Dynamics of transforming growth factor ß signaling and therapeutic efficacy.

Transforming growth factor ß (TGFß) is a multifunctional cytokine, and its signalling responses are exerted via integrated intracellular pathways and complex regulatory mechanisms. Due to its high potency, TGFß signalling is tightly controlled under normal circumstances, while its dysregulation in cancer favours metastasis. The recognised potential of TGFß as a therapeutic target led to emerging development of anti-TGFß reagents with preclinical success, yet these therapeutics failed to recapitulate their efficacy in experimental settings. In this review, possible reasons for this inconsistency are discussed, addressing the knowledge gap between theoretical and actual behaviours of TGFß signalling. Previous studies on oncogenic cells have demonstrated the spatiotemporal heterogeneity of TGFß signalling intensity. Under feedback mechanisms and exosomal ligand recycling, cancer cells may achieve cyclic TGFß signalling to facilitate dissemination and colonisation. This challenges the current presumption of persistently high TGFß signalling in cancer, pointing to a new direction of research on TGFß-targeted therapeutics.

The Interleukin-11/IL-11 Receptor Promotes Glioblastoma Survival and Invasion under Glucose-Starved Conditions through Enhanced Glutaminolysis.

Glioblastoma cells adapt to changes in glucose availability through metabolic plasticity allowing for cell survival and continued progression in low-glucose concentrations. However, the regulatory cytokine networks that govern the ability to survive in glucose-starved conditions are not fully defined. In the present study, we define a critical role for the IL-11/IL-11Rα signalling axis in glioblastoma survival, proliferation and invasion when cells are starved of glucose. We identified enhanced IL-11/IL-11Rα expression correlated with reduced overall survival in glioblastoma patients. Glioblastoma cell lines over-expressing IL-11Rα displayed greater survival, proliferation, migration and invasion in glucose-free conditions compared to their low-IL-11Rα-expressing counterparts, while knockdown of IL-11Rα reversed these pro-tumorigenic characteristics. In addition, these IL-11Rα-over-expressing cells displayed enhanced glutamine oxidation and glutamate production compared to their low-IL-11Rα-expressing counterparts, while knockdown of IL-11Rα or the pharmacological inhibition of several members of the glutaminolysis pathway resulted in reduced survival (enhanced apoptosis) and reduced migration and invasion. Furthermore, IL-11Rα expression in glioblastoma patient samples correlated with enhanced gene expression of the glutaminolysis pathway genes GLUD1, GSS and c-Myc. Overall, our study identified that the IL-11/IL-11Rα pathway promotes glioblastoma cell survival and enhances cell migration and invasion in environments of glucose starvation via glutaminolysis.

Cortical tension initiates the positive feedback loop between cadherin and F-actin.

Adherens junctions physically link two cells at their contact interface via extracellular binding between cadherin molecules and intracellular interactions between cadherins and the actin cytoskeleton. Cadherin and actomyosin cytoskeletal dynamics are regulated reciprocally by mechanical and chemical signals, which subsequently determine the strength of cell-cell adhesions and the emergent organization and stiffness of the tissues they form. However, an understanding of the integrated system is lacking. We present a new mechanistic computational model of intercellular junction maturation in a cell doublet to investigate the mechanochemical cross talk that regulates adherens junction formation and homeostasis. The model couples a two-dimensional lattice-based simulation of cadherin dynamics with a reaction-diffusion representation of the reorganising actomyosin network through its regulation by Rho signalling at the intracellular junction. We demonstrate that local immobilization of cadherin induces cluster formation in a cis-less-dependent manner. We then recapitulate the process of cell-cell contact formation. Our model suggests that cortical tension applied on the contact rim can explain the ring distribution of cadherin and actin filaments (F-actin) on the cell-cell contact of the cell doublet. Furthermore, we propose and test the hypothesis that cadherin and F-actin interact like a positive feedback loop, which is necessary for formation of the ring structure. Different patterns of cadherin distribution were observed as an emergent property of disturbances of this positive feedback loop. We discuss these findings in light of available experimental observations on underlying mechanisms related to cadherin/F-actin binding and the mechanical environment.

