Analysis and design of randomised clinical trials involving competing risks endpoints

<p>Abstract</p> <p>Background</p> <p>In randomised clinical trials involving time-to-event outcomes, the failures concerned may be events of an entirely different nature and as such define a classical competing risks framework. In designing and analysing clinical trials involving such endpoints, it is important to account for the competing events, and evaluate how each contributes to the overall failure. An appropriate choice of statistical model is important for adequate determination of sample size.</p> <p>Methods</p> <p>We describe how competing events may be summarised in such trials using cumulative incidence functions and Gray's test. The statistical modelling of competing events using proportional cause-specific and subdistribution hazard functions, and the corresponding procedures for sample size estimation are outlined. These are illustrated using data from a randomised clinical trial (SQNP01) of patients with advanced (non-metastatic) nasopharyngeal cancer.</p> <p>Results</p> <p>In this trial, treatment has no effect on the competing event of loco-regional recurrence. Thus the effects of treatment on the hazard of distant metastasis were similar via both the cause-specific (unadjusted <it>csHR </it>= 0.43, 95% CI 0.25 - 0.72) and subdistribution (unadjusted <it>subHR </it>0.43; 95% CI 0.25 - 0.76) hazard analyses, in favour of concurrent chemo-radiotherapy followed by adjuvant chemotherapy. Adjusting for nodal status and tumour size did not alter the results. The results of the logrank test (<it>p </it>= 0.002) comparing the cause-specific hazards and the Gray's test (<it>p </it>= 0.003) comparing the cumulative incidences also led to the same conclusion. However, the subdistribution hazard analysis requires many more subjects than the cause-specific hazard analysis to detect the same magnitude of effect.</p> <p>Conclusions</p> <p>The cause-specific hazard analysis is appropriate for analysing competing risks outcomes when treatment has no effect on the cause-specific hazard of the competing event. It requires fewer subjects than the subdistribution hazard analysis for a similar effect size. However, if the main and competing events are influenced in opposing directions by an intervention, a subdistribution hazard analysis may be warranted.</p

Microsecond Time-Resolved Absorption Spectroscopy Used to Study CO Compounds of Cytochrome bd from Escherichia coli

Cytochrome bd is a tri-heme (b558, b595, d) respiratory oxygen reductase that is found in many bacteria including pathogenic species. It couples the electron transfer from quinol to O2 with generation of an electrochemical proton gradient. We examined photolysis and subsequent recombination of CO with isolated cytochrome bd from Escherichia coli in oneelectron reduced (MV) and fully reduced (R) states by microsecond time-resolved absorption spectroscopy at 532-nm excitation. Both Soret and visible band regions were examined. CO photodissociation from MV enzyme possibly causes fast (t,1.5 ms) electron transfer from heme d to heme b595 in a small fraction of the protein, not reported earlier. Then the electron migrates to heme b558 (t,16 ms). It returns from the b-hemes to heme d with t,180 ms. Unlike cytochrome bd in the R state, in MV enzyme the apparent contribution of absorbance changes associated with CO dissociation from heme d is small, if any. Photodissociation of CO from heme d in MV enzyme is suggested to be accompanied by the binding of an internal ligand (L) at the opposite side of the heme. CO recombines with heme d (t,16 ms) yielding a transient hexacoordinate state (CO-Fe2+ -L). Then the ligand slowly (t,30 ms) dissociates from heme d. Recombination of CO with a reduced heme b in a fraction of the MV sample may also contribute to the 30-ms phase. In R enzyme, CO recombines to heme d (t,20 ms), some heme b558 (t,0.2–3 ms), and finally migrates from heme d to heme b595 (t,24 ms) in ,5% of the enzyme population. Data are consistent with the recent nanosecond study of Rappaport et al. conducted on the membranes at 640-nm excitation but limited to the Soret band. The additional phases were revealed due to differences in excitation and other experimental conditions

The Terminal Oxidase Cytochrome bd Promotes Sulfide-resistant Bacterial Respiration and Growth

Hydrogen sulfide (H2S) impairs mitochondrial respiration by potently inhibiting the heme-copper cytochrome c oxidase. Since many prokaryotes, including Escherichia (E.) coli, generate H2S and encounter high H2S levels particularly in the human gut, herein we tested whether bacteria can sustain sulfide-resistant O2-dependent respiration. E. coli has three respiratory oxidases, the cyanide-sensitive heme-copper bo3 enzyme and two bd oxidases much less sensitive to cyanide. Working on the isolated enzymes, we found that, whereas the bo3 oxidase is inhibited by sulfide with half-maximal inhibitory concentration IC50=1.1±0.1μM, under identical experimental conditions both bd oxidases are insensitive to sulfide up to 58μM. In E. coli respiratory mutants, both O2-consumption and aerobic growth proved to be severely impaired by sulfide when respiration was sustained by the bo3 oxidase alone, but unaffected by ≤200μM sulfide when either bd enzyme acted as the only terminal oxidase. Accordingly, wild-type E. coli showed sulfide-insensitive respiration and growth under conditions favouring the expression of bd oxidases. In all tested conditions, cyanide mimicked the functional effect of sulfide on bacterial respiration. We conclude that bd oxidases promote sulfide-resistant O2- consumption and growth in E. coli and possibly other bacteria. The impact of this discovery is discussed

