Machine Learning Interpretation of Extended Human Papillomavirus Genotyping by Onclarity in an Asian Cervical Cancer Screening Population.

This study aimed (i) to compare the performance of the BD Onclarity human papillomavirus (HPV) assay with the Cobas HPV test in identifying cervical intraepithelial neoplasia 2/3 or above (CIN2/3+) in an Asian screening population and (ii) to explore improving the cervical cancer detection specificity of Onclarity by machine learning. We tested 605 stratified random archived samples of cervical liquid-based cytology samples with both assays. All samples had biopsy diagnosis or repeated negative cytology follow-up. Association rule mining (ARM) was employed to discover coinfection likely to give rise to CIN2/3+. Outcome classifiers interpreting the extended genotyping results of Onclarity were built with different underlying models. The sensitivities (Onclarity, 96.32%; Cobas, 95.71%) and specificities (Onclarity, 46.38%; Cobas, 45.25%) of the high-risk HPV (hrHPV) components of the two tests were not significantly different. When HPV16 and HPV18 were used to further interpret hrHPV-positive cases, Onclarity displayed significantly higher specificity (Onclarity, 87.10%; Cobas, 80.77%). Both hrHPV tests achieved the same sensitivities (Onclarity, 90.91%; Cobas, 90.91%) and similar specificities (Onclarity, 48.46%; Cobas, 51.98%) when used for triaging atypical squamous cells of undetermined significance. Positivity in both HPV16 and HPV33/58 of the Onclarity channels entails the highest probability of developing CIN2/3+. Incorporating other hrHPVs into the outcome classifiers improved the specificity of identifying CIN2/3 to up to 94.32%. The extended genotyping of Onclarity therefore can help to highlight patients having the highest risk of developing CIN2/3+, with the potential to reduce unnecessary colposcopy and negative psychosocial impact on women receiving the reports.

Amyloid Precursor Protein Overexpression in Down Syndrome Trophoblast Reduces Cell Invasiveness and Interferes with Syncytialization.

The placentas of Down syndrome (DS) pregnancies exhibit morphologic and functional abnormalities. Although the increase in dosage of certain genes on chromosome 21 has been associated with the DS phenotype, the effects on placenta have seldom been studied. Herein, we examine the expression of four dosage-sensitive genes (APP, ETS2, SOD1, and HMGN1) in normal and DS placentas. We demonstrated significant overexpression of amyloid precursor protein (APP) in DS placentas at RNA and protein levels by real-time quantitative PCR, Western blot analysis, and immunohistochemistry. Inducible APP overexpression trophoblast cell line models were established using a Tet-On system. APP induction in HTR-8/SVneo dose-dependently decelerated cell growth, enhanced apoptosis, and reduced cell migration and invasion when compared with the uninduced controls. Concomitantly, decreased ß-human chorionic gonadotropin in the culture medium was also detected on induction. Moreover, although forskolin treatment induced α/ß-human chorionic gonadotropin and syncytin expression in BeWo cells, such induction of syncytialization was inhibited by APP overexpression. E-cadherin immunofluorescence also demonstrated a decrease in syncytia formation in forskolin-treated BeWo-overexpressing APP. By liquid chromatography-tandem mass spectrometry, proteins related to cell-cell adhesion, protein translation, processing, and folding were found to be up-regulated in APP-induced HTR-8/SVneo clones. Our data demonstrated, for the first time, the effects of increased APP expression in DS placenta.

FBI-1 Is Overexpressed in Gestational Trophoblastic Disease and Promotes Tumor Growth and Cell Aggressiveness of Choriocarcinoma via PI3K/Akt Signaling.

