RMA with quantile normalization mixes biological signals between different sample groups in microarray data analysis

Quantile normalization (QN) is a technique for microarray data processing and is the default normalization method in the Robust Multi-array Average (RMA) procedure, which was primarily designed for analysing gene expression data from Affymetrix arrays. Given the abundance of Affymetrix microarrays and the popularity of the RMA method, it is crucially important that the normalization procedure is applied appropriately. In this study we carried out simulation experiments and also analysed real microarray data to investigate the suitability of RMA when it is applied to dataset with different groups of biological samples. From our experiments, we showed that RMA with QN does not preserve the biological signal included in each group, but rather it would mix the signals between the groups. We also showed that the Median Polish method in the summarization step of RMA has similar mixing effect. RMA is one of the most widely used methods in microarray data processing and has been applied to a vast volume of data in biomedical research. The problematic behaviour of this method suggests that previous studies employing RMA could have been misadvised or adversely affected. Therefore we think it is crucially important that the research community recognizes the issue and starts to address it. The two core elements of the RMA method, quantile normalization and Median Polish, both have the undesirable effects of mixing biological signals between different sample groups, which can be detrimental to drawing valid biological conclusions and to any subsequent analyses. Based on the evidence presented here and that in the literature, we recommend exercising caution when using RMA as a method of processing microarray gene expression data, particularly in situations where there are likely to be unknown subgroups of samples.

The Type IIb SN 2011dh: Two years of observations and modelling of the lightcurves

We present optical and near-infrared (NIR) photometry and spectroscopy as well as modelling of the lightcurves of the Type IIb supernova (SN) 2011dh. Our extensive dataset, for which we present the observations obtained after day 100, spans two years, and complemented with Spitzer mid-infrared (MIR) data, we use it to build an optical-to-MIR bolometric lightcurve between days 3 and 732. To model the bolometric lightcurve before day 400 we use a grid of hydrodynamical SN models, which allows us to determine the errors in the derived quantities, and a bolometric correction determined with steady-state non-local thermodynamic equilibrium (NLTE) modelling. Using this method we find a helium core mass of 3.1^+0.7<inf>-0.4</inf> M<inf>⊙</inf> for SN 2011dh, consistent within error bars with previous results obtained using the bolometric lightcurve before day 80. We compute bolometric and broad-band lightcurves between days 100 and 500 from spectral steady-state NLTE models, presented and discussed in a companion paper. The preferred 12 M<inf>⊙</inf> (initial mass) model, previously found to agree well with the observed spectra, shows a good overall agreement with the observed lightcurves, although some discrepancies exist. Time-dependent NLTE modelling shows that after day ∼600 a steady-state assumption is no longer valid. The radioactive energy deposition in this phase is likely dominated by the positrons emitted in the decay of ⁵⁶Co, but seems insufficient to reproduce the lightcurves, and what energy source is dominating the emitted flux is unclear. We find an excess in the K and the MIR bands developing between days 100 and 250, during which an increase in the optical decline rate is also observed. A local origin of the excess is suggested by the depth of the He I 20 581 Å absorption. Steady-state NLTE models with a modest dust opacity in the core (τ = 0.44), turned on during this period, reproduce the observed behaviour, but an additional excess in the Spitzer 4.5 μm band remains. Carbon-monoxide (CO) first-overtone band emission is detected at day 206, and possibly at day 89, and assuming the additional excess to bedominated by CO fundamental band emission, we find fundamental to first-overtone band ratios considerably higher than observed in SN 1987A. The profiles of the [O i] 6300 Å and Mg i] 4571 Å lines show a remarkable similarity, suggesting that these lines originate from a common nuclear burning zone (O/Ne/Mg), and using small scale fluctuations in the line profiles we estimate a filling factor of ≲ 0.07 for the emitting material. This paper concludes our extensive observational and modelling work on SN 2011dh. The results from hydrodynamical modelling, steady-state NLTE modelling, and stellar evolutionary progenitor analysis are all consistent, and suggest an initial mass of ∼12 M<inf>⊙</inf> for the progenitor.

