived xenografts in vivo. Collectively, our work revealed the oncogenic potential of the LIF/LIFR axis in EC and support the utility of LIFR inhibitor, EC359, as a novel targeted therapy for EC via the inhibition of LIF/LIFR oncogenic signaling.

A rectovaginal septum mass in a BRCA1 positive patient years after risk reducing surgery: A case report.

•Rectovaginal septum mass in BRCA1 positive patient after risk reducing BSO years prior.•Papillary serous carcinoma presenting as a rectovaginal septum mass.•PAOLA-1 trial discussion for rectovaginal septum mass in BRCA1 positive patient.

PAI-1-Dependent Inactivation of SMAD4-Modulated Junction and Adhesion Complex in Obese Endometrial Cancer.

While plasminogen activator inhibitor-1 (PAI-1) is known to potentiate cellular migration via proteolytic regulation, this adipokine is implicated as an oncogenic ligand in the tumor microenvironment. To understand the underlying paracrine mechanism, here, we conduct transcriptomic analysis of 1,898 endometrial epithelial cells (EECs) exposed and unexposed to PAI-1-secreting adipose stromal cells. The PAI-1-dependent action deregulates crosstalk among tumor-promoting and tumor-repressing pathways, including transforming growth factor ß (TGF-ß). When PAI-1 is tethered to lipoprotein receptor-related protein 1 (LRP1), the internalized signaling causes downregulation of SMAD4 at the transcriptional and post-translational levels that attenuates TGF-ß-related transcription programs. Repression of genes encoding the junction and adhesion complex preferentially occurs in SMAD4-underexpressed EECs of persons with obesity. The findings highlight a role of PAI-1 signaling that renders ineffective intercellular communication for the development of adiposity-associated endometrial cancer.

Incidental Microscopic Granulosa Cell Proliferations: Tumorlet or Non-neoplastic?

Microscopic granulosa cell proliferations are rare incidental findings in ovarian and extraovarian tissue, and can simulate granulosa cell tumors. We report a case of a 22-yr-old nulliparous woman with incidental microscopic granulosa cell proliferation in the wall of an endometriotic cyst. Excision of the cyst revealed extensive endometriosis and incidental microscopic nests and cords of granulosa cells measuring 1.5 mm and positive for inhibin A and calretinin. A rare Call-Exner body was noted. An extensive literature review of both ovarian and extraovarian granulosa cell proliferations were performed to determine their possible origin, variable morphologic features and differential diagnosis. Forty-seven cases were identified occurring in pregnant (26%) and nonpregnant (74%) settings. The nonpregnant cases reveal an association with endometriosis and endometrioid adenocarcinomas. Follow-up data is very limited, but no reports of subsequent granulosa cell tumors are reported. We hypothesize that possible anovulation may play a role in the development of these proliferations. Because of unknown pathogenesis and limited follow-up data being available, their clinical significance and risk of neoplasia remain unclear.

Endometrial Polyp of the Placental Fetal Membranes: Report of a Case.

Endometrial polyps embedded in the fetal membranes have only rarely been described. A review of the English literature showed only one abstract describing this occurrence and to the best of our knowledge, there have been no other publications of this entity. Herein we present a case of a 37-yr-old woman with a history prior abortion and complicated pregnancy (type 2 diabetes mellitus and preeclampsia) who delivered by cesarean section. Although the placenta did not show hypertensive vasculopathic changes or other pathologic findings, an endometrial polyp embedded within the fetal membranes was present. Recognition of this rarely reported entity is important in order to avoid confusion with a significant neoplastic process.

Adipokines Deregulate Cellular Communication via Epigenetic Repression of Gap Junction Loci in Obese Endometrial Cancer.

Emerging evidence indicates that adipose stromal cells (ASC) are recruited to enhance cancer development. In this study, we examined the role these adipocyte progenitors play relating to intercellular communication in obesity-associated endometrial cancer. This is particularly relevant given that gap junctions have been implicated in tumor suppression. Examining the effects of ASCs on the transcriptome of endometrial epithelial cells (EEC) in an in vitro coculture system revealed transcriptional repression of GJA1 (encoding the gap junction protein Cx43) and other genes related to intercellular communication. This repression was recapitulated in an obesity mouse model of endometrial cancer. Furthermore, inhibition of plasminogen activator inhibitor 1 (PAI-1), which was the most abundant ASC adipokine, led to reversal of cellular distribution associated with the GJA1 repression profile, suggesting that PAI-1 may mediate actions of ASC on transcriptional regulation in EEC. In an endometrial cancer cohort (n = 141), DNA hypermethylation of GJA1 and related loci TJP2 and PRKCA was observed in primary endometrial endometrioid tumors and was associated with obesity. Pharmacologic reversal of DNA methylation enhanced gap-junction intercellular communication and cell-cell interactions in vitro. Restoring Cx43 expression in endometrial cancer cells reduced cellular migration; conversely, depletion of Cx43 increased cell migration in immortalized normal EEC. Our data suggest that persistent repression by ASC adipokines leads to promoter hypermethylation of GJA1 and related genes in the endometrium, triggering long-term silencing of these loci in endometrial tumors of obese patients. SIGNIFICANCE: Studies reveal that adipose-derived stem cells in endometrial cancer pathogenesis influence epigenetic repression of gap junction loci, which suggests targeting of gap junction activity as a preventive strategy for obesity-associated endometrial cancer.