Human placental trophoblasts can be considered pseudomalignant, with tightly controlled proliferation, apoptosis, and invasiveness. Gestational trophoblastic disease (GTD) represents a family of heterogeneous trophoblastic lesions with aberrant apoptotic and proliferative activities and dysregulation of cell signaling pathways. We characterize the oncogenic effects of factor that binds to the inducer of short transcripts of HIV-1 [FBI-1, alias POZ and Krüppel erythroid myeloid ontogenic factor (POKEMON)/ZBTB7A] in GTD and its role in promoting cell aggressiveness in vitro and tumor growth in vivo. IHC studies showed increased nuclear expression of FBI-1, including hydatidiform moles, choriocarcinoma (CCA), and placental site trophoblastic tumor, in GTD. In JAR and JEG-3 CCA cells, ectopic FBI-1 expression opposed apoptosis through repression of proapoptotic genes (eg, BAK1, FAS, and CASP8). FBI-1 overexpression also promoted Akt activation, as indicated by Akt-pS473 phosphorylation. FBI-1 overexpression promoted mobility and invasiveness of JEG-3 and JAR, but not in the presence of the phosphoinositide 3-kinase inhibitor LY294002. These findings suggest that FBI-1 could promote cell migration and invasion via phosphoinositide 3-kinase/Akt signaling. In vivo, nude mice injected with CCA cells with stable FBI-1 knockdown demonstrated reduced tumor growth compared with that in control groups. These findings suggest that FBI-1 is clinically associated with the progression of, and may be a therapeutic target in, GTD, owing to its diverse oncogenic effects on dysregulated trophoblasts.

Downregulation of ASPP2 in choriocarcinoma contributes to increased migratory potential through Src signaling pathway activation.

Gestational choriocarcinoma is a malignant tumor derived from placental trophoblast and the most aggressive member of gestational trophoblastic disease (GTD). Apoptosis-stimulating protein of p53-2 (ASPP2) is a member of ASPP family that transactivates p53 and thereby functions as a tumor suppressor. In this study, the expression profile of ASPP2 in choriocarcinoma was examined in comparison with normal placentas and hydatidiform moles, the latter being a type of GTD that carries malignant potential. Downregulation of ASPP2 messenger RNA and protein was demonstrated in choriocarcinoma by quantitative PCR and immunohistochemistry. ASPP2-transfected choriocarcinoma cells (JEG-3 and JAR) showed an increase in apoptosis and a decrease in cell migration as detected by TdT-mediated dUTP nick end labeling and wound healing assays, respectively, illustrating the complex action of ASPP2 on cell functions other than programmed cell death. Activated Src is known to be important in tumor progression. Transfection of ASPP2 but not ASPP1, another tumor-suppressive ASPP, was found to be related to subsequent decreased Src-pY416 phosphorylation, suggesting an inactivating effect of ASPP2 on Src. Moreover, this ASPP2-mediated inactivation of Src could be abolished by RNA interference with C-terminal Src kinase (Csk), a kinase that can inhibit Src activation. Our findings suggested that the ability of ASPP2 to attenuate Src activation was specific to ASPP2 in a Csk-dependent manner. Taken together, we demonstrated a loss of tumor-suppressive ASPP2 in choriocarcinoma with effects on cell migration and apoptosis. We also unveiled a possible mechanistic link between ASPP2 and Csk/Src signaling pathway, implicating the multiple cellular functions of ASPP2.

p21-activated kinase 4 regulates ovarian cancer cell proliferation, migration, and invasion and contributes to poor prognosis in patients.

Ovarian cancer is a lethal gynecological malignancy, and to improve survival, it is important to identify novel prognostic and therapeutic targets. In this study, we present a role for p21-activated kinase 4 (Pak4) in ovarian cancer progression. We show a significant association between increased expression of Pak4 and its activated form, phosphorylated (p)-Pak4 Ser(474), with metastasis of ovarian cancers, shorter overall and disease-free survival, advanced stage and high-grade cancers, serous/clear cell histological subtypes, and reduced chemosensitivity. Pak4 overexpression was also observed in ovarian cancer cell lines. Pak4 and p-Pak4 expression were detected both in the nucleus and cytoplasm of ovarian cancer cells, in vitro as well as in vivo. Stable knockdown of Pak4 in ovarian cancer cell lines led to reduced cell migration, invasion, and proliferation, along with reduced c-Src, ERK1/2, and epidermal growth factor receptor (EGFR) activation and decreased matrix metalloproteinase 2 (MMP2) expression. Conversely, Pak4 overexpression promoted ovarian cancer cell migration and invasion in a c-Src, MEK-1, MMP2, and kinase-dependent manner, and induced cell proliferation through the Pak4/c-Src/EGFR pathway that controls cyclin D1 and CDC25A expression. Stable knockdown of Pak4 also impeded tumor growth and dissemination in nude mice. This report reveals the association between Pak4 and important clinicopathologic parameters, suggesting Pak4 to be a significant prognostic marker and potential therapeutic molecular target in ovarian cancer. The implied possible cross-talk between Pak4 and EGFR suggests the potential of dual targeting of EGFR and Pak4 as a unique therapeutic approach for cancer therapy.

