Large-scale interlaboratory study to develop, analytically validate and apply highly multiplexed, quantitative peptide assays to measure cancer-relevant proteins in plasma

There is an increasing need in biology and clinical medicine to robustly and reliably measure tens-to-hundreds of peptides and proteins in clinical and biological samples with high sensitivity, specificity, reproducibility and repeatability. Previously, we demonstrated that LC-MRM-MS with isotope dilution has suitable performance for quantitative measurements of small numbers of relatively abundant proteins in human plasma, and that the resulting assays can be transferred across laboratories while maintaining high reproducibility and quantitative precision. Here we significantly extend that earlier work, demonstrating that 11 laboratories using 14 LC-MS systems can develop, determine analytical figures of merit, and apply highly multiplexed MRM-MS assays targeting 125 peptides derived from 27 cancer-relevant proteins and 7 control proteins to precisely and reproducibly measure the analytes in human plasma. To ensure consistent generation of high quality data we incorporated a system suitability protocol (SSP) into our experimental design. The SSP enabled real-time monitoring of LC-MRM-MS performance during assay development and implementation, facilitating early detection and correction of chromatographic and instrumental problems. Low to sub-nanogram/mL sensitivity for proteins in plasma was achieved by one-step immunoaffinity depletion of 14 abundant plasma proteins prior to analysis. Median intra- and inter-laboratory reproducibility was <20%, sufficient for most biological studies and candidate protein biomarker verification. Digestion recovery of peptides was assessed and quantitative accuracy improved using heavy isotope labeled versions of the proteins as internal standards. Using the highly multiplexed assay, participating laboratories were able to precisely and reproducibly determine the levels of a series of analytes in blinded samples used to simulate an inter-laboratory clinical study of patient samples. Our study further establishes that LC-MRM-MS using stable isotope dilution, with appropriate attention to analytical validation and appropriate quality c`ontrol measures, enables sensitive, specific, reproducible and quantitative measurements of proteins and peptides in complex biological matrices such as plasma.

Prior and posterior linear pooling for combining expert opinions: Uses and impact on Bayesian Networks

The use of expert knowledge to quantify a Bayesian Network (BN) is necessary when data is not available. This however raises questions regarding how opinions from multiple experts can be used in a BN. Linear pooling is a popular method for combining probability assessments from multiple experts. In particular, Prior Linear Pooling (PrLP), which pools opinions then places them into the BN is a common method. This paper firstly proposes an alternative pooling method, Posterior Linear Pooling (PoLP). This method constructs a BN for each expert, then pools the resulting probabilities at the nodes of interest. Secondly, it investigates the advantages and disadvantages of using these pooling methods to combine the opinions of multiple experts. Finally, the methods are applied to an existing BN, the Wayfinding Bayesian Network Model, to investigate the behaviour of different groups of people and how these different methods may be able to capture such differences. The paper focusses on 6 nodes Human Factors, Environmental Factors, Wayfinding, Communication, Visual Elements of Communication and Navigation Pathway, and three subgroups Gender (female, male),Travel Experience (experienced, inexperienced), and Travel Purpose (business, personal) and finds that different behaviors can indeed be captured by the different methods.

Ants, apples and the ABCs: The use of commercial phonics programmes in prior-to-school children's services

Commercial phonics programmes (e.g. Jolly Phonics and Letterland) are becoming widely used in the early years of school. These programmes claim to use a systematic explicit approach, considered as the preferred method of phonics instruction for teaching alphabetic code-breaking skills in Australia and the UK in the first years of school (Department of Education, Science and Training, 2005; Rose, 2006). However, little is known about the extent to which they are being used in prior-to-school settings, and the reasons behind decisions to use them. This study surveyed 283 early childhood staff in Sydney, Australia and found that commercial phonics programmes were being used in 36% of the early childhood settings surveyed. Staff with early childhood univer- sity qualifications and staff working in not-for-profit service types were less likely to use a commercial phonics programme than staff without university qualifications and staff working in for-profit services. Staff with less than 10 years’ experience were also more likely to use a commercial phonics programme. The rationale behind decisions deter- mining whether or not staff used the programmes ranged from pragmatic reasons, such as parent pressure or higher management decisions, to pedagogical reasons, such as teacher beliefs about how children learn to read and write. The practices staff engage in to teach phonics are explored.