Leiomyosarcoma of the uterine cervix associated with pregnancy: A case report and review of literature.

•Management of cervical leiomyosarcoma in pregnancy requires a multidisciplinary approach.•Ovarian preservation is preferred in young patients with early stage cervical leiomyosarcoma.•Routine lymphadenectomy in patients with early stage cervical leiomyosarcoma is not useful.

Clinical and Pathologic Features of Hispanic Endometrial Cancer Patients With Loss of Mismatch Repair Expression.

OBJECTIVES: Approximately 3% to 5% of endometrial cancers (EC) are associated with Lynch syndrome (LS). The clinical characteristics and prevalence of LS have not been well studied in the US Hispanic population. Hispanics are the largest and fastest growing ethnic minority group in the United States. We sought to characterize the demographics, tumor characteristics, and prevalence of loss of mismatch repair (MMR) protein expression in a large Hispanic population with EC. METHODS: From January 1, 2005, to August 1, 2012, 83 women of Hispanic ethnicity diagnosed with EC 50 years and younger were identified. Clinical and pathologic data were abstracted from the electronic medical record. Tumor studies included immunohistochemistry of MLH1, MSH2, MSH6, and PMS2 and methylation of the MLH1 promoter. RESULTS: Ninety-five percent of patients were overweight or obese. The mean body mass index was 40.1 kg/m, 75% had irregular menses, 36% had diabetes, 46% were nulliparous, and 95% had endometrioid histology. Thirteen patients (15.7%) had tumor MMR deficiency due to a presumed germline mutation (9 MSH6, 3 MSH2, and 1 MLH1). The pattern of MMR protein loss was consistent with the expected binding properties of the MMR heterodimer complexes. No significant difference was found in clinical or pathological variables between patients with and without MMR deficient tumors. CONCLUSIONS: The prevalence of molecular findings consistent with LS was at least as high as other populations of varied geography, race, and ethnicity. We found no reliable factors to include body mass index, family history, synchronous tumors, or pathologic tumor features to serve as triage markers for which ECs should be screened for MMR protein loss. Our findings support a recommendation for universal screening of ECs utilizing 2-antibody testing with MLH1 promoter methylation testing as indicated up to 60 years or older. Our recommendations should be generalizable to other Hispanic populations in the Southern United States.

Extrauterine Pelvic Serous Carcinomas: Current Update on Pathology and Cross-sectional Imaging Findings.

The spectrum of extrauterine pelvic serous carcinomas includes ovarian serous carcinoma, primary peritoneal serous carcinoma, and primary fallopian tube carcinoma. Ovarian serous carcinoma, the most common ovarian malignant epithelial neoplasm, consists of two distinct entities: high-grade and low-grade serous carcinomas. Primary peritoneal serous carcinoma and primary fallopian tube carcinoma are rare malignancies that share many characteristics of high-grade serous carcinomas. Recent advances in the genetics and molecular biology of gynecologic cancers have suggested a common origin of many extrauterine pelvic serous carcinomas from fallopian tube epithelium. With the exception of low-grade serous carcinomas, which arise from cortical inclusion cysts lined by tubal epithelium, most extrauterine pelvic serous carcinomas are believed to originate from serous tubal intraepithelial carcinomas and show similar clinical-biologic behaviors and natural histories. Indeed, the International Federation of Gynecology and Obstetrics Committee on Gynecologic Oncology recently recognized that these cancers should be considered collectively, with a common system of staging and management strategies for ovarian, primary peritoneal, and fallopian tube cancers. A paradigm shift has occurred in our understanding of the pathogenesis of extrauterine pelvic serous carcinomas that has the potential to change current strategies for screening, prevention, diagnosis, and management. Ultrasonography (US), computed tomography (CT), magnetic resonance imaging, and combined positron emission tomography and CT are pivotal in screening, initial diagnosis, and treatment follow-up; however, because of this paradigm shift, new radiologic techniques, such as contrast material-enhanced US and molecular US imaging, and various optical imaging techniques are being investigated as important screening and diagnostic tools. Because of evolving knowledge of genetic and molecular changes underlying the pathogenesis of extrauterine pelvic serous carcinomas, new targeted therapies are being developed to improve patient prognosis. (©)RSNA, 2016.

Successful pregnancy following myomectomy for uterine smooth muscle tumor of uncertain malignant potential: A case report and review of the literature.

