Thy-1 predicts poor prognosis and is associated with self-renewal in ovarian cancer.

BACKGROUND: Ovarian cancer is the leading cause of gynecologic cancer death in the United States despite effective first-line systemic chemotherapy. Cancer stem cells (CSCs) retain the ability to self-renew and proliferate and may be a means of harboring disease that evades standard treatment strategies. We previously performed a high-throughput screen to assess differential protein expression in ovarian CSCs compared to non-CSCs and observed that Thy-1 was more highly expressed in CSCs. Our primary aim was to validate Thy-1 (CD90) as a cancer stem cell (CSC) marker in epithelial ovarian cancer (EOC), correlate with clinical outcomes, and assess as a potential therapeutic target. RESULTS: Kaplan Meier (KM) Plotter data were correlated with survival outcomes. Quantitative real-time PCR, flow cytometry, and immunoblots assessed RNA and protein expression. Limiting dilution assays assessed self-renewal capacity and proliferation assays assessed proliferative capacity. RNA in-situ hybridization was performed on patient specimens to assess feasibility. Thy-1 (CD90) is more highly expressed in ovarian CSCs than non-CSCs, in EOC compared to benign ovarian epithelium (P < 0.001), and is highest in serous EOC (P < 0.05). Serous ovarian cancers with high Thy-1 expression have poorer outcomes (median PFS 15.8 vs. 18.3 months, P = 0 < 0.001; median OS 40.1 v. 45.8 months, P = 0.036). Endometrioid ovarian cancers with high Thy-1 have poorer PFS, but no difference in OS (upper quartile PFS 34 v. 11 months, P = 0.013; quartile OS not reached, P = 0.69). In vitro, Thy-1 expression is higher in CSCs versus non-CSCs. EOC cells with high Thy-1 expression demonstrate increased proliferation and self-renewal. Thy-1 knockdown in EOC cells decreases proliferative capacity and self-renewal capacity, and knockdown is associated with decreased expression of stem cell transcription factors NANOG and SOX2. RNA in situ hybridization is feasible in ovarian cancer tissue specimens. CONCLUSIONS: Thy-1 is a marker of ovarian CSCs. Increased expression of Thy-1 in EOC predicts poor prognosis and is associated with increased proliferative and self-renewal capacity. Thy-1 knockdown decreases proliferative and self-renewal capacity, and represents a potential therapeutic target.

Cx26 drives self-renewal in triple-negative breast cancer via interaction with NANOG and focal adhesion kinase.

Tumors adapt their phenotypes during growth and in response to therapies through dynamic changes in cellular processes. Connexin proteins enable such dynamic changes during development, and their dysregulation leads to disease states. The gap junction communication channels formed by connexins have been reported to exhibit tumor-suppressive functions, including in triple-negative breast cancer (TNBC). However, we find that connexin 26 (Cx26) is elevated in self-renewing cancer stem cells (CSCs) and is necessary and sufficient for their maintenance. Cx26 promotes CSC self-renewal by forming a signaling complex with the pluripotency transcription factor NANOG and focal adhesion kinase (FAK), resulting in NANOG stabilization and FAK activation. This FAK/NANOG-containing complex is not formed in mammary epithelial or luminal breast cancer cells. These findings challenge the paradigm that connexins are tumor suppressors in TNBC and reveal a unique function for Cx26 in regulating the core self-renewal signaling that controls CSC maintenance.

CD55 regulates self-renewal and cisplatin resistance in endometrioid tumors.

Effective targeting of cancer stem cells (CSCs) requires neutralization of self-renewal and chemoresistance, but these phenotypes are often regulated by distinct molecular mechanisms. Here we report the ability to target both of these phenotypes via CD55, an intrinsic cell surface complement inhibitor, which was identified in a comparative analysis between CSCs and non-CSCs in endometrioid cancer models. In this context, CD55 functions in a complement-independent manner and required lipid raft localization for CSC maintenance and cisplatin resistance. CD55 regulated self-renewal and core pluripotency genes via ROR2/JNK signaling and in parallel cisplatin resistance via lymphocyte-specific protein tyrosine kinase (LCK) signaling, which induced DNA repair genes. Targeting LCK signaling via saracatinib, an inhibitor currently undergoing clinical evaluation, sensitized chemoresistant cells to cisplatin. Collectively, our findings identify CD55 as a unique signaling node that drives self-renewal and therapeutic resistance through a bifurcating signaling axis and provides an opportunity to target both signaling pathways in endometrioid tumors.