Integrin genetic variants and stage-specific tumor recurrence in patients with stage II and III colon cancer

Integrins (ITGs) are key elements in cancer biology, regulating tumor growth, angiogenesis and lymphangiogenesis through interactions of the tumor cells with the microenvironment. Moving from the hypothesis that ITGs could have different effects in stage II and III colon cancer, we tested whether a comprehensive panel of germline single-nucleotide polymorphisms (SNPs) in ITG genes could predict stage-specific time to tumor recurrence (TTR). A total of 234 patients treated with 5-fluorouracil-based chemotherapy at the University of Southern California were included in this study. Whole-blood samples were analyzed for germline SNPs in ITG genes using PCR-restriction fragment length polymorphism or direct DNA sequencing. In the multivariable analysis, stage II colon cancer patients with at least one G allele for ITGB3 rs4642 had higher risk of recurrence (hazard ratio (HR)=4.027, 95% confidence interval (95% CI) 1.556-10.421, P=0.004). This association was also significant in the combined stage II-III cohort (HR=1.975, 95% CI 1.194-3.269, P=0.008). The predominant role of ITGB3 rs4642 in stage II diseases was confirmed using recursive partitioning, showing that ITGB3 rs4642 was the most important factor in stage II diseases. In contrast, in stage III diseases the combined analysis of ITGB1 rs2298141 and ITGA4 rs7562325 allowed to identify three distinct prognostic subgroups (P=0.009). The interaction between stage and the combined ITGB1 rs2298141 and ITGA4 rs7562325 on TTR was significant (P=0.025). This study identifies germline polymorphisms in ITG genes as independent stage-specific prognostic markers for stage II and III colon cancer. These data may help to select subgroups of patients who may benefit from ITG-targeted treatments.

Integration of copy number and transcriptomics provides risk stratification in prostate cancer: A discovery and validation cohort study

BACKGROUND: Understanding the heterogeneous genotypes and phenotypes of prostate cancer is fundamental to improving the way we treat this disease. As yet, there are no validated descriptions of prostate cancer subgroups derived from integrated genomics linked with clinical outcome.

METHODS: In a study of 482 tumour, benign and germline samples from 259 men with primary prostate cancer, we used integrative analysis of copy number alterations (CNA) and array transcriptomics to identify genomic loci that affect expression levels of mRNA in an expression quantitative trait loci (eQTL) approach, to stratify patients into subgroups that we then associated with future clinical behaviour, and compared with either CNA or transcriptomics alone.

FINDINGS: We identified five separate patient subgroups with distinct genomic alterations and expression profiles based on 100 discriminating genes in our separate discovery and validation sets of 125 and 103 men. These subgroups were able to consistently predict biochemical relapse (p = 0.0017 and p = 0.016 respectively) and were further validated in a third cohort with long-term follow-up (p = 0.027). We show the relative contributions of gene expression and copy number data on phenotype, and demonstrate the improved power gained from integrative analyses. We confirm alterations in six genes previously associated with prostate cancer (MAP3K7, MELK, RCBTB2, ELAC2, TPD52, ZBTB4), and also identify 94 genes not previously linked to prostate cancer progression that would not have been detected using either transcript or copy number data alone. We confirm a number of previously published molecular changes associated with high risk disease, including MYC amplification, and NKX3-1, RB1 and PTEN deletions, as well as over-expression of PCA3 and AMACR, and loss of MSMB in tumour tissue. A subset of the 100 genes outperforms established clinical predictors of poor prognosis (PSA, Gleason score), as well as previously published gene signatures (p = 0.0001). We further show how our molecular profiles can be used for the early detection of aggressive cases in a clinical setting, and inform treatment decisions.

INTERPRETATION: For the first time in prostate cancer this study demonstrates the importance of integrated genomic analyses incorporating both benign and tumour tissue data in identifying molecular alterations leading to the generation of robust gene sets that are predictive of clinical outcome in independent patient cohorts.

Protocolized versus non-protocolized weaning for reducing the duration of invasive mechanical ventilation in newborn infants: review

Background
Mechanical ventilation is a life-saving intervention for critically ill newborn infants with respiratory failure admitted to a neonatal intensive care unit (NICU). Ventilating newborn infants can be challenging due to small tidal volumes, high breathing frequencies, and the use of uncuffed endotracheal tubes. Mechanical ventilation has several short-term, as well as long-term complications. To prevent complications, weaning from the ventilator is started as soon as possible. Weaning aims to support the transfer from full mechanical ventilation support to spontaneous breathing activity.