Overexpressed PAK4 promotes proliferation, migration and invasion of choriocarcinoma.

Gestational trophoblastic disease (GTD) includes frankly malignant choriocarcinoma (CCA) and placental site trophoblastic tumor and potentially malignant hydatidiform mole. p21-Activated kinase (PAK) 4 promotes cell motility. This study investigated the role of PAK4 in the pathogenesis of GTD. PAK4 messenger RNA and protein expressions in clinical samples and cell lines of normal placentas and GTD were determined by quantitative real-time polymerase chain reaction and western blot, respectively. The effects of human chorionic gonadotropin (hCG) and phosphoinositide 3 kinase (PI3K) on the expression and activation of PAK4 were investigated by treating CCA JEG3 and JAR cells with anti-hCG antibody and PI3K inhibitor, respectively. The effects of PAK4 on CCA cell proliferation, migration and invasion were assessed by corresponding functional assays. We demonstrated overexpression of PAK4 in GTD and CCA cell lines at both RNA and protein level. hCG is one of the upstream regulators of PAK4 expression, whereas activation of PAK4 is PI3K/PKB dependent in JEG3 and JAR cells. Significant correlation was found between PAK4 expression and proliferation index minichromosome maintenance complex component 7 (P = 0.007). In JEG3 and JAR cells, stably transfected PAK4 increased proliferation, migration and invasion, whereas small interfering RNA knockdown of PAK4 decreased proliferation, migration and invasion along with downregulated CDK6 and membrane-type 1 matrix metalloproteinase (MT1-MMP) and upregulated p16. We further found PAK4-mediated transcription of MT1-MMP in CCA cells by luciferase reporter assay. Our results demonstrated for the first time that overexpressed PAK4 was involved in the pathogenesis of GTD, promoting proliferation and enhancing cell migration and invasion in CCA cells.

Downregulation of ASPP1 in gestational trophoblastic disease: correlation with hypermethylation, apoptotic activity and clinical outcome.

Gestational trophoblastic disease encompasses a spectrum of trophoblastic lesions including true neoplasms such as choriocarcinomas and the potentially malignant hydatidiform moles, which may develop persistent disease requiring chemotherapy. ASPP1, a member of apoptosis-stimulating proteins of p53 (ASPPs), is a proapoptotic protein that can stimulate apoptosis through its interaction with p53. We evaluated the promoter methylation and expression profiles of ASPP1 in different trophoblastic tissues and its in vitro functional effect on two choriocarcinoma cell lines, namely JEG-3 and JAR. Significant downregulation of ASPP1 mRNA and protein levels was demonstrated in hydatidiform moles and choriocarcinomas, when compared with normal placentas by quantitative-PCR and immunohistochemistry. The ASPP1 mRNA level was significantly correlated with its hypermethylation status, evaluated with methylation-specific PCR, in placenta and gestational trophoblastic disease samples (P=0.024). Moreover, lower ASPP1 immunoreactivity was shown in hydatidiform moles that progressed to persistent gestational trophoblastic neoplasms than in those that regressed (P=0.045). A significant correlation was also found between expression of ASPP1 and proliferative indices (assessed by Ki67 and MCM7), apoptotic activity (M30 CytoDeath antibody), p53 and caspase-8 immunoreactivities. An in vitro study showed that ectopic expression of ASPP1 could trigger apoptosis through intrinsic and extrinsic pathways as indicated by an increase in cleaved caspase-9 and Fas ligand protein expression. The latter suggests a hitherto unreported novel link between ASPP1 and the extrinsic pathway of apoptosis. Our findings suggest that downregulation of ASPP1 by hypermethylation may be involved in the pathogenesis and progress of gestational trophoblastic disease, probably through its effect on apoptosis.

Overexpression of dedicator of cytokinesis I (Dock180) in ovarian cancer correlated with aggressive phenotype and poor patient survival.

