Correction: Elevated plasma levels of cardiac troponin-I predict left ventricular systolic dysfunction in patients with myotonic dystrophy type 1: A multicentre cohort follow-up study

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Elevated plasma levels of cardiac troponin-I predict left ventricular systolic dysfunction in patients with myotonic dystrophy type 1:A multicentre cohort follow-up study

Objective: High sensitivity plasma cardiac troponin-I (cTnI) is emerging as a strong predictor of cardiac events in a variety of settings. We have explored its utility in patients with myotonic dystrophy type 1 (DM1). Methods: 117 patients with DM1 were recruited from routine outpatient clinics across three health boards. A single measurement of cTnI was made using the ARCHITECT STAT Troponin I assay. Demographic, ECG, echocardiographic and other clinical data were obtained from electronic medical records. Follow up was for a mean of 23 months. Results: Fifty five females and 62 males (mean age 47.7 years) were included. Complete data were available for ECG in 107, echocardiography in 53. Muscle Impairment Rating Scale score was recorded for all patients. A highly significant excess (p = 0.0007) of DM1 patients presented with cTnI levels greater than the 99th centile of the range usually observed in the general population (9 patients; 7.6%). Three patients with elevated troponin were found to have left ventricular systolic dysfunction (LVSD), compared with four of those with normal range cTnI (33.3% versus 3.7%; p = 0.001). Sixty two patients had a cTnI level < 5ng/L, of whom only one had documented evidence of LVSD. Elevated cTnI was not predictive of severe conduction abnormalities on ECG, or presence of a cardiac device, nor did cTnI level correlate with muscle strength expressed by Muscle Impairment Rating Scale score. Conclusions: Plasma cTnI is highly elevated in some ambulatory patients with DM1 and shows promise as a tool to aid cardiac risk stratification, possibly by detecting myocardial involvement. Further studies with larger patient numbers are warranted to assess its utility in this setting

Case Reports1. A Late Presentation of Loeys-Dietz Syndrome: Beware of TGFβ Receptor Mutations in Benign Joint Hypermobility

Background: Thoracic aortic aneurysms (TAA) and dissections are not uncommon causes of sudden death in young adults. Loeys-Dietz syndrome (LDS) is a rare, recently described, autosomal dominant, connective tissue disease characterized by aggressive arterial aneurysms, resulting from mutations in the transforming growth factor beta (TGFβ) receptor genes TGFBR1 and TGFBR2. Mean age at death is 26.1 years, most often due to aortic dissection. We report an unusually late presentation of LDS, diagnosed following elective surgery in a female with a long history of joint hypermobility. Methods: A 51-year-old Caucasian lady complained of chest pain and headache following a dural leak from spinal anaesthesia for an elective ankle arthroscopy. CT scan and echocardiography demonstrated a dilated aortic root and significant aortic regurgitation. MRA demonstrated aortic tortuosity, an infrarenal aortic aneurysm and aneurysms in the left renal and right internal mammary arteries. She underwent aortic root repair and aortic valve replacement. She had a background of long-standing joint pains secondary to hypermobility, easy bruising, unusual fracture susceptibility and mild bronchiectasis. She had one healthy child age 32, after which she suffered a uterine prolapse. Examination revealed mild Marfanoid features. Uvula, skin and ophthalmological examination was normal. Results: Fibrillin-1 testing for Marfan syndrome (MFS) was negative. Detection of a c.1270G > C (p.Gly424Arg) TGFBR2 mutation confirmed the diagnosis of LDS. Losartan was started for vascular protection. Conclusions: LDS is a severe inherited vasculopathy that usually presents in childhood. It is characterized by aortic root dilatation and ascending aneurysms. There is a higher risk of aortic dissection compared with MFS. Clinical features overlap with MFS and Ehlers Danlos syndrome Type IV, but differentiating dysmorphogenic features include ocular hypertelorism, bifid uvula and cleft palate. Echocardiography and MRA or CT scanning from head to pelvis is recommended to establish the extent of vascular involvement. Management involves early surgical intervention, including early valve-sparing aortic root replacement, genetic counselling and close monitoring in pregnancy. Despite being caused by loss of function mutations in either TGFβ receptor, paradoxical activation of TGFβ signalling is seen, suggesting that TGFβ antagonism may confer disease modifying effects similar to those observed in MFS. TGFβ antagonism can be achieved with angiotensin antagonists, such as Losartan, which is able to delay aortic aneurysm development in preclinical models and in patients with MFS. Our case emphasizes the importance of timely recognition of vasculopathy syndromes in patients with hypermobility and the need for early surgical intervention. It also highlights their heterogeneity and the potential for late presentation. Disclosures: The authors have declared no conflicts of interes