Objectives
To assess the efficacy of protocolized versus non-protocolized ventilator weaning for newborn infants in reducing the duration of invasive mechanical ventilation, the duration of weaning, and shortening the NICU and hospital length of stay. To determine efficacy in predefined subgroups including: gestational age and birth weight; type of protocol; and type of protocol delivery. To establish whether protocolized weaning is safe and clinically effective in reducing the duration of mechanical ventilation without increasing the risk of adverse events.

Search methods
We searched the Cochrane Central Register of Controlled trials (CENTRAL; the Cochrane Library; 2015, Issue 7); MEDLINE In-Process and other Non-Indexed Citations and OVID MEDLINE (1950 to 31 July 2015); CINAHL (1982 to 31 July 2015); EMBASE (1988 to 31 July 2015); and Web of Science (1990 to 15 July 2015). We did not restrict language of publication. We contacted authors of studies with a subgroup of newborn infants in their study, and experts in the field regarding this subject. In addition, we searched abstracts from conference proceedings, theses, dissertations, and reference lists of all identified studies for further relevant studies.

Selection criteria
Randomized, quasi-randomized or cluster-randomized controlled trials that compared protocolized with non-protocolized ventilator weaning practices in newborn infants with a gestational age of 24 weeks or more, who were enrolled in the study before the postnatal age of 28 completed days after the expected date of birth.

Data collection and analysis
Four authors, in pairs, independently reviewed titles and abstracts identified by electronic searches. We retrieved full-text versions of potentially relevant studies.

Main results
Our search yielded 1752 records. We removed duplicates (1062) and irrelevant studies (843). We did not find any randomized, quasi-randomized or cluster-randomized controlled trials conducted on weaning from mechanical ventilation in newborn infants. Two randomized controlled trials met the inclusion criteria on type of study and type of intervention, but only included a proportion of newborns. The study authors could not provide data needed for subgroup analysis; we excluded both studies.

Authors' conclusions
Based on the results of this review, there is no evidence to support or refute the superiority or inferiority of weaning by protocol over non-protocol weaning on duration of invasive mechanical ventilation in newborn infants.

Survival of patients with skin melanoma in Europe increases further: Results of the EUROCARE-5 study

Background In Europe skin melanoma (SM) survival has increased over time. The aims were to evaluate recent trends and differences between countries and regions of Europe.

Methods Relative survival (RS) estimates and geographical comparisons were based on 241,485 patients aged 15 years and over with a diagnosis of invasive SM in Europe (2000-2007). Survival time trends during 1999-2007 were estimated using the period approach, for 213,101 patients. Age, gender, sub-sites and morphology subgroups were considered. 

Results In European patients, estimated 5-year RS was 83% (95% confidence interval, CI 83-84%). The highest values were found for patients resident in Northern (88%; 87-88%) and Central (88%; 87-88%) Europe, followed by Ireland and United Kingdom (UK) (86%; 85-86%) and Southern Europe (83%; 82-83%). The lowest survival was in Eastern Europe (74%; 74-75%). Within regions the intercountry absolute difference in percentage points of RS varied from 4% (North) to 34% (East). RS decreased markedly with patients' age and was higher in women than men. Differences according to SM morphology and skin sub-sites also emerged. Survival has slightly increased from 1999 to 2007, with a small improvement in Northern and the most pronounced improvement in Eastern Europe. 

Discussion SM survival is high and still increasing in European patients. The gap between Northern and Southern and especially Eastern European countries, although still present, diminished over time. Differences in stage distribution at diagnosis may explain most of the geographical differences. However, part of the improvement in survival may be attributed to overdiagnosis from early diagnosis practices.

Epidermal growth factor receptor immunohistochemistry: new opportunities in metastatic colorectal cancer

The treatment of cancer is becoming more precise, targeting specific oncogenic drivers with targeted molecular therapies. The epidermal growth factor receptor has been found to be over-expressed in a multitude of solid tumours. Immunohistochemistry is widely used in the fields of diagnostic and personalised medicine to localise and visualise disease specific proteins. To date the clinical utility of epidermal growth factor receptor immunohistochemistry in determining monoclonal antibody efficacy has remained somewhat inconclusive. The lack of an agreed reproducible scoring criteria for epidermal growth factor receptor immunohistochemistry has, in various clinical trials yielded conflicting results as to the use of epidermal growth factor receptor immunohistochemistry assay as a companion diagnostic. This has resulted in this test being removed from the licence for the drug panitumumab and not performed in clinical practice for cetuximab. In this review we explore the reasons behind this with a particular emphasis on colorectal cancer, and to suggest a way of resolving the situation through improving the precision of epidermal growth factor receptor immunohistochemistry with quantitative image analysis of digitised images complemented with companion molecular morphological techniques such as in situ hybridisation and section based gene mutation analysis.