Indicators of river system hydromorphological character and dynamics: understanding current conditions and guiding sustainable river management

The work leading to this paper received funding from the EU’s FP7 programme under Grant Agreement No. 282656 (REFORM). The Indicators were developed within the context of REFORM deliverable D2.1, therefore all partners involved in this deliverable contributed to some extent to their discussion and development

Pulmonary oxygen uptake and muscle deoxygenation kinetics during recovery in trained and untrained male adolescents

Previous studies have demonstrated faster pulmonary oxygen uptake ( V ˙ O 2 ) kinetics in the trained state during the transition to and from moderate-intensity exercise in adults. Whilst a similar effect of training status has previously been observed during the on-transition in adolescents, whether this is also observed during recovery from exercise is presently unknown. The aim of the present study was therefore to examine V ˙ O 2 kinetics in trained and untrained male adolescents during recovery from moderate-intensity exercise. 15 trained (15 ± 0.8 years, V ˙ O 2max 54.9 ± 6.4 mL kg−1 min−1) and 8 untrained (15 ± 0.5 years, V ˙ O 2max 44.0 ± 4.6 mL kg−1 min−1) male adolescents performed two 6-min exercise off-transitions to 10 W from a preceding “baseline” of exercise at a workload equivalent to 80% lactate threshold; V ˙ O 2 (breath-by-breath) and muscle deoxyhaemoglobin (near-infrared spectroscopy) were measured continuously. The time constant of the fundamental phase of V ˙ O 2 off-kinetics was not different between trained and untrained (trained 27.8 ± 5.9 s vs. untrained 28.9 ± 7.6 s, P = 0.71). However, the time constant (trained 17.0 ± 7.5 s vs. untrained 32 ± 11 s, P < 0.01) and mean response time (trained 24.2 ± 9.2 s vs. untrained 34 ± 13 s, P = 0.05) of muscle deoxyhaemoglobin off-kinetics was faster in the trained subjects compared to the untrained subjects. V ˙ O 2 kinetics was unaffected by training status; the faster muscle deoxyhaemoglobin kinetics in the trained subjects thus indicates slower blood flow kinetics during recovery from exercise compared to the untrained subjects

Spatial Anisotropies and Temporal Fluctuations in Extracellular Matrix Network Texture during Early Embryogenesis

Early stages of vertebrate embryogenesis are characterized by a remarkable series of shape changes. The resulting morphological complexity is driven by molecular, cellular, and tissue-scale biophysical alterations. Operating at the cellular level, extracellular matrix (ECM) networks facilitate cell motility. At the tissue level, ECM networks provide material properties required to accommodate the large-scale deformations and forces that shape amniote embryos. In other words, the primordial biomaterial from which reptilian, avian, and mammalian embryos are molded is a dynamic composite comprised of cells and ECM. Despite its central importance during early morphogenesis we know little about the intrinsic micrometer-scale surface properties of primordial ECM networks. Here we computed, using avian embryos, five textural properties of fluorescently tagged ECM networks — (a) inertia, (b) correlation, (c) uniformity, (d) homogeneity, and (e) entropy. We analyzed fibronectin and fibrillin-2 as examples of fibrous ECM constituents. Our quantitative data demonstrated differences in the surface texture between the fibronectin and fibrillin-2 network in Day 1 (gastrulating) embryos, with the fibronectin network being relatively coarse compared to the fibrillin-2 network. Stage-specific regional anisotropy in fibronectin texture was also discovered. Relatively smooth fibronectin texture was exhibited in medial regions adjoining the primitive streak (PS) compared with the fibronectin network investing the lateral plate mesoderm (LPM), at embryonic stage 5. However, the texture differences had changed by embryonic stage 6, with the LPM fibronectin network exhibiting a relatively smooth texture compared with the medial PS-oriented network. Our data identify, and partially characterize, stage-specific regional anisotropy of fibronectin texture within tissues of a warm-blooded embryo. The data suggest that changes in ECM textural properties reflect orderly time-dependent rearrangements of a primordial biomaterial. We conclude that the ECM microenvironment changes markedly in time and space during the most important period of amniote morphogenesis—as determined by fluctuating textural properties

A feasibility study exploring the role of pre-operative assessment when examining the mechanism of ‘chemo-brain’ in breast cancer patients