Basic science232. Certolizumab pegol prevents pro-inflammatory alterations in endothelial cell function

Background: Cardiovascular disease is a major comorbidity of rheumatoid arthritis (RA) and a leading cause of death. Chronic systemic inflammation involving tumour necrosis factor alpha (TNF) could contribute to endothelial activation and atherogenesis. A number of anti-TNF therapies are in current use for the treatment of RA, including certolizumab pegol (CZP), (Cimzia ®; UCB, Belgium). Anti-TNF therapy has been associated with reduced clinical cardiovascular disease risk and ameliorated vascular function in RA patients. However, the specific effects of TNF inhibitors on endothelial cell function are largely unknown. Our aim was to investigate the mechanisms underpinning CZP effects on TNF-activated human endothelial cells. Methods: Human aortic endothelial cells (HAoECs) were cultured in vitro and exposed to a) TNF alone, b) TNF plus CZP, or c) neither agent. Microarray analysis was used to examine the transcriptional profile of cells treated for 6 hrs and quantitative polymerase chain reaction (qPCR) analysed gene expression at 1, 3, 6 and 24 hrs. NF-κB localization and IκB degradation were investigated using immunocytochemistry, high content analysis and western blotting. Flow cytometry was conducted to detect microparticle release from HAoECs. Results: Transcriptional profiling revealed that while TNF alone had strong effects on endothelial gene expression, TNF and CZP in combination produced a global gene expression pattern similar to untreated control. The two most highly up-regulated genes in response to TNF treatment were adhesion molecules E-selectin and VCAM-1 (q 0.2 compared to control; p > 0.05 compared to TNF alone). The NF-κB pathway was confirmed as a downstream target of TNF-induced HAoEC activation, via nuclear translocation of NF-κB and degradation of IκB, effects which were abolished by treatment with CZP. In addition, flow cytometry detected an increased production of endothelial microparticles in TNF-activated HAoECs, which was prevented by treatment with CZP. Conclusions: We have found at a cellular level that a clinically available TNF inhibitor, CZP reduces the expression of adhesion molecule expression, and prevents TNF-induced activation of the NF-κB pathway. Furthermore, CZP prevents the production of microparticles by activated endothelial cells. This could be central to the prevention of inflammatory environments underlying these conditions and measurement of microparticles has potential as a novel prognostic marker for future cardiovascular events in this patient group. Disclosure statement: Y.A. received a research grant from UCB. I.B. received a research grant from UCB. S.H. received a research grant from UCB. All other authors have declared no conflicts of interes

Adding 6 months of androgen deprivation therapy to postoperative radiotherapy for prostate cancer: a comparison of short-course versus no androgen deprivation therapy in the RADICALS-HD randomised controlled trial

Background Previous evidence indicates that adjuvant, short-course androgen deprivation therapy (ADT) improves metastasis-free survival when given with primary radiotherapy for intermediate-risk and high-risk localised prostate cancer. However, the value of ADT with postoperative radiotherapy after radical prostatectomy is unclear. Methods RADICALS-HD was an international randomised controlled trial to test the efficacy of ADT used in combination with postoperative radiotherapy for prostate cancer. Key eligibility criteria were indication for radiotherapy after radical prostatectomy for prostate cancer, prostate-specific antigen less than 5 ng/mL, absence of metastatic disease, and written consent. Participants were randomly assigned (1:1) to radiotherapy alone (no ADT) or radiotherapy with 6 months of ADT (short-course ADT), using monthly subcutaneous gonadotropin-releasing hormone analogue injections, daily oral bicalutamide monotherapy 150 mg, or monthly subcutaneous degarelix. Randomisation was done centrally through minimisation with a random element, stratified by Gleason score, positive margins, radiotherapy timing, planned radiotherapy schedule, and planned type of ADT, in a computerised system. The allocated treatment was not masked. The primary outcome measure was metastasis-free survival, defined as distant metastasis arising from prostate cancer or death from any cause. Standard survival analysis methods were used, accounting for randomisation stratification factors. The trial had 80% power with two-sided α of 5% to detect an absolute increase in 10-year metastasis-free survival from 80% to 86% (hazard ratio [HR] 0·67). Analyses followed the intention-to-treat principle. The trial is registered with the ISRCTN registry, ISRCTN40814031, and ClinicalTrials.gov, NCT00541047. Findings Between Nov 22, 2007, and June 29, 2015, 1480 patients (median age 66 years [IQR 61–69]) were randomly assigned to receive no ADT (n=737) or short-course ADT (n=743) in addition to postoperative radiotherapy at 121 centres in Canada, Denmark, Ireland, and the UK. With a median follow-up of 9·0 years (IQR 7·1–10·1), metastasis-free survival events were reported for 268 participants (142 in the no ADT group and 126 in the short-course ADT group; HR 0·886 [95% CI 0·688–1·140], p=0·35). 10-year metastasis-free survival was 79·2% (95% CI 75·4–82·5) in the no ADT group and 80·4% (76·6–83·6) in the short-course ADT group. Toxicity of grade 3 or higher was reported for 121 (17%) of 737 participants in the no ADT group and 100 (14%) of 743 in the short-course ADT group (p=0·15), with no treatment-related deaths. Interpretation Metastatic disease is uncommon following postoperative bed radiotherapy after radical prostatectomy. Adding 6 months of ADT to this radiotherapy did not improve metastasis-free survival compared with no ADT. These findings do not support the use of short-course ADT with postoperative radiotherapy in this patient population