All the World's a Stage: Facilitating Discovery Science and Improved Cancer Care through the Global Alliance for Genomics and Health

The recent explosion of genetic and clinical data generated from tumor genome analysis presents an unparalleled opportunity to enhance our understanding of cancer, but this opportunity is compromised by the reluctance of many in the scientific community to share datasets and the lack of interoperability between different data platforms. The Global Alliance for Genomics and Health is addressing these barriers and challenges through a cooperative framework that encourages "team science" and responsible data sharing, complemented by the development of a series of application program interfaces that link different data platforms, thus breaking down traditional silos and liberating the data to enable new discoveries and ultimately benefit patients.

Validation of night blindness reports among children and women in a vitamin A deficient population in rural Tanzania.

OBJECTIVE: This study validates different definitions of reported night blindness (XN) in a vitamin A deficient African population with no local term for XN. DESIGN: Case-control study with follow-up after treatment. SETTING: Eight primary schools and health centres in rural Tanzania. SUBJECTS: A total of 1214 participants were screened for reported XN and other eye signs of xerophthalmia: 461 children aged 24-71 months, 562 primary school-age children and 191 pregnant or breast-feeding women. All 152 cases of reported XN were selected for the validation study and group matched with 321 controls who did not complain of XN. XN reports were validated against serum retinol concentrations and pupillary dark adaptation measurements in cases and controls. INTERVENTION: All children and women who reported XN or had other signs of active xerophthalmia were treated with vitamin A and followed up 3-4 weeks later. Half of the untreated control group who had their serum retinol examined in the baseline examination were also followed up. RESULTS: The overall prevalence of reported XN was 12.5%. At baseline, mean pupillary threshold (-1.52 vs -1.55 log cd/m(2), P=0.501) and median serum retinol concentrations (0.95 vs 0.93 micromol/l, P=0.734) were not significantly different in cases and controls either overall or in each population group. More restricted case definitions reduced the prevalence of reported XN to 5.5% (P<0.001), but there was still no significant difference between cases and controls although the results were in the expected direction. After treatment, the median serum retinol concentration improved significantly only in the most deficient group, the young children. Dark adaptation improved in all the subgroups but the difference was only significant for young children and primary school-age children when the restricted case definitions were used. CONCLUSIONS: XN reports are a poor indicator of vitamin A deficiency in this population. SPONSORSHIP: Task Force Sight and Life, Basel, Switzerland.

Higher plasma levels of complement C3a, C4a and C5a increase the risk of subretinal fibrosis in neovascular age-related macular degeneration.

Background: The aim of this study was to investigate the plasma levels of complement C3a, C4a, and C5a in different types of neovascular age-related macular degeneration (nAMD) and whether the levels were related to patients’ responsiveness to anti-VEGF therapy.

Results: Ninety-six nAMD patients (including 61 with choroidal neovascularisation (CNV), 17 with retinal angiomatous proliferation (RAP), 14 with polypoidal choroidal vasculopathy (PCV) and 4 unclassified patients) and 43 controls were recruited to this case–control study. Subretinal fibrosis was observed in 45 nAMD patients and was absent in 51 nAMD patients. In addition, the responsiveness to anti-VEGF (Lucentis) therapy was also evaluated in nAMD patients. Forty-four patients were complete responders, 48 were partially responders, and only 4 patients did not respond to the therapy. The plasma levels of C3a, C4a and C5a were significantly higher in nAMD patients compared to
controls. Further analysis of nAMD subgroups showed that the levels of C3a, C4a and C5a were significantly increased in patients with CNV but not RAP and PCV. Significantly increased levels of C3a, C4a and C5a were also observed in nAMD patients with subretinal fibrosis but not in those without subretinal fibrosis. Higher levels of C3a were observed in nAMD patients who responded partially to anti-VEGF therapy.