Cisplatin induces stemness in ovarian cancer.

The mainstay of treatment for ovarian cancer is platinum-based cytotoxic chemotherapy. However, therapeutic resistance and recurrence is a common eventuality for nearly all ovarian cancer patients, resulting in poor median survival. Recurrence is postulated to be driven by a population of self-renewing, therapeutically resistant cancer stem cells (CSCs). A current limitation in CSC studies is the inability to interrogate their dynamic changes in real time. Here we utilized a GFP reporter driven by the NANOG-promoter to enrich and track ovarian CSCs. Using this approach, we identified a population of cells with CSC properties including enhanced expression of stem cell transcription factors, self-renewal, and tumor initiation. We also observed elevations in CSC properties in cisplatin-resistant ovarian cancer cells as compared to cisplatin-naïve ovarian cancer cells. CD49f, a marker for CSCs in other solid tumors, enriched CSCs in cisplatin-resistant and -naïve cells. NANOG-GFP enriched CSCs (GFP+ cells) were more resistant to cisplatin as compared to GFP-negative cells. Moreover, upon cisplatin treatment, the GFP signal intensity and NANOG expression increased in GFP-negative cells, indicating that cisplatin was able to induce the CSC state. Taken together, we describe a reporter-based strategy that allows for determination of the CSC state in real time and can be used to detect the induction of the CSC state upon cisplatin treatment. As cisplatin may provide an inductive stress for the stem cell state, future efforts should focus on combining cytotoxic chemotherapy with a CSC targeted therapy for greater clinical utility.

The Impact of Obesity on the 30-day Morbidity and Mortality After Surgery for Ovarian Cancer.

OBJECTIVES: To examine the effect of body mass index (BMI) on postoperative 30-day morbidity and mortality after surgery for ovarian cancer (OC). METHODS: Patients with OC were identified from the American College of Surgeons National Surgical Quality Improvement Program from 2005 to 2011. Women were divided into 3 groups: nonobese (BMI <30 kg/m), obese (30 to <40 kg/m), and morbidly obese (≥40 kg/m). Multivariable logistic regression models were performed. RESULTS: Of 2061 women included in this study, 1336 (65%) were nonobese, 560 (27%) were obese, and 165 (8%) were morbidly obese. The overall 30-day mortality and morbidity rates for the entire cohort were 2% and 31%, respectively. In multivariate analyses adjusting for confounders, both obese (odds ratio [OR], 0.9; 95% confidence interval [CI], 0.4-2.0; P = 0.87) and morbid obesity (OR, 0.8; 95% CI, 0.1-3.0; P = 0.73) were not significant predictors of increased 30-day postoperative mortality. Likewise, rates of any complication in 30 days were comparable between nonobese, obese, and morbidly obese patients (31% vs. 28% vs. 33%, respectively; P = 0.35) with no significant difference even after adjusting for other confounders (OR, 0.9; 95% CI, 0.7-1.1; P = 0.26 and OR, 1.1; 95% CI, 0.7-1.6; P = 0.70, respectively). Obese and morbidly obese patients were more likely to have diabetes, hypertension requiring medications, cardiac morbidities, higher American Society of Anesthesiologists class, and leukocytosis and less likely to have weight loss before surgery. CONCLUSIONS: With appropriate control for confounding comorbidities, the 30-day morbidity and mortality rates for the obese and morbidly obese patients undergoing surgery for OC do not seem to differ. Therefore, reported inferior long-term survival for these patients is likely related to a different phase of their disease and treatment process and is deserving of further investigation.