Blocking Muscarinic Receptor 3 Attenuates Tumor Growth and Decreases Immunosuppressive and Cholinergic Markers in an Orthotopic Mouse Model of Colorectal Cancer.

Tumor cells have evolved to express immunosuppressive molecules allowing their evasion from the host's immune system. These molecules include programmed death ligands 1 and 2 (PD-L1 and PD-L2). Cancer cells can also produce acetylcholine (ACh), which plays a role in tumor development. Moreover, tumor innervation can stimulate vascularization leading to tumor growth and metastasis. The effects of atropine and muscarinic receptor 3 (M3R) blocker, 1,1-dimethyl-4-diphenylacetoxypiperidinium iodide (4-DAMP), on cancer growth and spread were evaluated in vitro using murine colon cancer cell line, CT-26, and in vivo in an orthotopic mouse model of colorectal cancer. In the in vitro model, atropine and 4-DAMP significantly inhibited CT-26 cell proliferation in a dose dependent manner and induced apoptosis. Atropine attenuated immunosuppressive markers and M3R via inhibition of EGFR/AKT/ERK signaling pathways. However, 4-DAMP showed no effect on the expression of PD-L1, PD-L2, and choline acetyltransferase (ChAT) on CT-26 cells but attenuated M3R by suppressing the phosphorylation of AKT and ERK. Blocking of M3R in vivo decreased tumor growth and expression of immunosuppressive, cholinergic, and angiogenic markers through inhibition of AKT and ERK, leading to an improved immune response against cancer. The expression of immunosuppressive and cholinergic markers may hold potential in determining prognosis and treatment regimens for colorectal cancer patients. This study's results demonstrate that blocking M3R has pronounced antitumor effects via several mechanisms, including inhibition of immunosuppressive molecules, enhancement of antitumor immune response, and suppression of tumor angiogenesis via suppression of the AKT/ERK signaling pathway. These findings suggest a crosstalk between the cholinergic and immune systems during cancer development. In addition, the cholinergic system influences cancer evasion from the host's immunity.

Extracellular Vesicles Secreted by Glioma Stem Cells Are Involved in Radiation Resistance and Glioma Progression.

Glioblastoma is the most aggressive brain tumour with short survival, partly due to resistance to conventional therapy. Glioma stem cells (GSC) are likely to be involved in treatment resistance, by releasing extracellular vesicles (EVs) containing specific molecular cargoes. Here, we studied the EVs secreted by glioma stem cells (GSC-EVs) and their effects on radiation resistance and glioma progression. EVs were isolated from 3 GSCs by serial centrifugation. NanoSight measurement, cryo-electron microscopy and live imaging were used to study the EVs size, morphology and uptake, respectively. The non-GSC glioma cell lines LN229 and U118 were utilised as a recipient cell model. Wound healing assays were performed to detect cell migration. Colony formation, cell viability and invadopodium assays were conducted to detect cell survival of irradiated recipient cells and cell invasion post GSC-EV treatment. NanoString miRNA global profiling was used to select for the GSC-EVs' specific miRNAs. All three GSC cell lines secreted different amounts of EVs, and all expressed consistent levels of CD9 but different level of Alix, TSG101 and CD81. EVs were taken up by both LN229 and U118 recipient cells. In the presence of GSC-EVs, these recipient cells survived radiation exposure and initiated colony formation. After GSC-EVs exposure, LN229 and U118 cells exhibited an invasive phenotype, as indicated by an increase in cell migration. We also identified 25 highly expressed miRNAs in the GSC-EVs examined, and 8 of these miRNAs can target PTEN. It is likely that GSC-EVs and their specific miRNAs induced the phenotypic changes in the recipient cells due to the activation of the PTEN/Akt pathway. This study demonstrated that GSC-EVs have the potential to induce radiation resistance and modulate the tumour microenvironment to promote glioma progression. Future therapeutic studies should be designed to interfere with these GSC-EVs and their specific miRNAs.

Elucidating the role of long intergenic non-coding RNA 339 in human endometrium and endometriosis.