Conclusions: Our results suggest increased systemic complement activation in nAMD patients with CNV but not RAP and PCV. Our results also suggest that higher levels of systemic complement activation may increase the risk of subretinal fibrosis in nAMD patients

Psychosocial interventions to improve quality of life and emotional wellbeing for recently diagnosed cancer patients

BACKGROUND:
A cancer diagnosis may lead to significant psychological distress in up to 75% of cases. There is a lack of clarity about the most effective ways to address this psychological distress.
OBJECTIVES:
To assess the effects of psychosocial interventions to improve quality of life (QoL) and general psychological distress in the 12-month phase following an initial cancer diagnosis.
SEARCH METHODS:
We searched the Cochrane Central Register of Controlled Trials (CENTRAL) (The Cochrane Library 2010, Issue 4), MEDLINE, EMBASE, and PsycINFO up to January 2011. We also searched registers of clinical trials, abstracts of scientific meetings and reference lists of included studies. Electronic searches were carried out across all primary sources of peer-reviewed publications using detailed criteria. No language restrictions were imposed.
SELECTION CRITERIA:
Randomised controlled trials of psychosocial interventions involving interpersonal dialogue between a 'trained helper' and individual newly diagnosed cancer patients were selected. Only trials measuring QoL and general psychological distress were included. Trials involving a combination of pharmacological therapy and interpersonal dialogue were excluded, as were trials involving couples, family members or group formats.
DATA COLLECTION AND ANALYSIS:
Trial data were examined and selected by two authors in pairs with mediation from a third author where required. Where possible, outcome data were extracted for combining in a meta-analyses. Continuous outcomes were compared using standardised mean differences and 95% confidence intervals, using a random-effects model. The primary outcome, QoL, was examined in subgroups by outcome measurement, cancer site, theoretical basis for intervention, mode of delivery and discipline of trained helper. The secondary outcome, general psychological distress (including anxiety and depression), was examined according to specified outcome measures.
MAIN RESULTS:
A total of 3309 records were identified, examined and the trials subjected to selection criteria; 30 trials were included in the review. No significant effects were observed for QoL at 6-month follow up (in 9 studies, SMD 0.11; 95% CI -0.00 to 0.22); however, a small improvement in QoL was observed when QoL was measured using cancer-specific measures (in 6 studies, SMD 0.16; 95% CI 0.02 to 0.30). General psychological distress as assessed by 'mood measures' improved also (in 8 studies, SMD - 0.81; 95% CI -1.44 to - 0.18), but no significant effect was observed when measures of depression or anxiety were used to assess distress (in 6 studies, depression SMD 0.12; 95% CI -0.07 to 0.31; in 4 studies, anxiety SMD 0.05; 95% CI -0.13 to 0.22). Psychoeducational and nurse-delivered interventions that were administered face to face and by telephone with breast cancer patients produced small positive significant effects on QoL (in 2 studies, SMD 0.23; 95% CI 0.04 to 0.43).
AUTHORS' CONCLUSIONS:
The significant variation that was observed across participants, mode of delivery, discipline of 'trained helper' and intervention content makes it difficult to arrive at a firm conclusion regarding the effectiveness of psychosocial interventions for cancer patients. It can be tentatively concluded that nurse-delivered interventions comprising information combined with supportive attention may have a beneficial impact on mood in an undifferentiated population of newly diagnosed cancer patients.

Abdominal Aortic Aneurysm Genetic Associations: Mostly False? A Systematic Review and Meta-analysis

OBJECTIVE/BACKGROUND: Many associations between abdominal aortic aneurysm (AAA) and genetic polymorphisms have been reported. It is unclear which are genuine and which may be caused by type 1 errors, biases, and flexible study design. The objectives of the study were to identify associations supported by current evidence and to investigate the effect of study design on reporting associations.

METHODS: Data sources were MEDLINE, Embase, and Web of Science. Reports were dual-reviewed for relevance and inclusion against predefined criteria (studies of genetic polymorphisms and AAA risk). Study characteristics and data were extracted using an agreed tool and reports assessed for quality. Heterogeneity was assessed using I(2) and fixed- and random-effects meta-analyses were conducted for variants that were reported at least twice, if any had reported an association. Strength of evidence was assessed using a standard guideline.

RESULTS: Searches identified 467 unique articles, of which 97 were included. Of 97 studies, 63 reported at least one association. Of 92 studies that conducted multiple tests, only 27% corrected their analyses. In total, 263 genes were investigated, and associations were reported in polymorphisms in 87 genes. Associations in CDKN2BAS, SORT1, LRP1, IL6R, MMP3, AGTR1, ACE, and APOA1 were supported by meta-analyses.