Impact of Age on 30-Day Mortality and Morbidity in Patients Undergoing Surgery for Ovarian Cancer.

OBJECTIVE: To examine the effect of age on postoperative 30-day morbidity and mortality after surgery for ovarian cancer. METHODS: The American College of Surgeons National Surgical Quality Improvement Program files were used to identify patients with ovarian cancer who underwent surgery in 2005 to 2011. Women were divided into 4 age groups: <60, 60 to 69, 70 to 79, and ≥80 years. Multivariable logistic regression models were performed. RESULTS: Of 2087 patients included, 47% were younger than 60 years, 28% were 60 to 69 years old, 18% were 70 to 79 years old, and 7% were 80 years or older. Overall 30-day mortality and morbidity rates were 2% and 30%. Elderly patients 80 years or older were more likely to die within 30 days compared with patients younger than 60 years, 60 to 69 years old, and 70 to 79 years old (9.2% vs. 0.6% vs .2.8% vs 2.5%, P < 0.001). Elderly patient aged 80 years or older were more likely to develop pulmonary (9% vs 2% vs 5% vs 3%, P < 0.001) and septic (9% vs 3% vs 5% vs 4%, P = 0.01) complications compared with patients younger than 60 years, 60 to 69 years old, and 70 to 79 years old, respectively. No difference in the risk of renal (0.2% vs 1% vs 1% vs 1%, P = 0.20) complications and surgical reexploration (4% vs 4% vs 3% vs 5%, P = 0.80) between the 4 age groups. In multivariable analyses after adjusting for other confounders, age was a significant predictor of 30-day postoperative mortality and morbidity. Compared with younger patients, octogenarians were 9-times more likely to die and 70% more likely to develop complications within 30 days after surgery. Other significant predictors of 30-day mortality were higher American Society of Anesthesiologists class and hypoalbuminemia (serum albumin ≤ 3 g/dL), whereas, surgical complexity, higher American Society of Anesthesiologists class, longer operative time, and hypoalbuminemia were other significant predictors of 30-day morbidity. CONCLUSIONS: Elderly patients have a higher risk of perioperative mortality and morbidity within 30 days. Therefore, those patients should be counseled thoroughly about the risk of primary debulking surgery vs neoadjuvant chemotherapy.

Development of a Fluorescent Reporter System to Delineate Cancer Stem Cells in Triple-Negative Breast Cancer.

Advanced cancers display cellular heterogeneity driven by self-renewing, tumorigenic cancer stem cells (CSCs). The use of cell lines to model CSCs is challenging due to the difficulty of identifying and isolating cell populations that possess differences in self-renewal and tumor initiation. To overcome these barriers in triple-negative breast cancer (TNBC), we developed a CSC system using a green fluorescent protein (GFP) reporter for the promoter of the well-established pluripotency gene NANOG. NANOG-GFP+ cells gave rise to both GFP+ and GFP(-) cells, and GFP+ cells possessed increased levels of the embryonic stem cell transcription factors NANOG, SOX2, and OCT4 and elevated self-renewal and tumor initiation capacities. GFP+ cells also expressed mesenchymal markers and demonstrated increased invasion. Compared with the well-established CSC markers CD24(-) /CD44(+) , CD49f, and aldehyde dehydrogenase (ALDH) activity, our NANOG-GFP reporter system demonstrated increased enrichment for CSCs. To explore the utility of this system as a screening platform, we performed a flow cytometry screen that confirmed increased CSC marker expression in the GFP+ population and identified new cell surface markers elevated in TNBC CSCs, including junctional adhesion molecule-A (JAM-A). JAM-A was highly expressed in GFP+ cells and patient-derived xenograft ALDH+ CSCs compared with the GFP(-) and ALDH(-) cells, respectively. Depletion of JAM-A compromised self-renewal, whereas JAM-A overexpression induced self-renewal in GFP(-) cells. Our data indicate that we have defined and developed a robust system to monitor differences between CSCs and non-CSCs in TNBC that can be used to identify CSC-specific targets for the development of future therapeutic strategies.