Duration of androgen deprivation therapy with postoperative radiotherapy for prostate cancer: a comparison of long-course versus short-course androgen deprivation therapy in the RADICALS-HD randomised trial

Background Previous evidence supports androgen deprivation therapy (ADT) with primary radiotherapy as initial treatment for intermediate-risk and high-risk localised prostate cancer. However, the use and optimal duration of ADT with postoperative radiotherapy after radical prostatectomy remains uncertain. Methods RADICALS-HD was a randomised controlled trial of ADT duration within the RADICALS protocol. Here, we report on the comparison of short-course versus long-course ADT. Key eligibility criteria were indication for radiotherapy after previous radical prostatectomy for prostate cancer, prostate-specific antigen less than 5 ng/mL, absence of metastatic disease, and written consent. Participants were randomly assigned (1:1) to add 6 months of ADT (short-course ADT) or 24 months of ADT (long-course ADT) to radiotherapy, using subcutaneous gonadotrophin-releasing hormone analogue (monthly in the short-course ADT group and 3-monthly in the long-course ADT group), daily oral bicalutamide monotherapy 150 mg, or monthly subcutaneous degarelix. Randomisation was done centrally through minimisation with a random element, stratified by Gleason score, positive margins, radiotherapy timing, planned radiotherapy schedule, and planned type of ADT, in a computerised system. The allocated treatment was not masked. The primary outcome measure was metastasis-free survival, defined as metastasis arising from prostate cancer or death from any cause. The comparison had more than 80% power with two-sided α of 5% to detect an absolute increase in 10-year metastasis-free survival from 75% to 81% (hazard ratio [HR] 0·72). Standard time-to-event analyses were used. Analyses followed intention-to-treat principle. The trial is registered with the ISRCTN registry, ISRCTN40814031, and ClinicalTrials.gov , NCT00541047 . Findings Between Jan 30, 2008, and July 7, 2015, 1523 patients (median age 65 years, IQR 60–69) were randomly assigned to receive short-course ADT (n=761) or long-course ADT (n=762) in addition to postoperative radiotherapy at 138 centres in Canada, Denmark, Ireland, and the UK. With a median follow-up of 8·9 years (7·0–10·0), 313 metastasis-free survival events were reported overall (174 in the short-course ADT group and 139 in the long-course ADT group; HR 0·773 [95% CI 0·612–0·975]; p=0·029). 10-year metastasis-free survival was 71·9% (95% CI 67·6–75·7) in the short-course ADT group and 78·1% (74·2–81·5) in the long-course ADT group. Toxicity of grade 3 or higher was reported for 105 (14%) of 753 participants in the short-course ADT group and 142 (19%) of 757 participants in the long-course ADT group (p=0·025), with no treatment-related deaths. Interpretation Compared with adding 6 months of ADT, adding 24 months of ADT improved metastasis-free survival in people receiving postoperative radiotherapy. For individuals who can accept the additional duration of adverse effects, long-course ADT should be offered with postoperative radiotherapy. Funding Cancer Research UK, UK Research and Innovation (formerly Medical Research Council), and Canadian Cancer Society