Pre-packaging preschool literacy: What drives early childhood teachers to use commercially produced phonics programs in prior to school settings

Language-rich environments are key to overall quality in early childhood settings, including frequent child–staff interactions around picture books and dramatic play. In a language-rich environment, explicit teaching of literacy concepts, such as phonics, is embedded in authentic and meaningful situations where alphabet letters and sounds are taught in a context meaningful to the child. Recent research, however, suggests that the use of commercial pre-packaged phonics programs (such as Letterland and Jolly Phonics) is widespread in prior to school settings in Sydney, Australia. Little is known about why early childhood teachers choose to use such programs with children aged five and under. In the present study, thematic analysis of data from interviews with five early childhood teachers using commercial phonics programs found that their reasons were pragmatic rather than pedagogical. Motivations included the idea that the programs reduced their workload, provided tangible evidence to parents of their child’s ‘school readiness’, and served as a marketing tool to attract parents. Further analysis found that the teachers were unable to articulate what phonics and phonological awareness are and how they are learnt in early childhood.

Feeling the pressure: Early childhood educators’ reported views about learning and teaching phonics in Australian prior-to-school settings

Early childhood educators’ beliefs about literacy teaching can impact on the types of phonics experiences educators provide for children in prior-to-school settings. The Australian Early Years Learning Framework supports a play-based, intentional approach to teaching phonics, however little is known about what Australian early childhood educators believe is important in teaching phonics in the prior-to-school years. Using a qualitative content analysis, this research study investigates 115 early childhood educators’ views about how phonics should be taught and the use of commercially produced phonics programs (e.g. Jolly Phonics and Letterland) in prior-to-school settings. This study further investigates educators’ perceived pressures to include structured phonics lessons, as a way of addressing parental notions of ‘school readiness’. The results of this study indicate conflicting views were held about how phonics should be taught. Some educators also experienced external pressures to engage in literacy practices that may be in opposition with their own beliefs about how literacy is learnt. This study provides insights into the pedagogical practices early childhood educators believe are appropriate when teaching phonics. The educational implications are discussed.

Being here: Designing for distributed hands-on collaboration in blended interaction spaces

This paper describes a concept for supporting distributed hands-on collaboration through interaction design for the physical and the digital workspace. The Blended Interaction Spaces concept creates distributed work environments in which collaborating parties all feel that they are present “here” rather than “there”. We describe thinking and inspirations behind the Blended Interaction Spaces concept, and summarize findings from fieldwork activities informing our design. We then exemplify the Blended Interaction Spaces concept through a prototype implementation of one of four concepts.

A phase II trial of brivanib in recurrent or persistent endometrial cancer: An NRG oncology/gynecologic oncology group study

PURPOSE Brivanib, an oral, multi-targeted tyrosine kinase inhibitor with activity against vascular endothelial growth factor (VEGF) and fibroblast growth factor receptor (FGFR) was investigated as a single agent in a phase II trial to assess the activity and tolerability in recurrent or persistent endometrial cancer (EMC). PATIENTS AND METHODS Eligible patients had persistent or recurrent EMC after receiving one to two prior cytotoxic regimens, measurable disease, and performance status of ≤2. Treatment consisted of brivanib 800 mg orally every day until disease progression or prohibitive toxicity. Primary endpoints were progression-free survival (PFS) at six months and objective tumor response. Expression of multiple angiogenic proteins and FGFR2 mutation status was assessed. RESULTS Forty-five patients were enrolled. Forty-three patients were eligible and evaluable. Median age was 64 years. Twenty-four patients (55.8%) received prior radiation. Median number of cycles was two (range 1-24). No GI perforations but one rectal fistula were seen. Nine patients had grade 3 hypertension, with one experiencing grade 4 confusion. Eight patients (18.6%; 90% CI 9.6%-31.7%) had responses (one CR and seven PRs), and 13 patients (30.2%; 90% CI 18.9%-43.9%) were PFS at six months. Median PFS and overall survival (OS) were 3.3 and 10.7 months, respectively. When modeled jointly, VEGF and angiopoietin-2 expression may diametrically predict PFS. Estrogen receptor-α (ER) expression was positively correlated with OS. CONCLUSION Brivanib is reasonably well tolerated and worthy of further investigation based on PFS at six months in recurrent or persistent EMC.

Bayesian computational methods for spatial analysis of images

This thesis introduces a new way of using prior information in a spatial model and develops scalable algorithms for fitting this model to large imaging datasets. These methods are employed for image-guided radiation therapy and satellite based classification of land use and water quality. This study has utilized a pre-computation step to achieve a hundredfold improvement in the elapsed runtime for model fitting. This makes it much more feasible to apply these models to real-world problems, and enables full Bayesian inference for images with a million or more pixels.