CONCLUSION: Uncorrected multiple testing and flexible study design (particularly testing many inheritance models and subgroups, and failure to check for Hardy-Weinberg equilibrium) contributed to apparently false associations being reported. Heterogeneity, possibly due to the case mix, geographical, temporal, and environmental variation between different studies, was evident. Polymorphisms in nine genes had strong or moderate support on the basis of the literature at this time. Suggestions are made for improving AAA genetics study design and conduct.

Glycosylation of Vitamin D Binding Protein Reduced in Juvenile Idiopathic Arthritis Patients At Risk of Disease Extension.

Background/Purpose:Juvenile idiopathic arthritis (JIA) comprises a poorly understood group of chronic, childhood onset, autoimmune diseases with variable clinical outcomes. We investigated whether profiling of the synovial fluid (SF) proteome by a fluorescent dye based, two-dimensional gel (DIGE) approach could distinguish the subset of patients in whom inflammation extends to affect a large number of joints, early in the disease process. The post-translational modifications to candidate protein markers were verified by a novel deglycosylation strategy.Methods:SF samples from 57 patients were obtained around time of initial diagnosis of JIA. At 1 year from inclusion patients were categorized according to ILAR criteria as oligoarticular arthritis (n=26), extended oligoarticular (n=8) and polyarticular disease (n=18). SF samples were labeled with Cy dyes and separated by two-dimensional electrophoresis. Multivariate analyses were used to isolate a panel of proteins which distinguish patient subgroups. Proteins were identified using MALDI-TOF mass spectrometry with vitamin D binding protein (VDBP) expression and siaylation further verified by immunohistochemistry, ELISA test and immunoprecipitation. Candidate biomarkers were compared to conventional inflammation measure C-reactive protein (CRP). Sialic acid residues were enzymatically cleaved from immunopurified SF VDBP, enriched by hydrophilic interaction liquid chromatography (HILIC) and analysed by mass spectrometry.Results:Hierarchical clustering based on the expression levels of a set of 23 proteins segregated the extended-to-be oligoarticular from the oligoarticular patients. A cleaved isoform of VDBP, spot 873, is present at significantly reduced levels in the SF of oligoarticular patients at risk of disease extension, relative to other subgroups (p<0.05). Conversely total levels of vitamin D binding protein are elevated in plasma and ROC curves indicate an improved diagnostic sensitivity to detect patients at risk of disease extension, over both spot 873 and CRP levels. Sialysed forms of intact immunopurified VDBP were more prevalent in persistent oligoarticular patient synovial fluids.Conclusion:The data indicate that a subset of the synovial fluid proteome may be used to stratify patients to determine risk of disease extension. Reduced conversion of VDBP to a macrophage activation factor may represent a novel pathway contributing to increased risk of disease extension in JIA patients.

Alpha-2 agonists for sedation of mechanically ventilated adults in intensive care units: a systematic review

BACKGROUND: Care of critically ill patients in intensive care units (ICUs) often requires potentially invasive or uncomfortable procedures, such as mechanical ventilation (MV). Sedation can alleviate pain and discomfort, provide protection from stressful or harmful events, prevent anxiety and promote sleep. Various sedative agents are available for use in ICUs. In the UK, the most commonly used sedatives are propofol (Diprivan(®), AstraZeneca), benzodiazepines [e.g. midazolam (Hypnovel(®), Roche) and lorazepam (Ativan(®), Pfizer)] and alpha-2 adrenergic receptor agonists [e.g. dexmedetomidine (Dexdor(®), Orion Corporation) and clonidine (Catapres(®), Boehringer Ingelheim)]. Sedative agents vary in onset/duration of effects and in their side effects. The pattern of sedation of alpha-2 agonists is quite different from that of other sedatives in that patients can be aroused readily and their cognitive performance on psychometric tests is usually preserved. Moreover, respiratory depression is less frequent after alpha-2 agonists than after other sedative agents.

OBJECTIVES: To conduct a systematic review to evaluate the comparative effects of alpha-2 agonists (dexmedetomidine and clonidine) and propofol or benzodiazepines (midazolam and lorazepam) in mechanically ventilated adults admitted to ICUs.

DATA SOURCES: We searched major electronic databases (e.g. MEDLINE without revisions, MEDLINE In-Process & Other Non-Indexed Citations, EMBASE and Cochrane Central Register of Controlled Trials) from 1999 to 2014.

