Investigating the role of CBX2 in ER- positive breast cancer

Breast cancer is the most common form of cancer in women, with oestrogen receptor (ER) positive breast cancers being the most common subtype. Although there are targeted endocrine therapies for this receptor, resistance mechanisms mean that they are not always effective. Post-translational modifications of histone proteins are important for regulating gene expression. It is known that the pattern of modifications is different in cancerous tissue compared to normal tissue. Epigenetic reader proteins recognise histone post-translational modifications and help remodel the adjacent chromatin landscape, resulting in gene expression or repression. This means epigenetic reader proteins are possible novel therapeutic targets. CBX2 is an example of an epigenetic reader protein which is overexpressed in ER-positive breast cancer. The aim of this study was to analyse the role of CBX2 in ER-positive breast cancer. This was assessed by knocking down CBX2 gene and protein expression, using siRNAs in MCF-7 and T47D cell lines and analysing changes to cellular phenotype and gene expression regulation. It was determined that CBX2 in the breast cancer cell lines was a nuclear activated phosphorylated form of the protein, and that knockdown of CBX2 has little effect on H2AK119ub and H3K27me3, but causes decrease of H3K27ac. Phenotypic experiments analysed the effect of CBX2 on cell growth, using MTS and apoptosis assays, and showed that CBX2 knockdown reduced cell number and increased cell death. RNA- Sequencing analysis identified that CBX2 has a role in regulating genes within the cell cycle and ER-signalling pathway. The effect on ER-target genes was validated by quantitative-PCR. Additional investigation of the RNA-Seq data will further validate the role of CBX2 in ER-positive breast cancer. Continued research is important for developing therapies for the future treatment of this cancer

The Epigenetic Regulatory Protein CBX2 Promotes mTORC1 Signalling and Inhibits DREAM Complex Activity to Drive Breast Cancer Cell Growth

Chromobox 2 (CBX2) is a chromatin-binding component of polycomb repressive complex 1, which causes gene silencing. CBX2 expression is elevated in triple-negative breast cancer (TNBC), for which there are few therapeutic options. Here, we aimed to investigate the functional role of CBX2 in TNBC. CBX2 knockdown in TNBC models reduced cell numbers, which was rescued by ectopic expression of wild-type CBX2 but not a chromatin binding-deficient mutant. Blocking CBX2 chromatin interactions using the inhibitor SW2_152F also reduced cell growth, suggesting CBX2 chromatin binding is crucial for TNBC progression. RNA sequencing and gene set enrichment analysis of CBX2-depleted cells identified downregulation of oncogenic signalling pathways, including mTORC1 and E2F signalling. Subsequent analysis identified that CBX2 represses the expression of mTORC1 inhibitors and the tumour suppressor RBL2. RBL2 repression, in turn, inhibits DREAM complex activity. The DREAM complex inhibits E2F signalling, causing cell senescence; therefore, inhibition of the DREAM complex via CBX2 may be a key oncogenic driver. We observed similar effects in oestrogen receptor-positive breast cancer, and analysis of patient datasets suggested CBX2 inhibits RBL2 activity in other cancer types. Therapeutic inhibition of CBX2 could therefore repress mTORC1 activation and promote DREAM complex-mediated senescence in TNBC and could have similar effects in other cancer types

The lncRNA CASC15 regulates SOX4 expression in RUNX1-rearranged acute leukemia

Abstract Background Long non-coding RNAs (lncRNAs) play a variety of cellular roles, including regulation of transcription and translation, leading to alterations in gene expression. Some lncRNAs modulate the expression of chromosomally adjacent genes. Here, we assess the roles of the lncRNA CASC15 in regulation of a chromosomally nearby gene, SOX4, and its function in RUNX1/AML translocated leukemia. Results CASC15 is a conserved lncRNA that was upregulated in pediatric B-acute lymphoblastic leukemia (B-ALL) with t (12; 21) as well as pediatric acute myeloid leukemia (AML) with t (8; 21), both of which are associated with relatively better prognosis. Enforced expression of CASC15 led to a myeloid bias in development, and overall, decreased engraftment and colony formation. At the cellular level, CASC15 regulated cellular survival, proliferation, and the expression of its chromosomally adjacent gene, SOX4. Differentially regulated genes following CASC15 knockdown were enriched for predicted transcriptional targets of the Yin and Yang-1 (YY1) transcription factor. Interestingly, we found that CASC15 enhances YY1-mediated regulation of the SOX4 promoter. Conclusions Our findings represent the first characterization of this CASC15 in RUNX1-translocated leukemia, and point towards a mechanistic basis for its action