Patient, treatment and discharge factors associated with hospital readmission within 30 days after surgical cytoreduction for epithelial ovarian carcinoma.

OBJECTIVE: Hospital readmissions are common, costly and increasingly viewed as adverse events. In gynecologic oncology, data on readmissions are limited. The goal of this study was to examine the patient, treatment and discharge factors associated with unplanned readmission after cytoreductive surgery. METHODS: We identified all patients with stages II-IV ovarian cancer who underwent surgical cytoreduction at our institution between 2003 and 2011. A retrospective chart review was performed, and clinical variables were extracted. Utilizing linear and logistic regression, these clinical variables were correlated with risk of readmission. RESULTS: A total of 460 patients were included in the analysis, with the majority having a stage IIIC high grade serous cancer. Optimal cytoreduction (<1.0 cm residual disease) was obtained in 368 patients (81%), and 233 patients (50%) underwent at least one radical procedure. Perioperative complications were observed in 148 patients (32%). A large proportion of our cohort was discharged to rehabilitation facilities (12%) or with a visiting nurse (38%). Fifty five patients (12%) were readmitted within 30 days. On multivariate logistic regression, reoperation and perioperative cardiopulmonary event were the only factors associated with readmission (OR=3.2, 95% CI=1.7-6.0). Discharge home with ancillary services was not protective against readmission, even when controlling for perioperative complications (OR=1.18, 95% CI=0.53-2.64). CONCLUSIONS: Readmission after surgical cytoreduction affected 12% of our population. Multivariate analyses suggested perioperative complications, particularly reoperation and cardiopulmonary event, placed the patient at the greatest risk. Age, comorbidities, surgical radicality and discharge with visiting nurse services/rehabilitation facility did not affect the likelihood of readmission.

Management of a skin metastasis in a patient with advanced ovarian cancer.

► Skin metastasis of ovarian cancer is rare, often nodular in appearance, and conveys a poor prognosis. ► This patient developed an unusual maculo-papular rash which was biopsy-proven to be metastatic endometrioid adenocarcinoma. ► Pruritic symptoms from skin metastases should be palliated; SSRIs, local radiation, and topical creams all may play a role.

Genomewide RNA expression profiling in lung identifies distinct signatures in idiopathic pulmonary arterial hypertension and secondary pulmonary hypertension.

Idiopathic pulmonary arterial hypertension (PAH) is a life-threatening condition characterized by pulmonary arteriolar remodeling. This investigation aimed to identify genes involved specifically in the pathogenesis of PAH and not other forms of pulmonary hypertension (PH). Using genomewide microarray analysis, we generated the largest data set to date of RNA expression profiles from lung tissue specimens from 1) 18 PAH subjects and 2) 8 subjects with PH secondary to idiopathic pulmonary fibrosis (IPF) and 3) 13 normal subjects. A molecular signature of 4,734 genes discriminated among these three cohorts. We identified significant novel biological changes that were likely to contribute to the pathogenesis of PAH, including regulation of actin-based motility, protein ubiquitination, and cAMP, transforming growth factor-beta, MAPK, estrogen receptor, nitric oxide, and PDGF signaling. Bone morphogenic protein receptor type II expression was downregulated, even in subjects without a mutation in this gene. Women with PAH had higher expression levels of estrogen receptor 1 than normal women. Real-time quantitative PCR confirmed differential expression of the following genes in PAH relative to both normal controls and PH secondary to IPF: a disintegrin-like and metalloprotease with thrombospondin type 1 motif 9, cell adhesion molecule with homology to L1CAM, cytochrome b(558) and beta-polypeptide, coagulation factor II receptor-like 3, A-myb myeloblastosis viral oncogene homolog 1, nuclear receptor coactivator 2, purinergic receptor P2Y, platelet factor 4, phospholamban, and tropomodulin 3. This study shows that PAH and PH secondary to IPF are characterized by distinct gene expression signatures, implying distinct pathophysiological mechanisms.