AIMS: Dedicator of cytokinesis I (Dock180) is a novel guanine nucleotide exchange factor for Rho guanosine triphosphates (GTPases) important for cell migration. The aim of this study was to evaluate the role of Dock180 in ovarian carcinogenesis. METHODS AND RESULTS: Using immunohistochemistry, real-time polymerase chain reaction and Western blotting, overexpression of Dock180 RNA and protein was demonstrated in the nucleus and cytoplasm of ovarian cancer cell lines (n = 5) and clinical samples of ovarian borderline tumours (n = 21) and invasive cancers (n = 108) when compared with ovarian epithelial cell lines (n = 3) and benign cystadenomas (n = 10) (P < 0.05). High Dock180 cytoplasmic expression in ovarian cancer (n = 108) was associated significantly with serous histological type, high-grade cancer and advanced stage (P < 0.05), as well as poor overall and disease-free survival (P = 0.004). Using multivariate progression analysis, high Dock180 cytoplasmic expression and advanced cancer stage were found to be independent prognostic factors for short overall survival and disease-free survival (P < 0.05). Exogenous expression of Dock180 by transient transfection enhanced cancer cell migration and invasion, whereas knockdown of Dock180 by an siRNA approach retarded cancer cell migration and invasion in association with down-regulation of matrix metalloproteinase 2. CONCLUSIONS: Our findings suggest that Dock180 contributes to ovarian carcinogenesis and dissemination and is a potential prognostic marker and therapeutic target.

Targeting DNA Damage Response Pathway in Ovarian Clear Cell Carcinoma.

Ovarian clear cell carcinoma (OCCC) is one of the major types of ovarian cancer and is of higher relative prevalence in Asians. It also shows higher possibility of resistance to cisplatin-based chemotherapy leading to poor prognosis. This may be attributed to the relative lack of mutations and aberrations in homologous recombination-associated genes, which are crucial in DNA damage response (DDR), such as BRCA1, BRCA2, p53, RAD51, and genes in the Fanconi anemia pathway. On the other hand, OCCC is characterized by a number of genetic defects rendering it vulnerable to DDR-targeting therapy, which is emerging as a potent treatment strategy for various cancer types. Mutations of ARID1A, PIK3CA, PTEN, and catenin beta 1 (CTNNB1), as well as overexpression of transcription factor hepatocyte nuclear factor-1ß (HNF-1ß), and microsatellite instability are common in OCCC. Of particular note is the loss-of-function mutations in ARID1A, which is found in approximately 50% of OCCC. ARID1A is crucial for processing of DNA double-strand break (DSB) and for sustaining DNA damage signaling, rendering ARID1A-deficient cells prone to impaired DNA damage checkpoint regulation and hence sensitive to poly ADP ribose polymerase (PARP) inhibitors. However, while preclinical studies have demonstrated the possibility to exploit DDR deficiency in OCCC for therapeutic purpose, progress in clinical application is lagging. In this review, we will recapitulate the preclinical studies supporting the potential of DDR targeting in OCCC treatment, with emphasis on the role of ARID1A in DDR. Companion diagnostic tests (CDx) for predicting susceptibility to PARP inhibitors are rapidly being developed for solid tumors including ovarian cancers and may readily be applicable on OCCC. The potential of various available DDR-targeting drugs for treating OCCC by drawing analogies with other solid tumors sharing similar genetic characteristics with OCCC will also be discussed.

Overexpression of proto-oncogene FBI-1 activates membrane type 1-matrix metalloproteinase in association with adverse outcome in ovarian cancers.