The Long Non-Coding RNA H19 Drives the Proliferation of Diffuse Intrinsic Pontine Glioma with H3K27 Mutation

Diffuse intrinsic pontine glioma (DIPG) is an incurable paediatric malignancy. Identifying the molecular drivers of DIPG progression is of the utmost importance. Long non-coding RNAs (lncRNAs) represent a large family of disease- and tissue-specific transcripts, whose functions have not yet been elucidated in DIPG. Herein, we studied the oncogenic role of the development-associated H19 lncRNA in DIPG. Bioinformatic analyses of clinical datasets were used to measure the expression of H19 lncRNA in paediatric high-grade gliomas (pedHGGs). The expression and sub-cellular location of H19 lncRNA were validated in DIPG cell lines. Locked nucleic acid antisense oligonucleotides were designed to test the function of H19 in DIPG cells. We found that H19 expression was higher in DIPG vs. normal brain tissue and other pedHGGs. H19 knockdown resulted in decreased cell proliferation and survival in DIPG cells. Mechanistically, H19 buffers let-7 microRNAs, resulting in the up-regulation of oncogenic let-7 target (e.g., SULF2 and OSMR). H19 is the first functionally characterized lncRNA in DIPG and a promising therapeutic candidate for treating this incurable cancer

Public Sector Poetry Journal: Telling Stories about Health, Education and Society

A collection of ethnographic poems by public sector workers across the UK, using poetry to articulate their lived experiences within the sector. Issue 2 of the magazine is a collection of 15 selected poems from a submission of 187 poems; all poems will be used as qualitative data and explored using thematic analysis and findings will support recommendations to advocate change and improvement in the sector

Obeticholic acid for the treatment of non-alcoholic steatohepatitis: interim analysis from a multicentre, randomised, placebo-controlled phase 3 trial

Background Non-alcoholic steatohepatitis (NASH) is a common type of chronic liver disease that can lead to cirrhosis. Obeticholic acid, a farnesoid X receptor agonist, has been shown to improve the histological features of NASH. Here we report results from a planned interim analysis of an ongoing, phase 3 study of obeticholic acid for NASH. Methods In this multicentre, randomised, double-blind, placebo-controlled study, adult patients with definite NASH,non-alcoholic fatty liver disease (NAFLD) activity score of at least 4, and fibrosis stages F2–F3, or F1 with at least oneaccompanying comorbidity, were randomly assigned using an interactive web response system in a 1:1:1 ratio to receive oral placebo, obeticholic acid 10 mg, or obeticholic acid 25 mg daily. Patients were excluded if cirrhosis, other chronic liver disease, elevated alcohol consumption, or confounding conditions were present. The primary endpointsfor the month-18 interim analysis were fibrosis improvement (≥1 stage) with no worsening of NASH, or NASH resolution with no worsening of fibrosis, with the study considered successful if either primary endpoint was met. Primary analyses were done by intention to treat, in patients with fibrosis stage F2–F3 who received at least one dose of treatment and reached, or would have reached, the month 18 visit by the prespecified interim analysis cutoff date. The study also evaluated other histological and biochemical markers of NASH and fibrosis, and safety. This study is ongoing, and registered with ClinicalTrials.gov, NCT02548351, and EudraCT, 20150-025601-6. Findings Between Dec 9, 2015, and Oct 26, 2018, 1968 patients with stage F1–F3 fibrosis were enrolled and received at least one dose of study treatment; 931 patients with stage F2–F3 fibrosis were included in the primary analysis (311 in the placebo group, 312 in the obeticholic acid 10 mg group, and 308 in the obeticholic acid 25 mg group). The fibrosis improvement endpoint was achieved by 37 (12%) patients in the placebo group, 55 (18%) in the obeticholic acid 10 mg group (p=0·045), and 71 (23%) in the obeticholic acid 25 mg group (p=0·0002). The NASH resolution endpoint was not met (25 [8%] patients in the placebo group, 35 [11%] in the obeticholic acid 10 mg group [p=0·18], and 36 [12%] in the obeticholic acid 25 mg group [p=0·13]). In the safety population (1968 patients with fibrosis stages F1–F3), the most common adverse event was pruritus (123 [19%] in the placebo group, 183 [28%] in the obeticholic acid 10 mg group, and 336 [51%] in the obeticholic acid 25 mg group); incidence was generally mild to moderate in severity. The overall safety profile was similar to that in previous studies, and incidence of serious adverse events was similar across treatment groups (75 [11%] patients in the placebo group, 72 [11%] in the obeticholic acid 10 mg group, and 93 [14%] in the obeticholic acid 25 mg group). Interpretation Obeticholic acid 25 mg significantly improved fibrosis and key components of NASH disease activity among patients with NASH. The results from this planned interim analysis show clinically significant histological improvement that is reasonably likely to predict clinical benefit. This study is ongoing to assess clinical outcomes