Background: Women receiving chemotherapy treatment for breast cancer may experience problems with their memory and attention (cognition), which is distressing and interferes with quality of life. It is unclear what causes or contributes to the problems they report: psychological distress, fatigue, coping style, or specific biological changes for example to pro inflammatory cytokines. Research shows however, that approximately a third of women with breast cancer perform poorly on tests of cognition before commencing chemotherapy. We aimed to examine the acceptability and relevance of pre-surgical assessments (bloods, brain imaging, cognitive tests and self-report questionnaires) when investigating the phenomenon of ‘chemo-brain’ and investigate whether inflammatory markers mediate chemotherapy-induced neuropsychological impairments in women treated for breast cancer. Methods: Women with early stage breast cancer completed neuropsychological and quality of life assessments at T1 (pre-surgery), T2 (post-surgery before chemotherapy) and T3 (6 months later). Blood cytokine levels were measured at the same time points and brain imaging was performed at T1 and T3. Results: In total, 14/58 women participated (8 chemotherapy, 6 non-chemotherapy). Prior to the start of chemotherapy a decline in cognitive performance compared to baseline was observed in one participant. At T3 women who received chemotherapy reported poorer quality of life and greater fatigue. Increases in soluble tumour necrosis factor receptor II (sTNFRII), interleukin-6, interleukin-10 and vascular endothelial growth factor occurred post chemotherapy only. Levels of sTNFRII were inversely correlated with grey matter volume (GMV) of the right posterior insula in both groups. At T3, the chemotherapy group displayed a greater reduction in GMV in the subgenual and dorsal anterior cingulate, and the inferior temporal gyrus. Conclusions: Pre-operative recruitment to the study was challenging; however, the lack of significant changes in blood cytokine levels and neuropsychological tests at T2 implies that post surgery may be a valid baseline assessment, but this needs further investigation in a larger study. The preliminary results support the hypothesis that chemotherapy induced fatigue is mediated by a change in peripheral cytokine levels which could explain some symptoms of ‘chemo brain’ experienced by patients

A framework for teaching epistemic insight in schools

This paper gives the rationale and a draft outline for a framework for education to teach epistemic insight into schools in England. The motivation to research and propose a strategy to teach and assess epistemic insight followed research that investigated how students and teachers in primary and secondary schools respond to big questions about the nature of reality and human personhood. The research revealed that there are pressures in schools that dampen students’ expressed curiosity in these types of questions and limit their developing epistemic insight into how science, religion and the wider humanities relate. These findings prompted the construction of a framework for education for students aged 5–16 designed to encourage students’ expressed interest in big questions and develop their understanding of the ways that science interacts with other ways of knowing. The centrepiece of the framework is a sequence of learning objectives for epistemic insight, organised into three categories. The categories are, firstly, the nature of science in real world contexts and multidisciplinary arenas; secondly, ways of knowing and how they interact; and thirdly, the relationships between science and religion. Our current version of the Framework is constructed to respond to the way that teaching is organised in England. The key principles and many of the activities could be adopted and tailored to work in many other countries

Increased autophagy in EphrinB2-deficient osteocytes is associated with elevated secondary mineralization and brittle bone

Mineralized bone forms when collagen-containing osteoid accrues mineral crystals. This is initiated rapidly (primary mineralization), and continues slowly (secondary mineralization) until bone is remodeled. The interconnected osteocyte network within the bone matrix differentiates from bone-forming osteoblasts; although osteoblast differentiation requires EphrinB2, osteocytes retain its expression. Here we report brittle bones in mice with osteocyte-targeted EphrinB2 deletion. This is not caused by low bone mass, but by defective bone material. While osteoid mineralization is initiated at normal rate, mineral accrual is accelerated, indicating that EphrinB2 in osteocytes limits mineral accumulation. No known regulators of mineralization are modified in the brittle cortical bone but a cluster of autophagy-associated genes are dysregulated. EphrinB2-deficient osteocytes displayed more autophagosomes in vivo and in vitro, and EphrinB2-Fc treatment suppresses autophagy in a RhoA-ROCK dependent manner. We conclude that secondary mineralization involves EphrinB2-RhoA-limited autophagy in osteocytes, and disruption leads to a bone fragility independent of bone mass

p21 as a Transcriptional Co-Repressor of S-Phase and Mitotic Control Genes

It has been previously described that p21 functions not only as a CDK inhibitor but also as a transcriptional co-repressor in some systems. To investigate the roles of p21 in transcriptional control, we studied the gene expression changes in two human cell systems. Using a human leukemia cell line (K562) with inducible p21 expression and human primary keratinocytes with adenoviral-mediated p21 expression, we carried out microarray-based gene expression profiling. We found that p21 rapidly and strongly repressed the mRNA levels of a number of genes involved in cell cycle and mitosis. One of the most strongly down-regulated genes was CCNE2 (cyclin E2 gene). Mutational analysis in K562 cells showed that the N-terminal region of p21 is required for repression of gene expression of CCNE2 and other genes. Chromatin immunoprecipitation assays indicated that p21 was bound to human CCNE2 and other p21-repressed genes gene in the vicinity of the transcription start site. Moreover, p21 repressed human CCNE2 promoter-luciferase constructs in K562 cells. Bioinformatic analysis revealed that the CDE motif is present in most of the promoters of the p21-regulated genes. Altogether, the results suggest that p21 exerts a repressive effect on a relevant number of genes controlling S phase and mitosis. Thus, p21 activity as inhibitor of cell cycle progression would be mediated not only by the inhibition of CDKs but also by the transcriptional down-regulation of key genes