•STUMPs are rare smooth muscle tumors with an overall favorable prognosis.•Pregnancy is possible after diagnosis of STUMP treated with myomectomy•Management of patients desiring fertility with STUMPs requires a multidisciplinary approach.

Impaired Development of Early Endometriotic Lesions in CD44 Knockout Mice.

Previous studies have shown endometrial cell (EC) CD44 and peritoneal mesothelial cell (PMC)-associated hyaluronan (hyaluronic acid [HA]) are involved in the attachment of endometrial stroma and epithelial cells to peritoneal mesothelium. Here we assess the CD44-HA interaction in the formation of the early endometriotic lesion using CD44(-/-) (knockout) mice. Using an established murine model and crossover technique, endometrial tissue from donor mice (wild type [WT] and CD44(-/-)) was used to induce endometriosis in recipient mice (WT and CD44(-/-)). Endometriotic lesions were visualized by fluorescent microscopy and confirmed by hematoxylin and eosin staining. Early endometriotic lesions were decreased when CD44(-/-) endometrium was placed in WT recipients and when WT endometrium was placed in CD44(-/-) recipients (P = .002). Early endometriotic lesions were also significantly decreased when both peritoneal and endometrial tissues lacked CD44 expression (P < .01). These studies demonstrate that both EC and PMC CD44 play a role in the development of early endometriotic lesion.

The frequency of herpes simplex virus changes in anal Pap smear and its association with squamous intraepithelial lesions in high-risk male patients.

While there are studies postulating a model of synergism between human papillomavirus (HPV) and herpes simplex virus (HSV) in cervical carcinogenesis, the frequency of anal herpes as well as its association with anal squamous intraepithelial lesions (ASILs) has been understudied in men. This study evaluates the frequency of HSV changes in anal Pap smears and its association with ASILs in a high-risk population. A computerized search for specimens associated with anal cytology that had positive findings of HSV was performed. The electronic medical records were examined for past diagnosis of herpes, HSV serology prior to or after cytology, and if the patient received treatment after cytologic diagnosis of HSV. Of the 470 anal Pap smears (Thin-prep) examined, seven had cellular changes consistent with HSV infection. All patients were asymptomatic human immunodeficiency virus (HIV) positive males with no prior HSV serology tests. Two patients had prior diagnoses of HSV infection. Cytologic abnormalities were identified in 86% ranging from atypical squamous cells of undetermined significance to high grade squamous intraepithelial lesion. Three patients were treated after the HSV cytologic diagnosis. The frequency of HSV changes in anal Pap smear is low (1.48%), but the presence of concomitant cytologic abnormalities is high (86%). While our findings suggest the possible role of HSV as a HPV co-factor in ASILs, larger studies are needed to support this. Identification of HSV infection on anal Pap smear is important for institution of patient treatment and subsequent reduction of transmission.

Isolated vaginal myeloid sarcoma in a 16-year-old girl.

Involvement of the female genital tract by myeloid sarcoma as the initial presentation is extremely uncommon, especially in the vagina. The lack of specific histologic features and the unusual location can be a diagnostic challenge to both the surgical pathologist and the clinician. The very few reported cases of myeloid sarcoma occurring in the vagina have been exclusively seen in adults. We report a 16-year-old girl who presented with a vaginal mass of 4 weeks duration. The initial clinical impression was a Bartholin cyst vs an abscess. However, because of persistence of the vaginal mass after a full course of antibiotic treatment, a biopsy was performed. Immunohistochemistry supported the diagnosis of myeloid sarcoma. Peripheral blood and bone marrow studies were normal. The patient received 4 cycles of chemotherapy and remained disease free 5 months from therapy completion. The clinical course, diagnostic workup, and differential diagnosis of our patient are discussed. Reported cases of myeloid sarcoma occurring in the vagina are reviewed and summarized.

Micropapillary carcinoma of the urinary bladder: report of a case and review of its cytologic features.

Micropapillary carcinoma (MPC) is an aggressive variant of urothelial carcinoma. MPC has a propensity to invade lymphovascular spaces and detrusor muscle early in the disease that often leads to upstaging and/or lymph node metastasis in many cases at cystectomy. Its association with the usual high-grade urothelial carcinoma provides an easy recognition of malignancy in cytology specimens without attempt at separating or identifying the MPC component. This may be related to our limited familiarity of its cytologic features with only 4 cases described in the literature. We report another case of MPC and highlight its features in cytologic preparations including the presence of singly scattered tumor cells with high nuclear to cytoplasmic ratio and pleomorphic nuclei, clustered cells devoid of fibrovascular core (micropapillae), 3-dimensional cell aggregates, cytoplasmic vacuoles, and micropapillae exhibiting some features of low-grade urothelial neoplasm. Appreciation of these features may help facilitate its early diagnosis and hopefully a better outcome for these aggressive tumors.