Endometriosis is a complex disease, influenced by genetic factors. Genetic markers associated with endometriosis exist at chromosome 1p36.12 and lead to altered expression of the long intergenic non-coding RNA 339 (LINC00339), however, the role of LINC00339 in endometriosis pathophysiology remains unknown. The aim of this work was to characterize the expression patterns of LINC00339 mRNA in endometrium and endometriotic lesions in situ and to determine the functional role of LINC00339 in human endometrium. We employed RNA-sequencing (RNA-seq), quantitative RT-PCR and in situ hybridization to investigate the abundance of LINC00339 transcripts in endometrium and endometrial cell lines and to describe the pattern and localization of LINC00339 expression in endometrium and endometriotic lesions. LINC00339 mRNA expression was manipulated (overexpressed and silenced) in endometrial stromal cell lines and RNA-seq data from overexpression models were analysed using online bioinformatics platforms (STRING and Ingenuity Pathway Analysis) to determine functional processes. We demonstrated the expression of LINC00339 in endometriotic lesions for the first time; we found LINC00339 expression was restricted to the lesion foci and absent in surrounding non-lesion tissue. Furthermore, manipulation of LINC00339 expression in endometrial stromal cell lines significantly impacted the expression of genes involved in immune defence pathways. These studies identify a novel mechanism for LINC00339 activity in endometrium and endometriosis, paving the way for future work, which is essential for understanding the pathogenesis of endometriosis.

Carfilzomib Promotes the Unfolded Protein Response and Apoptosis in Cetuximab-Resistant Colorectal Cancer.

Cetuximab is a common treatment option for patients with wild-type K-Ras colorectal carcinoma. However, patients often display intrinsic resistance or acquire resistance to cetuximab following treatment. Here we generate two human CRC cells with acquired resistance to cetuximab that are derived from cetuximab-sensitive parental cell lines. These cetuximab-resistant cells display greater in vitro proliferation, colony formation and migration, and in vivo tumour growth compared with their parental counterparts. To evaluate potential alternative therapeutics to cetuximab-acquired-resistant cells, we tested the efficacy of 38 current FDA-approved agents against our cetuximab-acquired-resistant clones. We identified carfilzomib, a selective proteosome inhibitor to be most effective against our cell lines. Carfilzomib displayed potent antiproliferative effects, induced the unfolded protein response as determined by enhanced CHOP expression and ATF6 activity, and enhanced apoptosis as determined by enhanced caspase-3/7 activity. Overall, our results indicate a potentially novel indication for carfilzomib: that of a potential alternative agent to treat cetuximab-resistant colorectal cancer.

Antitumor and Antiangiogenic Properties of Gold(III) Complexes Containing Cycloaurated Triphenylphosphine Sulfide Ligands.

A family of stable anticancer gold(III)-based therapeutic complexes containing cyclometalated triphenylphosphine sulfide ligands have been prepared. The anticancer properties of the newly developed complexes [AuCl2{κ2-2-C6H4P(S)Ph2}] (1), [Au(κ2-S2CNEt2){κ2-2-C6H4P(S)Ph2}]PF6 (2), [AuCl(dppe){κC-2-C6H4P(S)Ph2}]Cl (3), and [Au(dppe){κ2-2-C6H4P(S)Ph2}][PF6]2 (4) were investigated toward five human cancer cell lines [cervical (HeLa), lung (A549), prostate (PC3), fibrosarcoma (HT1080), and breast (MDA-MB-231)]. In vitro cytotoxicity studies revealed that compounds 2-4 displayed potent cell growth inhibition (IC50 values in the range of 0.17-2.50 µM), comparable to, or better than, clinically used cisplatin (0.63-6.35 µM). Preliminary mechanistic studies using HeLa cells indicate that the cytotoxic effects of the compounds involve apoptosis induction through ROS accumulation. Compound 2 also demonstrated significant inhibition of endothelial cell migration and tube formation in the angiogenesis process. Evaluation of the in vivo antitumor activity of compound 2 in nude mice bearing cervical cancer cell (HeLa) xenografts indicated significant tumor growth inhibition (55%) with 1 mg/kg dose (every 3 days) compared with the same dose of cisplatin (28%). These results demonstrate the potential of gold(III) complexes containing cyclometalated triphenylphosphine sulfide ligands as novel metal-based anticancer agents.