METHODS: Evidence was considered from randomised controlled trials (RCTs) comparing dexmedetomidine with clonidine or dexmedetomidine or clonidine with propofol or benzodiazepines such as midazolam, lorazepam and diazepam (Diazemuls(®), Actavis UK Limited). Primary outcomes included mortality, duration of MV, length of ICU stay and adverse events. One reviewer extracted data and assessed the risk of bias of included trials. A second reviewer cross-checked all the data extracted. Random-effects meta-analyses were used for data synthesis.

RESULTS: Eighteen RCTs (2489 adult patients) were included. One trial at unclear risk of bias compared dexmedetomidine with clonidine and found that target sedation was achieved in a higher number of patients treated with dexmedetomidine with lesser need for additional sedation. The remaining 17 trials compared dexmedetomidine with propofol or benzodiazepines (midazolam or lorazepam). Trials varied considerably with regard to clinical population, type of comparators, dose of sedative agents, outcome measures and length of follow-up. Overall, risk of bias was generally high or unclear. In particular, few trials blinded outcome assessors. Compared with propofol or benzodiazepines (midazolam or lorazepam), dexmedetomidine had no significant effects on mortality [risk ratio (RR) 1.03, 95% confidence interval (CI) 0.85 to 1.24, I (2) = 0%; p = 0.78]. Length of ICU stay (mean difference -1.26 days, 95% CI -1.96 to -0.55 days, I (2) = 31%; p = 0.0004) and time to extubation (mean difference -1.85 days, 95% CI -2.61 to -1.09 days, I (2) = 0%; p < 0.00001) were significantly shorter among patients who received dexmedetomidine. No difference in time to target sedation range was observed between sedative interventions (I (2) = 0%; p = 0.14). Dexmedetomidine was associated with a higher risk of bradycardia (RR 1.88, 95% CI 1.28 to 2.77, I (2) = 46%; p = 0.001).

LIMITATIONS: Trials varied considerably with regard to participants, type of comparators, dose of sedative agents, outcome measures and length of follow-up. Overall, risk of bias was generally high or unclear. In particular, few trials blinded assessors.

CONCLUSIONS: Evidence on the use of clonidine in ICUs is very limited. Dexmedetomidine may be effective in reducing ICU length of stay and time to extubation in critically ill ICU patients. Risk of bradycardia but not of overall mortality is higher among patients treated with dexmedetomidine. Well-designed RCTs are needed to assess the use of clonidine in ICUs and identify subgroups of patients that are more likely to benefit from the use of dexmedetomidine.

STUDY REGISTRATION: This study is registered as PROSPERO CRD42014014101.

FUNDING: The National Institute for Health Research Health Technology Assessment programme. The Health Services Research Unit is core funded by the Chief Scientist Office of the Scottish Government Health and Social Care Directorates.

Challenging the cancer molecular stratification dogma: Intratumoral heterogeneity undermines consensus molecular subtypes and potential diagnostic value in colorectal cancer

Purpose:
A number of independent gene expression profiling studies have identified transcriptional subtypes in colorectal cancer (CRC) with potential diagnostic utility, culminating in publication of a CRC Consensus Molecular Subtype classification. The worst prognostic subtype has been defined by genes associated with stem-like biology. Recently, it has been shown that the majority of genes associated with this poor prognostic group are stromal-derived. We investigated the potential for tumor misclassification into multiple diagnostic subgroups based on tumoral region sampled.

Experimental Design:
We performed multi-region tissue RNA extraction/transcriptomic analysis using Colorectal Specific Arrays on invasive front, central tumor and lymph node regions selected from tissue samples from 25 CRC patients.

Results:
We identified a consensus 30 gene list which represents the intratumoral heterogeneity within a cohort of primary CRC tumors. Using a series of online datasets, we showed that this gene list displays prognostic potential (HR=2.914 (CI 0.9286-9.162) in stage II/III CRC patients, but in addition we demonstrated that these genes are stromal derived, challenging the assumption that poor prognosis tumors with stem-like biology have undergone a widespread Epithelial Mesenchymal Transition (EMT). Most importantly, we showed that patients can be simultaneously classified into multiple diagnostically relevant subgroups based purely on the tumoral region analysed.

Conclusions:
Gene expression profiles derived from the non-malignant stromal region can influence assignment of CRC transcriptional subtypes, questioning the current molecular classification dogma and highlighting the need to consider pathology sampling region and degree of stromal infiltration when employing transcription-based classifiers to underpin clinical decision-making in CRC.