Modelling human choices: MADeM and decision‑making

Research supported by FAPESP 2015/50122-0 and DFG-GRTK 1740/2. RP and AR are also part of the Research, Innovation and Dissemination Center for Neuromathematics FAPESP grant (2013/07699-0). RP is supported by a FAPESP scholarship (2013/25667-8). ACR is partially supported by a CNPq fellowship (grant 306251/2014-0)

Identification of novel factors involved in y-globin gene regulation

Typescript (photocopy)Cover title: Identification of novel factors involved in gamma-globin gene regulationRotary Bone Marrow Research Laboratories, Royal Melbourne HospitalThesis (PhD) -- University of Melbourne, Faculty of Medicine, Dentistry and Health Sciences, 2004Includes bibliographical references (leaves 175-201)The ? to ? globin switch that occurs at birth also marks the onset of the adult globin disorders including ? thalassemia and Sickle Cell Disease. These are long-term chronic disorders requiring intense therapy. It was discovered that re-activating ? globin in patients can lead to amelioration of the symptoms. Therefore understanding the regulation of the ? to P switch and the factors that are involved is important as it may identify targets for reversing the switch and re-activating ? globin in adults. NF-E4 is a powerful activator of ? globin, however, its mechanism of activation is unknown This thesis concentrated on identifying partner proteins of NF-E4 and determining the complex that NF-E4 forms to regulate ? globin. A candidate gene approach determined that NF-E4 interacts with the co-activator and histone acetyltransferase protein CBP in vivo. NF- E4 also interacts in vivo with the histone deacetylase factor and general repressor, HDAC1. All 3 factors bind to the ? promoter but HDAC1 appears to bind preferentially when the ? promoter is in a repressed state. NF-E4 also interacts in vitro with TAF70 and the TBP from the TFIID complex. These interactions provide evidence that NF-E4 functions both as an activator of ? along with CBP and the TFIID complex as well as functioning as a repressor with HDAC1. In order to identify novel partner proteins of NF-E4 an unbiased yeast-2-hybrid screen was performed with NF-E4 as the bait. Over 4 x 105 genes were screened using cDNA libraries constructed from tissues expressing ? globin and 36 interacting factors were isolated. Included were 2 factors of particular interest, Nmi and SHARP. Nmi interacts with CBP to increase transcription and SHARP interacts with HDAC1 and the NuRD complex to silence genes. Further characterisation of the interactions between NF-E4 and Nmi determined that the 2 factors form a complex in vitro and in vivo. Nmi locates to the nucleus of ? expressing K562 cells and is bound to the ? promoter during y expression. A mutant form of NF-E4 that does not interact with Nmi cannot induce high levels of ? globin showing a dependence on Nmi for NF-E4 function. Nmi also appears to stabilise NF-E4 in a process that may be affected by the acetylation status of NF-E4. SHARP interacts with NF-E4 in vitro and preliminary experiments suggest SHARP acts as a repressor of the ? globin gene. In conclusion the role of NF-E4 in ? regulation appears to be twofold. During the foetal stage, when ? levels are highest NF-E4 is stabilised by Nmi. NF-E4 interacts with CBP to open chromatin surrounding the ? promoter or to generally increase transcription factor binding to activate transcription. NF-E4 also stabilises the TFIID complex at the y promoter where it would recruit RNA Po1II to initiate y gene transcription. During the adult stage when ? is silenced NF-E4 interacts with HDAC1 and SHARP at the ? promoter. HDAC1 and SHARP both interact with the NuRD repressor complex and together they would condense the ? locus into closed chromatin and silence the ? gene

CiIIder:A tool for predicting and analysing transcription factor binding sites

The availability of large amounts of high-throughput genomic, transcriptomic and epigenomic data has provided opportunity to understand regulation of the cellular transcriptome with an unprecedented level of detail. As a result, research has advanced from identifying gene expression patterns associated with particular conditions to elucidating signalling pathways that regulate expression. There are over 1,000 transcription factors (TFs) in vertebrates that play a role in this regulation. Determining which of these are likely to be controlling a set of genes can be assisted by computational prediction, utilising experimentally verified binding site motifs. Here we present CiiiDER, an integrated computational toolkit for transcription factor binding analysis, written in the Java programming language, to make it independent of computer operating system. It is operated through an intuitive graphical user interface with interactive, high-quality visual outputs, making it accessible to all researchers. CiiiDER predicts transcription factor binding sites (TFBSs) across regulatory regions of interest, such as promoters and enhancers derived from any species. It can perform an enrichment analysis to identify TFs that are significantly over- or under-represented in comparison to a bespoke background set and thereby elucidate pathways regulating sets of genes of pathophysiological importance