Investigating the potential role of a human long non-coding RNA with ‘accelerated evolution’ in neurodevelopment and gliomas

Long non-coding RNAs (lncRNAs) have a wide range of functions in health and disease, but many remain uncharacterised. Human accelerated region 1 (HAR1) and HAQER0035 are stretches of genomic DNA that, in humans, have significantly altered base sequences relative to chimpanzees. HAR1A and HAR1B are overlapping lncRNAs containing these sequences, and their expression has been shown to be regulated by the RE1 silencing transcription factor (REST). The functions of these lncRNAs are largely unknown, but they have been associated with neurodevelopment and neurological disorders. In cancers, low expression of HAR1A and HAR1B is correlated with glioma progression and poor patient prognosis. The aim of this study was to characterise and investigate the role of HAR1A and HAR1B in the context of the healthy or diseased human nervous system. The expression patterns of HAR1A, HAR1B and REST were investigated in silico using a developing human brain and clinical glioma datasets, revealing an inverse correlation between the lncRNAs and REST. The expression of the lncRNAs increases postnatally, in structures relevant to memory and sensory processing, and their low expression correlates with advanced glioma grade and poor prognosis. Gene ontology analysis found the most enriched process associated with HAR1 expression to be chemical synaptic signalling. Following the in silico analysis, the expression and function of HAR1A was analysed in a neural progenitor cell line (ReN VM) that could be readily differentiated into astrocytes and dopaminergic neurons, and in multiple brain cancer cell lines (U-373, SH-SY5Y, DIPGA, GIN28, GCE28). Key studies determined the subcellular localisation of HAR1Ato be nuclear. Upon ReN cell differentiation, the expression of HAR1Awas found to decrease within 12 hours, independent of the locus, though cell distribution did not change from baseline. HAR1Awas expressed via lentiviral overexpression, and its subcellular localisation confirmed to be retained in the nucleus. In ReN cells, proliferation, migration assays, KCl incubation and assessment of nuclear morphology by TEM identified little observable effect of HAR1A overexpression. In U-373 cells, HAR1A expression was increased with siRNA silencing of REST, but the reduction in cell proliferation observed was not correlated with HAR1A overexpression. Similar to ReN, little observable effect of HAR1Aoverexpression was found when analysing proliferation, migration and nuclear morphology. This study has revealed that HAR1A is not involved in widely measured phenotypes, however, the rapid degradation associated with differentiation suggest that HAR1A plays a role in maintaining a less specialised, or early neuronal cell type.</br

Comorbidity and treatment response in pediatric obsessive-compulsive disorder : a pilot study of group cognitive-behavioral treatment