BACKGROUND: FBI-1 (factor that binds to the inducer of short transcripts of human immunodeficiency virus-1) is a member of the POK (POZ and Kruppel) family of transcription factors and play important roles in cellular differentiation and oncogenesis. Recent evidence suggests that FBI-1 is expressed at high levels in a subset of human lymphomas and some epithelial solid tumors. However, the function of FBI-1 in human ovarian cancers remains elusive. RESULTS: In this study, we investigated the role of FBI-1 in human ovarian cancers, in particularly, its function in cancer cell invasion via modulating membrane type 1-matrix metalloproteinase (MT1-MMP). Significantly higher FBI-1 protein and mRNA expression levels were demonstrated in ovarian cancers samples and cell lines compared with borderline tumors and benign cystadenomas. Increased FBI-1 mRNA expression was correlated significantly with gene amplification (P = 0.037). Moreover, higher FBI-1 expression was found in metastatic foci (P = 0.036) and malignant ascites (P = 0.021), and was significantly associated with advanced stage (P = 0.012), shorter overall survival (P = 0.032) and disease-free survival (P = 0.016). In vitro, overexpressed FBI-1 significantly enhanced cell migration and invasion both in OVCA 420 and SKOV-3 ovarian carcinoma cells, irrespective of p53 status, accompanied with elevated expression of MT1-MMP, but not MMP-2 or TIMP-2. Moreover, knockdown of MT1-MMP abolished FBI-1-mediated cell migration and invasion. Conversely, stable knockdown of FBI-1 remarkably reduced the motility of these cells with decreased expression of MT1-MMP. Promoter assay and chromatin immunoprecipitation study indicated that FBI-1 could directly interact with the promoter spanning ~600 bp of the 5'-flanking sequence of MT1-MMP and enhanced its expression in a dose-dependent manner. Furthermore, stable knockdown and ectopic expression of FBI-1 decreased and increased cell proliferation respectively in OVCA 420, but not in the p53 null SKOV-3 cells. CONCLUSIONS: Our results suggested an important role of FBI-1 in ovarian cancer cell proliferation, cell mobility, and invasiveness, and that FBI-1 can be a potential target of chemotherapy.

Napsin A, Hepatocyte Nuclear Factor-1-Beta (HNF-1ß), Estrogen and Progesterone Receptors Expression in Arias-Stella Reaction.

BACKGROUND: The Arias-Stella reaction (ASR) can mimic endometrial clear cell carcinoma (ECCC) in small biopsies, especially when drug or pregnancy history is unknown. A panel of immunohistochemical markers comprising napsin A, hepatocyte nuclear factor-1-beta (HNF-1ß), estrogen and progesterone receptors (ER, PR) has been found useful in confirming a diagnosis of ECCC. However, the detailed characterization of how expression of this combination of markers in the ECCC mimics ASR has yet to be thoroughly evaluated. DESIGN: The frequency and extent of napsin A, HNF-1ß, ER, and PR expression in ASR were assessed in a large series. For napsin A, any cytoplasmic staining was considered positive while only nuclear staining was deemed to be positive for HNF-1ß, ER, and PR. Immunohistochemical histoscores based on the intensity and extent of staining were calculated. RESULTS: Forty cases were gestational and 10 were nongestational ASR. In 19 (38%), the reaction was extensive and involved >50% of the glands. A stromal decidual change was found in 31 (77.5%) of the gestational and 3 (30%) of the nongestational cases. Napsin A was positive in all gestational and 8 of 10 (80%) nongestational ASR. All ASR showed HNF-1ß expression. ER expression was reduced in 37 (92.5%) and lost in 3 (7.5%) gestational ASR, and reduced in 9 (90%) and lost in 1 (10%) of nongestational ASR. None of the ASR in our series expressed PR. CONCLUSIONS: Naspin A and HNF-1ß were frequently expressed in both gestational and nongestational ASR, and ER expression was usually either reduced or loss. Interpretation of these markers in small biopsies containing atypical clear cells should be made with caution.

HPV genotyping and E6/E7 transcript assays for cervical lesion detection in an Asian screening population-Cobas and Aptima HPV tests.

BACKGROUND: High-risk human papillomavirus (hrHPV) detection and genotyping by Cobas HPV test has become an important technical platform in cervical cancer screening. It may be used as a co-test with cervical cytology or as a standalone test. Aptima HPV assay (AHPV) is another hrHPV test detecting 13 genotypes through qPCR based amplification of viral E6/E7 transcripts. Partial genotyping with Aptima HPV 16 18/45 genotype assay (AHPV GT) on positive samples is possible. Evidence supporting the performance of AHPV in Asian populations is scarce. OBJECTIVE: To compare the performances of Cobas and AHPV in detection of cervical squamous intraepithelial lesions (SIL) and triage of cytologically equivocal smears in a cohort of Hong Kong women. STUDY DESIGN: 442 liquid based cytology (LBC) residues with biopsy confirmed diagnoses were evaluated by both AHPV and Cobas HPV tests. RESULTS: Overall, there was a moderate agreement between AHPV and Cobas (κ = 0.5082, 95% CI: 0.492-0.672). The sensitivities of AHPV and Cobas for detecting biopsy confirmed HSIL or worse lesions (HSIL+) were 96.71% (95% CI: 92.49%-98.92%) and 97.37% (95% CI: 93.40%-99.28%) respectively. AHPV demonstrated significantly higher specificity than Cobas (37.85% vs 23.96%, p < 0.0001). Both tests could identify all ASC-US and AGC cases with HSIL + in follow-up biopsies, but AHPV showed a significantly higher specificity in both settings (ASC-US: 28.81% vs 11.86%, p < 0.0001; AGC: 55.00% vs 20.00%, p = 0.0233). CONCLUSIONS: Both AHPV and Cobas were equally sensitive in detecting high-grade SIL in both scenarios of screening and ASC-US or AGC triage but AHPV showed a higher specificity.