The psychosocial purpose of driving and its relationship with the risky driving behaviour of young novice drivers

Background The overrepresentation of young drivers in road crashes, injuries and fatalities around the world has resulted in a breadth of injury prevention efforts including education, enforcement, engineering, and exposure control. Despite multifaceted intervention, the young driver problem remains a challenge for injury prevention researchers, practitioners and policy-makers. The intractable nature of young driver crash risks suggests that a deeper understanding of their car use – that is, the purpose of their driving – is required to inform the design of more effective young driver countermeasures. Aims This research examined the driving purpose reported by young drivers, including the relationship with self-reported risky driving behaviours including offences. Methods Young drivers with a Learner or Provisional licence participated in three online surveys (N1 = 656, 17–20 years; N2 = 1051, 17–20 years; N3 = 351, 17–21 years) as part of a larger state-wide project in Queensland, Australia. Results A driving purpose scale was developed (the PsychoSocial Purpose Driving Scale, PSPDS), revealing that young drivers drove for psychosocial reasons such as for a sense of freedom and to feel independent. Drivers who reported the greatest psychosocial purpose for driving were more likely to be male and to report more risky driving behaviours such as speeding. Drivers who deliberately avoided on-road police presence and reported a prior driving-related offence had significantly greater PSPDS scores, and higher reporting of psychosocial driving purposes was found over time as drivers transitioned from the supervised Learner licence phase to the independent Provisional (intermediate) licence phase. Discussion and conclusions The psychosocial needs met by driving suggest that effective intervention to prevent young driver injury requires further consideration of their driving purpose. Enforcement, education, and engineering efforts which consider the psychosocial purpose of the driving are likely to be more efficacious than those which presently do not. Road safety countermeasures could reduce the young driver’s exposure to risk through such mechanisms as encouraging the use of public transport.

Dr. Matthew Rimmer takes a closer look at Fair Use

For this week's Favorites of the Week, Dr. Matthew Rimmer wanted to focus in on some of the specific points that came up in the Congressional hearing on fair use earlier this week. While a bit different than our usual "favorites of the week" post, it's a really fantastic in-depth look at some of the issues, which I'm sure many of you will enjoy.

Australia's Biodiversity Conservation Strategy 2010-2020 Submission to the National Biodiversity Strategy Review Taskforce

This submission focuses on “Priority for Change 3: Knowledge for all” and “Priority for Change 5: Involving Indigenous Australians”. Our particular interest lies with ensuring that Indigenous knowledge holders are engaged with in a manner that recognises their prior rights over their own knowledge and intellectual property. As the Preamble of the recently endorsed United Nations Declaration on the Rights of Indigenous Peoples states, we need to; “Recognis[e] that respect for Indigenous Knowledge, cultures and traditional practises contributes to sustainable and equitable development and proper management of the environment”.

Damned to fame: The moral rights of the Beckett estate

The Company B production of Samuel Beckett's Waiting for Godot raises important questions about copyright law, moral rights, and dramatic works. The playwright's nephew and executor, Edward Beckett, threatened to bring a legal action against the Sydney company for breach of contract on the grounds that unauthorised music appeared in the production. The Company B production denied that the contract made any such express provisions. The director Neil Armfield complained: 'In coming here with its narrow prescriptions, its dead controlling hand, the Beckett estate seems to me to be the enemy of art'. In the biography Damned to fame, James Knowlson documents a number of other proceedings taken by Beckett and his agents to control the productions of his work: 'He was often represented as a tyrannical figure, an arch-controller of his work, ready to unleash fiery thunderbolts onto the head of any bold, innovative director, unwilling to follow his text and stage directions to the last counted dot and precisely timed pause.' However, Knowlson notes that Beckett was inconsistent in his willingness to use legal action: 'It made a tremendous difference if he liked and respected the persons involved or if he had been able to listen to their reasons for wanting to attempt something highly innovative or even slightly different'. Famously, in 1988, Beckett brought legal action against a Dutch theatre company, which wanted to stage a production of Waiting for Godot, with women acting all the roles. His lawyer argued that the integrity of the text was violated because actresses were substituted for the male actors asked for in the text. The judge in the Haarlem court ruled that the integrity of the play had not been violated, because the performance showed fidelity to the dialogue and the stage directions of the play. By contrast, in 1992, a French court held a stage director was liable for an infringement of Beckett's moral right of integrity because the director had staged Waiting for Godot with the two lead roles played by women. In 1998, a United States production of Waiting for Godot with a racially mixed cast attracted legal threats amid accusations it had 'injected race into the play'. In the 2000 New York Fringe Festival, a company made light of this ongoing conflict between the Beckett estate and artistic directors. The work was entitled: The complete lost works of Samuel Beckett as found in an envelope (partially burned) in a dustbin in Paris labelled 'Never to be performed. Never. Ever. EVER! Or I'll sue! I'LL SUE FROM THE GRAVE!'. The plot concerned a fight between three producers and the Beckett estate. In the wake of such disputes, Beckett and later his estate sought to tighten production contracts to state that no additions, omissions or alterations should be made to the text of the play or the stage directions and that no music, special effects or other supplements should be added without prior consent.