This pilot study evaluated the effectiveness of group cognitive-behavioral treatment (CBT) on treatment outcomes for children and adolescents who presented with obsessive–compulsive disorder (OCD) and complex comorbid conditions, including depression, attention deficit/hyperactivity disorder and pervasive developmental disorders (PDD). Specifically, the impact of comorbidity on treatment response rates and remission rates was examined. Forty-three youth (aged 7–17) with OCD participated in group family-based CBT. Assessments were conducted at pre- and post-treatment and 6 months. Eighty-six percent of youth presented with a secondary psychiatric disorder, and 74% presented with a tertiary psychiatric condition. Contrary to the expected, comorbidity was not associated with poorer treatment outcomes at post-assessment. At longer term follow-up (6 months), however, treatment outcomes were poorer for youth with multiple comorbid conditions and for those with attention deficit/hyperactivity disorder. The finding that group CBT is largely effective for youth with comorbid conditions is of clinical and practical significance. Group delivery of CBT provides an efficient and cost-effective approach, and alleviates strain on services and service providers. Continued efforts are needed to improve long-term outcomes for youth with multiple comorbid conditions and attention deficit/hyperactivity disorder. Examining treatment response as a function of comorbidity with larger clinical samples is important to extend this research.9 page(s

Responsibility beliefs, memory confidence, intolerance of uncertainty and the urge to check in childhood obsessive-compulsive disorder : an examination of cognitive theory

Objective: This study aimed to extend current research into cognitive models of obsessive–compulsive disorder (OCD) in a pediatric sample by examining the impact of perceived responsibility on memory confidence, intolerance of uncertainty (IU) and checking urge using an experimental design to manipulate perceived responsibility. It was hypothesised that the high responsibility condition would result in higher ratings of responsibility, lower memory confidence and higher IU, which would also result in higher ratings on urge to check. Moreover, it was hypothesised that adolescents would report significantly higher ratings of responsibility than children. Finally, it was hypothesised that the effect of perceived inflated responsibility on the urge to check in a high responsibility condition would be mediated by IU. Method: Twenty-seven children and adolescents diagnosed with OCD completed an experimental cognitive appraisal task (CAT) in which they heard two standardised vignettes presented in counterbalanced order; one in which participants were responsible and one in which they were not responsible for preventing harm to a friend's pet cat. Memory confidence, IU and checking urge were assessed after each scenario using Likert scales. Results: The manipulation of perceived responsibility was successful with children and adolescents rating increased responsibility in the high compared with the low responsibility scenario. There were no differences across high and low responsibility conditions, however, in ratings of memory confidence, IU or the urge to check. There were no significant age-related differences; however, there was a trend for adolescents to report higher ratings across all variables. Finally, the relationship between perceived inflated responsibility and the urge to check was not mediated by IU. Conclusions: Responsibility is not related to ratings of memory confidence, IU or the urge to check in a pediatric sample, suggesting that biases of responsibility may not be central to the formulation of childhood OCD. Results are discussed in terms of implications for cognitive formulations and cognitive approaches to treatment in pediatric OCD.15 page(s

Blood-Injection-Injury phobia and dog phobia in youth : psychological characteristics and associated features in a clinical sample

Blood-Injection-Injury (BII) phobia is a particularly debilitating condition that has been largely ignored in the child literature. The present study examined the clinical phenomenology of BII phobia in 27 youths, relative to 25 youths with dog phobia-one of the most common and well-studied phobia subtypes in youth. Children were compared on measures of phobia severity, functional impairment, comorbidity, threat appraisals (danger expectancies and coping), focus of fear, and physiological responding, as well as vulnerability factors including disgust sensitivity and family history. Children and adolescents with BII phobia had greater diagnostic severity. In addition, they were more likely to have a comorbid diagnosis of a physical health condition, to report more exaggerated danger expectancies, and to report fears that focused more on physical symptoms (e.g., faintness and nausea) in comparison to youth with dog phobia. The present study advances knowledge relating to this poorly understood condition in youth.13 page(s