Cell quiescence correlates with enhanced glioblastoma cell invasion and cytotoxic resistance.

Glioblastoma (GBM) tumor cells exhibit drug resistance and are highly infiltrative. GBM stem cells (GSCs), which have low proliferative capacity are thought to be one of the sources of resistant cells which result in relapse/recurrence. However, the molecular mechanisms regulating quiescent-specific tumor cell biology are not well understood. Using human GBM cell lines and patient-derived GBM cells, Oregon Green dye retention was used to identify and isolate the slow-cycling, quiescent-like cell subpopulation from the more proliferative cells in culture. Sensitivity of cell subpopulations to temozolomide and radiation, as well as the migration and invasive potential were measured. Differential expression analysis following RNAseq identified genes enriched in the quiescent cell subpopulation. Orthotopic transplantation of cells into mice was used to compare the in vivo malignancy and invasive capacity of the cells. Proliferative quiescence correlated with better TMZ resistance and enhanced cell invasion, in vitro and in vivo. RNAseq expression analysis identified genes involved in the regulation cell invasion/migration and a three-gene signature, TGFBI, IGFBP3, CHI3L1, overexpressed in quiescent cells which correlates with poor GBM patient survival.

Novel additive for sperm cryopreservation media: Holotheria parva coelomic cavity extract protects human spermatozoa against oxidative stress-A pilot study.

Cryopreservation is the most effective method for preserving semen for a long period of time. However, during the freeze-thaw process, production of reactive oxygen species (ROS) leads to a steep reduction in sperm fertility indices. In this study, we tested the effects of the extract of the coelomic cavity of five Holotheria parva, a marine organism rich in antioxidants, for its ROS-scavenging activity and cryoprotective effects on oxidative stress. Using a total of 50 semen samples, our results demonstrated that doses of 250 and 500 µg/ml of H. parva coelomic cavity extract significantly increased sperm vitality as compared to the control (p < .05). The addition of 250 µg/ml of the extract exerted a significant positive effect on sperm motility. Moreover, sperm DNA damage and ROS production were significantly reduced at extract concentrations of 250 and 500 µg/ml (p < .05). To the best of our knowledge, the results of this study represent the first demonstration of the possibility of improving sperm parameters and reducing ROS production and DNA damage by supplementing sperm freezing media with H. parva coelomic extract. Our results suggested that H. parva coelomic extract could be useful for improving the fertilising ability of frozen-thawed human semen.

Targeting Glioma Stem Cells by Functional Inhibition of Dynamin 2: A Novel Treatment Strategy for Glioblastoma.

Glioma stem cells (GSCs) play major roles in drug resistance, tumour maintenance and recurrence of glioblastoma. We investigated inhibition of the GTPase dynamin 2 as a therapy for glioblastoma. Glioma cell lines and patient-derived GSCs were treated with dynamin inhibitors, Dynole 34-2 and CyDyn 4-36. We studied about cell viability, and GSC neurosphere formation in vitro and orthotopic tumour growth in vivo. Dynamin inhibition reduced glioblastoma cell line viability and suppressed neurosphere formation and migration of GSCs. Tumour growth was reduced by CyDyn 4-36 treatment. Dynamin 2 inhibition therefore represents a novel approach for stem cell-directed Glioblastoma therapy.

Potent and Selective Cytotoxic and Anti-inflammatory Gold(III) Compounds Containing Cyclometalated Phosphine Sulfide Ligands.