Randomized Crossover Study Showing Nurse-Led Same Day Review Replacing Next Day Review in Uneventful Phacoemulsification to Be Safe and Efficacious.

Purpose. To study whether nurse led same-day review (SDR) after uneventful phacoemulsification can replace next-day review (NDR) in terms of safety and efficacy. Setting. Patients are recruited from an ophthalmology outpatient clinic in Hong Kong. Design. A prospective, randomized crossover study conducted from November 2012 to 2014. Methods. Inclusion criteria include cataract surgery naïve patients undergoing phacoemulsification under local anaesthesia. All patients were seen by our ophthalmic nurse 2 hours after surgery. Before undergoing phacoemulsification of the first eye, patients were randomized to be reviewed on day 1 or 7 after surgery. Surgeons and reviewing doctors were blinded to patient allocation. For the patients' second eye surgery, group allocation will cross over. Primary outcome measures include visual improvement and patient satisfaction questionnaire. Other measures include cataract characteristics, surgical details, and complications. Statistical tests include paired t-test, Wilcoxon signed rank test, and Chi-square test. Results. 164 eyes from 82 patients were available. Visual improvement, satisfaction, and complications were comparable between both groups. Conclusions. A nurse led SDR can replace NDR in uneventful phacoemulsification in terms of safety and efficacy. Patient satisfaction is also comparable in the setting of Asian culture and when transportation is not a major concern.

Stem cell transcription factor NANOG in cancers--is eternal youth a curse?

INTRODUCTION: Targeting cancer stem cells can be a more effective approach to treat cancer. NANOG is one of the key factors for maintaining the self-renewal ability and pluripotency of stem cells, including cancer stem cells. Overexpression of NANOG has been observed in various human malignancies. Several reports have suggested that NANOG contributes to carcinogenesis by initiating and preserving cancer stem cells. It is obvious that NANOG is also involved in establishing other hallmarks of cancer such as uncontrolled cell growth, chemoresistance, metastasis, and immune evasion. AREAS COVERED: This review will discuss the molecular properties and oncogenic roles of NANOG. The idea of using agents that inhibit the transcription factor to treat cancer is presented. Interfering with NANOG-mediated transcriptions using small interfering RNA, transcription factor decoy, genome editing, and small-molecule inhibitors may provide novel strategies to target cancer stem cells. EXPERT OPINION: As a pivotal controller in cancer stem cell maintenance and a positive regulator of various oncogenic pathways, NANOG may be an important target for cancer therapy. However, as a transcription factor, it is inherently difficult to target by pharmacological means. Novel approaches need to be explored before the inhibition of NANOG can be applied in a clinical setting.

Overexpression of forkhead box protein M1 (FOXM1) in ovarian cancer correlates with poor patient survival and contributes to paclitaxel resistance.

AIM: Deregulation of FOXM1 has been documented in various cancers. The aim of this study was to evaluate the role of FOXM1 in ovarian cancer tumorigenesis and paclitaxel resistance. EXPERIMENTAL DESIGN: Expression of FOXM1 was examined in 119 clinical samples by immunohistochemistry and correlated with clinicopathological parameters. Effects of FOXM1 knockdown on ovarian cancer cell migration, invasion and mitotic catastrophe were also studied. qPCR and ChIP-qPCR were used to establish KIF2C as a novel FOXM1 target gene implicated in chemoresistance. RESULTS: High nuclear FOXM1 expression in ovarian cancer patient samples was significantly associated with advanced stages (P = 0.035), shorter overall (P = 0.019) and disease-free (P = 0.014) survival. Multivariate analysis confirmed FOXM1 expression as an independent prognostic factor for ovarian cancer. FOXM1 knockdown significantly inhibited migration and invasion of ovarian cancer cells and enhanced paclitaxel-mediated cell death and mitotic catastrophe in a p53-independent manner. Bioinformatics analysis suggested a number of potential transcription targets of FOXM1. One of the potential targets, KIF2C, exhibited similar expression pattern to FOXM1 in chemosensitive and chemoresistant cells in response to paclitaxel treatment. FOXM1 could be detected at the promoter of KIF2C and FOXM1 silencing significantly down-regulated KIF2C. CONCLUSION: Our findings suggest that FOXM1 is associated with poor patient outcome and contributes to paclitaxel resistance by blocking mitotic catastrophe. KIF2C is identified as a novel FOXM1 transcriptional target that may be implicated in the acquisition of chemoresistance. FOXM1 should be further investigated as a potential prognostic marker and therapeutic target for ovarian cancer.