Eccentric knee-flexor strength and risk of hamstring injuries in Rugby Union: A prospective study

BACKGROUND Hamstring strain injuries (HSIs) represent the most common cause of lost playing time in rugby union. Eccentric knee-flexor weakness and between-limb imbalance in eccentric knee-flexor strength are associated with a heightened risk of hamstring injury in other sports; however these variables have not been explored in rugby union. PURPOSE To determine if lower levels of eccentric knee-flexor strength or greater between-limb imbalance in this parameter during the Nordic hamstring exercise are risk-factors for hamstring strain injury in rugby union. STUDY DESIGN Cohort study; level of evidence, 3. METHODS This prospective study was conducted over the 2014 Super Rugby and Queensland Rugby Union seasons. In total, 178 rugby union players (age, 22.6 ± 3.8 years; height, 185 ± 6.8 cm; mass, 96.5 ± 13.1 kg) had their eccentric knee-flexor strength assessed using a custom-made device during the pre-season. Reports of previous hamstring, quadriceps, groin, calf and anterior cruciate ligament injury were also obtained. The main outcome measure was prospective occurrence of hamstring strain injury. RESULTS Twenty players suffered at least one hamstring strain during the study period. Players with a history of hamstring strain injury had 4.1 fold (RR = 4.1, 95% CI = 1.9 to 8.9, p = 0.001) greater risk of subsequent hamstring injury than players without such history. Between-limb imbalance in eccentric knee-flexor strength of ≥ 15% and ≥ 20% increased the risk of hamstring strain injury 2.4 fold (RR = 2.4, 95% CI = 1.1 to 5.5, p = 0.033) and 3.4 fold (RR = 3.4, 95% CI = 1.5 to 7.6, p = 0.003), respectively. Lower eccentric knee flexor strength and other prior injuries were not associated with increased risk of future hamstring strain. Multivariate logistic regression revealed that the risk of re-injury was augmented in players with strength imbalances. CONCLUSION Previous hamstring strain injury and between-limb imbalance in eccentric knee-flexor strength were associated with an increased risk of future hamstring strain injury in rugby union. These results support the rationale for reducing imbalance, particularly in players who have suffered a prior hamstring injury, to mitigate the risk of future injury.

Progress in the genetics of ankylosing spondylitis

Ankylosing spondylitis (AS) is a common, highly heritable, inflammatory arthropathy. In addition to being strongly associated with HLA-B27, a further 13 genes have been robustly associated with the disease. These genes highlight the involvement of the IL-23 pathway in disease pathogenesis, and indicate overlaps between the pathogenesis of AS, and of inflammatory bowel disease. Genetic associations in B27-positive and -negative disease are similar, with the main exception of association with ERAP1, which is restricted in association to B27-positive cases. This restriction, and the known function of ERAP1 in peptide trimming prior to HLA Class I presentation, indicates that HLA-B27 is likely to operate in AS by a mechanism involving aberrant peptide handling. These advances point to several potential novel therapeutic approaches in AS.

The revolution in human monogenic disease mapping

The successful completion of the Human Genome Project (HGP) was an unprecedented scientific advance that has become an invaluable resource in the search for genes that cause monogenic and common (polygenic) diseases. Prior to the HGP, linkage analysis had successfully mapped many disease genes for monogenic disorders; however, the limitations of this approach were particularly evident for identifying causative genes in rare genetic disorders affecting lifespan and/or reproductive fitness, such as skeletal dysplasias. In this review, we illustrate the challenges of mapping disease genes in such conditions through the ultra-rare disorder fibrodysplasia ossificans progressiva (FOP) and we discuss the advances that are being made through current massively parallel (“next generation”) sequencing (MPS) technologies.