Four cycloaurated phosphine sulfide complexes, [Au{κ2 -2-C6 H4 P(S)Ph2 }2 ][AuX2 ] [X=Cl (2), Br (3), I (4)] and [Au{κ2 -2-C6 H4 P(S)Ph2 }2 ]PF6 (5), have been prepared and thoroughly characterized. The compounds were found to be stable under physiological-like conditions and showed excellent cytotoxicity against a broad range of cancer cell lines and remarkable cytotoxicity in 3D tumor spheroids. Mechanistic studies with cervical cancer (HeLa) cells indicated that the cytotoxic effects of the compounds involve the inhibition of thioredoxin reductase and induction of apoptosis through mitochondrial disruption. In vivo experiments in nude mice bearing HeLa xenografts showed that treatment with compounds 4 and 5 resulted in significant inhibition of tumor growth (35.8 and 46.9 %, respectively), better than that of cisplatin (29 %). The newly synthesized gold complexes were also evaluated for their in vitro and in vivo anti-inflammatory activity through the study of lipopolysaccharide (LPS)-activated macrophages and carrageenan-induced hind paw edema in rats, respectively.

Epidermal growth factor receptor: Structure-function informing the design of anticancer therapeutics.

Research on the epidermal growth factor (EGF) family and the family of receptors (EGFR) has progressed rapidly in recent times. New crystal structures of the ectodomains with different ligands, the activation of the kinase domain through oligomerisation and the use of fluorescence techniques have revealed profound conformational changes on ligand binding. The control of cell signaling from the EGFR-family is complex, with heterodimerisation, ligand affinity and signaling cross-talk influencing cellular outcomes. Analysis of tissue homeostasis indicates that the control of pro-ligand processing is likely to be as important as receptor activation events. Several members of the EGFR-family are overexpressed and/or mutated in cancer cells. The perturbation of EGFR-family signaling drives the malignant phenotype of many cancers and both inhibitors and antagonists of signaling from these receptors have already produced therapeutic benefits for patients. The design of affibodies, antibodies, small molecule inhibitors and even immunotherapeutic drugs targeting the EGFR-family has yielded promising new approaches to improving outcomes for cancer patients. In this review, we describe recent discoveries which have increased our understanding of the structure and dynamics of signaling from the EGFR-family, the roles of ligand processing and receptor cross-talk. We discuss the relevance of these studies to the development of strategies for designing more effective targeted treatments for cancer patients.

Effects of EphB4 receptor expression on colorectal cancer cells, tumor growth, vascularization and composition.

BACKGROUND: Colorectal cancer (CRC) is one of the most common causes of cancer-related deaths worldwide. Increased expression of the molecular target, EphB4 receptor, has been observed in several cancer types. However, studies on the role of EphB4 receptor in CRC have yielded contradictory results. The aim of this study was to investigate the influence of EphB4 expression levels on CRC cell behavior and its contribution to tumor growth and vascularization. METHODS: SW480, LIM2405 and CT26 CRC cell lines were transfected with EphB4 expression vector. High EphB4 expressing cells were compared to low EphB4 expressing empty vector controls. Proliferation and migration assays as well as EphrinB2-Fc cell stimulations were conducted in vitro and subcutaneous xenografts of CRC were analyzed in vivo. RESULTS: High EphB4 expression enhanced migratory ability of these CRC cell lines in vitro and contributed to a significant increase in tumor growth and vascularization in vivo. Tumours induced with high EphB4 expressing SW480 and LIM2405 cells yielded homogenous masses densely packed with cancer cells. EphrinB2-Fc cell stimulations induced cell clustering of high EphB4 expressing SW480 and LIM2405 in vitro. CONCLUSION: These results suggest that with enhanced vascularization and an increase in migratory abilities, the high EphB4 expressing cells may be able to metastasize more readily.

Defining the essential function of FBP/KSRP proteins: Drosophila Psi interacts with the mediator complex to modulate MYC transcription and tissue growth.

Despite two decades of research, the major function of FBP-family KH domain proteins during animal development remains controversial. The literature is divided between RNA processing and transcriptional functions for these single stranded nucleic acid binding proteins. Using Drosophila, where the three mammalian FBP proteins (FBP1-3) are represented by one ortholog, Psi, we demonstrate the primary developmental role is control of cell and tissue growth. Co-IP-mass spectrometry positioned Psi in an interactome predominantly comprised of RNA Polymerase II (RNA Pol II) transcriptional machinery and we demonstrate Psi is a potent transcriptional activator. The most striking interaction was between Psi and the transcriptional mediator (MED) complex, a known sensor of signaling inputs. Moreover, genetic manipulation of MED activity modified Psi-dependent growth, which suggests Psi interacts with MED to integrate developmental growth signals. Our data suggest the key target of the Psi/MED network in controlling developmentally regulated tissue growth is the transcription factor MYC. As FBP1 has been implicated in controlling expression of the MYC oncogene, we predict interaction between MED and FBP1 might also have implications for cancer initiation and progression.