SpliceArray profiling of breast cancer reveals a novel variant of NCOR2/SMRT that is associated with tamoxifen resistance and control of ERα transcriptional activity.

Gene expression profiling aimed at classifying and prognosing breast cancer has yielded signatures with little, if any, concordance. However, expression arrays used in these studies do not discriminate alternate RNA splice isoforms that vary widely in cancer and may resolve this problem. In this study, we profiled splice isoforms in a panel of tamoxifen-sensitive and -resistant cell lines, defining a novel variant (BQ323636.1) of the nuclear receptor corepressor 2 (NCOR2) that was associated with tamoxifen resistance. Overexpression of this variant in a tamoxifen-sensitive cell line induced its resistance to tamoxifen. We confirmed our initial findings from cell lines in 77 breast tumors from a Chinese cohort, where BQ323636.1 expression was higher in tamoxifen-resistant patients than tamoxifen-sensitive patients. For patients who were estrogen receptor (ER)-positive and had received tamoxifen treatment, higher BQ323636.1 expression level correlated with distant metastasis. High expression level of BQ323636.1 was found to be associated with poorer overall and disease-free survival for patients who had received tamoxifen treatment. Notably, higher BQ323636.1 versus NCOR2 wild-type ratio was also associated with negative ER and progesterone receptor (PR) status, and triple-negative status (ER-/PR-/HER2- receptor status). Mechanistic investigations showed that under conditions of tamoxifen exposure, BQ323636.1 suppressed the transcriptional activity of ERα, exhibiting promoter-regulating functions. Our findings highlight a novel splice variant of the ERα corepressor NCOR2 as a candidate biomarker in breast cancer that not only predicts tamoxifen response but may be targeted to overcome tamoxifen resistance.

AMPK activators suppress cervical cancer cell growth through inhibition of DVL3 mediated Wnt/ß-catenin signaling activity.

Recent evidence has suggested that AMPK activators may be applied as therapeutic drugs in suppressing cancer cell growth. However, the molecular mechanism of their suppressive function in cancer cells is still unclear. Here we show that AMPK activators impair cervical cancer cell growth through the reduction of DVL3, a positive regulator in Wnt/ß-catenin signaling and an oncogenic player in cervical cancer tumorigenesis. By western blot and immunohistochemical analyses, we demonstrated that DVL3 was frequently upregulated and significantly associated with elevated ß-catenin (P = 0.009) and CyclinD1 (P = 0.009) expressions in cervical cancer. Enforced expression of DVL3 elevated ß-catenin and augmented cervical cancer cell growth, verifying that DVL3-mediated Wnt/ß-catenin activation is involved in cervical cancer oncogenesis. On the other aspect, we noted that the cervical cancer cell growth was remarkably suppressed by AMPK activators and such cell growth inhibition was in concomitant with the reduction of DVL3 protein level in dose- and time-dependent manners. Besides, impaired mTOR signaling activity also reduced DVL3 expression. In contrast, co-treatment with Compound C (AMPK inhibitor) could significantly abrogate metformin induced DVL3 reduction. In addition, co-treatment with AM114 or MG132 (proteosomal inhibitors) could partially restore DVL3 expression under the treatment of metformin. Further in vivo ubiquitination assay revealed that metformin could reduce DVL3 by ubiquitin/proteasomal degradation. To our knowledge, this is the first report showing the probable molecular mechanisms of that the AMPK activators suppress cervical cancer cell growth by impairing DVL3 protein synthesis via AMPK/mTOR signaling and/or partially promoting the proteasomal degradation of DVL3.

Dysregulated stemness-related genes in gynecological malignancies.

In recent years, much attention has been paid to the concept of cancer stem cells (CSC) and self-renewal related pathways in cancer biology. This review outlines the dysregulated stemness-related genes or transcription factors in gynecological cancers. Hedgehog (Hh) and Notch signaling are important pathways in tissue pattern programming and cell fate determination during embryonic development. Hyperactivation of these two pathways was frequently observed in gynecological malignancies such as ovarian, endometrial and cervical cancers. In contrast, the expression profiles of pluripotency-regulating core transcriptional circuitry: Nanog, Oct4 and Sox2 appear heterogeneous. Among these transcription factors, overexpression of Nanog was found to exert a prominent effect in gynecological tumorigenesis, while dysregulations of Oct4 and Sox2 may vary in a context dependent manner. On the other hand, the isolation of putative CSC illustrates a hierarchy model of tumor heterogeneity, in which only a subset of cells among biologically distinct populations can initiate tumor growth. Re-activation of these pluripotent transcription factors (Nanog, Oct4 and/or Sox2) in association with distinct tumorigenic properties could be found in clones isolated from gynecological tumors using various approaches. Recent understanding on the roles of Hh and Notch signaling in enhancing CSC survival may help to better understand the mechanism of carcinogenesis and identify new pharmaceutical targets for gynecological malignancies.

iASPP and chemoresistance in ovarian cancers: effects on paclitaxel-mediated mitotic catastrophe.

PURPOSE: iASPP is a specific regulator of p53-mediated apoptosis. Herein, we provided the first report on the expression profile of iASPP in ovarian epithelial tumor and its effect on paclitaxel chemosensitivity. EXPERIMENTAL DESIGN: Expression and amplification status of iASPP was examined in 203 clinical samples and 17 cell lines using immunohistochemistry, quantitative real-time PCR, and immunoblotting, and correlated with clinicopathologic parameters. Changes in proliferation, mitotic catastrophe, apoptosis, and underlying mechanism in ovarian cancer cells of different p53 status following paclitaxel exposure were also analyzed. RESULTS: The protein and mRNA expression of iASPP was found to be significantly increased in ovarian cancer samples and cell lines. High iASPP expression was significantly associated with clear cell carcinoma subtype (P = 0.003), carboplatin and paclitaxel chemoresistance (P = 0.04), shorter overall (P = 0.003), and disease-free (P = 0.001) survival. Multivariate analysis confirmed iASPP expression as an independent prognostic factor. Increased iASPP mRNA expression was significantly correlated with gene amplification (P = 0.023). iASPP overexpression in ovarian cancer cells conferred resistance to paclitaxel by reducing mitotic catastrophe in a p53-independent manner via activation of separase, whereas knockdown of iASPP enhanced paclitaxel-mediated mitotic catastrophe through inactivating separase. Both securin and cyclin B1/CDK1 complex were involved in regulating separase by iASPP. Conversely, overexpressed iASPP inhibited apoptosis in a p53-dependent mode. CONCLUSIONS: Our data show an association of iASPP overexpression with gene amplification in ovarian cancer and suggest a role of iASPP in poor patient outcome and chemoresistance, through blocking mitotic catastrophe. iASPP should be explored further as a potential prognostic marker and target for chemotherapy.

TrkB as a therapeutic target for ovarian cancer.

BACKGROUND: In many countries, ovarian cancer is the most lethal gynecological malignancy. Its poor prognosis is mainly due to the late stage of disease with metastasis at presentation. The significant failure rate of chemotherapy in patients with advanced stage disease is also a main concern. As such, developing novel therapeutic targets is essential to improve long-term survival. Overexpression of Tropomyosin-related kinase B (TrkB), a tyrosine kinase receptor, has been documented in ovarian cancer and is found to be correlated with poor prognosis. OBJECTIVE/METHODS: We discuss the functional roles and the related downstream signaling pathways of TrkB and its ligand brain-derived neurotrophic factor (BDNF) in ovarian cancer. The possible crosstalk between TrkB/BDNF and other putative molecular targets in ovarian cancer is also discussed. RESULTS/CONCLUSIONS: All these latest findings shed light on the application of TrkB as a therapeutic target for ovarian cancer.