SPSB1, a Novel Negative Regulator of the Transforming Growth Factor-ß Signaling Pathway Targeting the Type II Receptor.

Appropriate cellular signaling is essential to control cell proliferation, differentiation, and cell death. Aberrant signaling can have devastating consequences and lead to disease states, including cancer. The transforming growth factor-ß (TGF-ß) signaling pathway is a prominent signaling pathway that has been tightly regulated in normal cells, whereas its deregulation strongly correlates with the progression of human cancers. The regulation of the TGF-ß signaling pathway involves a variety of physiological regulators. Many of these molecules act to alter the activity of Smad proteins. In contrast, the number of molecules known to affect the TGF-ß signaling pathway at the receptor level is relatively low, and there are no known direct modulators for the TGF-ß type II receptor (TßRII). Here we identify SPSB1 (a Spry domain-containing Socs box protein) as a novel regulator of the TGF-ß signaling pathway. SPSB1 negatively regulates the TGF-ß signaling pathway through its interaction with both endogenous and overexpressed TßRII (and not TßRI) via its Spry domain. As such, TßRII and SPSB1 co-localize on the cell membrane. SPSB1 maintains TßRII at a low level by enhancing the ubiquitination levels and degradation rates of TßRII through its Socs box. More importantly, silencing SPSB1 by siRNA results in enhanced TGF-ß signaling and migration and invasion of tumor cells.

Coalition of Oct4A and ß1 integrins in facilitating metastasis in ovarian cancer.

BACKGROUND: Ovarian cancer is a metastatic disease and one of the leading causes of gynaecology malignancy-related deaths in women. Cancer stem cells (CSCs) are key contributors of cancer metastasis and relapse. Integrins are a family of cell surface receptors which allow interactions between cells and their surrounding microenvironment and play a fundamental role in promoting metastasis. This study investigates the molecular mechanism which associates CSCs and integrins in ovarian cancer metastasis. METHODS: The expression of Oct4A in high-grade serous ovarian tumors and normal ovaries was determined by immunofluorescence analysis. The functional role of Oct4A was evaluated by generating stable knockdown (KD) of Oct4A clones in an established ovarian cancer cell line HEY using shRNA-mediated silencing. The expression of integrins in cell lines was evaluated by flow cytometry. Spheroid forming ability, adhesion and the activities of matrix metalloproteinases 9/2 (MMP-9/2) was measured by in vitro functional assays and gelatin zymography. These observations were further validated in in vivo mouse models using Balb/c nu/nu mice. RESULTS: We report significantly elevated expression of Oct4A in high-grade serous ovarian tumors compared to normal ovarian tissues. The expression of Oct4A in ovarian cancer cell lines correlated with their CSC-related sphere forming abilities. The suppression of Oct4A in HEY cells resulted in a significant diminution of integrin ß1 expression and associated α5 and α2 subunits compared to vector control cells. This was associated with a reduced adhesive ability on collagen and fibronectin and decreased secretion of pro-MMP2 in Oct4A KD cells compared to vector control cells. In vivo, Oct4A knock down (KD) cells produced tumors which were significantly smaller in size and weight compared to tumors derived from vector control cells. Immunohistochemical analyses of Oct4A KD tumor xenografts demonstrated a significant loss of cytokeratin 7 (CK7), Glut-1 as well as CD34 and CD31 compared to vector control cell-derived xenografts. CONCLUSION: The expression of Oct4A may be crucial to promote and sustain integrin-mediated extracellular matrix (ECM) remodeling requisite for tumor metastasis in ovarian cancer patients.

A critical role of Oct4A in mediating metastasis and disease-free survival in a mouse model of ovarian cancer.

BACKGROUND: High grade epithelial ovarian cancer (EOC) is commonly characterised by widespread peritoneal dissemination and ascites. Metastatic EOC tumour cells can attach directly to neighbouring organs or alternatively, maintain long term tumourigenicity and chemoresistance by forming cellular aggregates (spheroids). Cancer stem-like cells are proposed to facilitate this mechanism. This study aimed to investigate the role of Oct4A, an embryonic stem cell factor and known master regulator of pluripotency in EOC progression, metastasis and chemoresistance. METHODS: To investigate the expression of Oct4A in primary EOC tumours, IHC and qRT-PCR analyses were used. The expression of Oct4A in chemonaive and recurrent EOC patient ascites-derived tumour cells samples was investigated by qRT-PCR. The functional role of Oct4A in EOC was evaluated by generating stable knockdown Oct4A clones in the established EOC cell line HEY using shRNA-mediated silencing technology. Cellular proliferation, spheroid forming ability, migration and chemosensitivty following loss of Oct4A in HEY cells was measured by in vitro functional assays. These observations were further validated in an in vivo mouse model using intraperitoneal (IP) injection of established Oct4A KD clones into Balb/c nu/nu mice. RESULTS: We demonstrate that, compared to normal ovaries Oct4A expression significantly increases with tumour dedifferentiation. Oct4A expression was also significantly high in the ascites-derived tumour cells of recurrent EOC patients compared to chemonaive patients. Silencing of Oct4A in HEY cells resulted in decreased cellular proliferation, migration, spheroid formation and increased chemosensitivity to cisplatin in vitro. IP injection of Oct4A knockdown cells in vivo produced significantly reduced tumour burden, tumour size and invasiveness in mice, which overall resulted in significantly increased mouse survival rates compared to mice injected with control cells. CONCLUSIONS: This data highlights a crucial role for Oct4A in the progression and metastasis of EOC. Targeting Oct4A may prove to be an effective strategy in the treatment and management of epithelial ovarian tumours.

Single live cell TGF-ß signalling imaging: breast cancer cell motility and migration is driven by sub-populations of cells with dynamic TGF-ß-Smad3 activity.

BACKGROUND: Metastasis is a process where only a small subset of cells is capable of successfully migrating to and propagating at secondary sites. TGF-ß signalling is widely known for its role in cancer metastasis and is associated with cell migration in whole cell populations. FINDINGS: We extend these findings by investigating the role of TGF-ß signalling in promoting migration and motility by imaging the signalling activity in live, individual MDA-MB-231 cancer cells utilizing a novel Smad3 Td-Tomato reporter adenovirus. Here we find that not all MDA-MB-231 cancer cells have similar TGF-ß mediated Smad3 transcription activity and display at least two distinct migratory populations. Importantly, Smad3 activity was significantly higher within migratory cells compared to non-migrated cells in wound healing and transwell assays. Furthermore, time-lapse experiments showed that MDA-MB-231 cells displaying Smad3 activity moved faster and a greater distance compared to cells not displaying Smad3 reporter activity. Interestingly, despite being more motile than cells with undetectable levels of Smad3 activity, high Smad3 activity was detrimental to cell motility compared to low and medium level of Smad3 activity. CONCLUSIONS: We have developed a method enabling real-time visualization of TGF-ß signalling in single live cells. Breast cancer cell motility and migration is driven by sub-populations of cells with dynamic TGF-ß-Smad3 activity. Those sub-populations may be responsible for tumor invasion and metastasis.

MicroRNA as potential biomarkers in Glioblastoma.

Glioblastoma is the most aggressive and lethal tumour of the central nervous system and as such the identification of reliable prognostic and predictive biomarkers for patient survival and tumour recurrence is paramount. MicroRNA detection has rapidly emerged as potential biomarkers, in patients with glioblastoma. Over the last decade, analysis of miRNA in laboratory based studies have yielded several candidates as potential biomarkers however, the accepted use of these candidates in the clinic is yet to be validated. Here we will examine the use of miRNA signatures to improve glioblastoma stratification into subgroups and summarise recent advances made in miRNA examination as potential biomarkers for glioblastoma progression and recurrence.