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Abstract Background Diagnostic autopsy is the most reliable approach to definitively ascertain the cause of death and evaluate the accuracy of antemortem clinical diagnoses. Identifying diagnostic discrepancies is vital to understanding common gaps in antemortem clinical diagnoses and modifying antemortem diagnostic approaches to increase the accuracy of clinical diagnosis. The objective of this study was to determine the frequency of diagnostic discrepancies between antemortem clinical diagnoses and postmortem autopsies on lung pathologies and to understand the reasons for diagnostic discrepancies among cases included in Child Health and Mortality Prevention Surveillance (CHAMPS) in Ethiopia. Methods A clinical case series study of deaths among children under-five in the CHAMPS study at three sites in Ethiopia between October 2019 and April 2022 was conducted. The antemortem clinical diagnoses and postmortem pathological diagnoses of the lung were compared for each case. Two senior physicians assessed the findings for both agreement and disagreement. McNemar’s test was used to assess for statistically significant differences between antemortem and postmortem diagnoses. Results Seventy-five cases were included (73.3% male). Over half (54.7%) died between the 1st and 7th day of life. Sepsis (66.7%), pneumonia (6.7%), and meconium aspiration syndrome (5.0%) were the most common immediate causes of death. Half (52%) of cases were correctly diagnosed antemortem. The magnitude of diagnostic discrepancy was 35% (95% CI: 20-47%). The most common contributing factors to diagnostic discrepancy were gaps in knowledge (22/75, 35.5%) and problems in consultation and teamwork (22/75, 35.5%). Conclusions Misdiagnoses were common among young children who died with positive lung pathology findings. In-service education initiatives and multidisciplinary collaboration are needed to mitigate high rates of diagnostic discrepancies among young children to potentially prevent future deaths.

IntroductionThe therapeutic potential of bispecific antibodies is becoming widely recognised, with over a hundred formats already described. For many applications, enhanced tissue penetration is sought, so bispecifics with low molecular weight may offer a route to enhanced potency. Here we report the design of bi- and tri-specific antibody-based constructs with molecular weights as low as 14.5 and 22 kDa respectively.MethodsAutonomous bovine ultra-long CDR H3 (knob domain peptide) modules have been engineered with artificial coiled-coil stalks derived from Sin Nombre orthohantavirus nucleocapsid protein and human Beclin-1, and joined in series to produce bi- and tri-specific antibody-based constructs with exceptionally low molecular weights.ResultsKnob domain peptides with coiled-coil stalks retain high, independent antigen binding affinity, exhibit exceptional levels of thermal stability, and can be readily joined head-to-tail yielding the smallest described multi-specific antibody format. The resulting constructs are able to bind simultaneously to all their targets with no interference.DiscussionCompared to existing bispecific formats, the reduced molecular weight of the knob domain fusions may enable enhanced tissue penetration and facilitate binding to cryptic epitopes that are inaccessible to conventional antibodies. Furthermore, they can be easily produced at high yield as recombinant products and are free from the heavy-light chain mispairing issue. Taken together, our approach offers an efficient route to modular construction of minimalistic bi- and multi-specifics, thereby further broadening the therapeutic scope for knob domain peptides.

In resource-limited settings where vital registration and medical death certificates are unavailable or incomplete, verbal autopsy (VA) is often used to attribute causes of death (CoD) and prioritize resource allocation and interventions. We aimed to determine the CoD concordance between InterVA and CHAMPS's method. The causes of death (CoDs) of children

ABSTRACTGremlin-1, a high-affinity antagonist of bone morphogenetic proteins (BMP)-2, −4, and −7, is implicated in tumor initiation and progression. Increased gremlin-1 expression, and therefore suppressed BMP signaling, correlates with poor prognosis in a range of cancer types. A lack of published work using therapeutic modalities has precluded the testing of the hypothesis that blocking the gremlin-1/BMP interaction will provide benefits to patients. To address this shortfall, we developed ginisortamab (UCB6114), a first-in-class clinical anti-human gremlin-1 antibody, currently in clinical development for the treatment of cancer, along with its murine analog antibody Ab7326 mouse immunoglobulin G1 (mIgG1). Surface plasmon resonance assays revealed that ginisortamab and Ab7326 mIgG1 had similar affinities for human and mouse gremlin-1, with mean equilibrium dissociation constants of 87 pM and 61 pM, respectively. The gremlin-1/Ab7326 antigen-binding fragment (Fab) crystal structure revealed a gremlin-1 dimer with a Fab molecule bound to each monomer that blocked BMP binding. In cell culture experiments, ginisortamab fully blocked the activity of recombinant human gremlin-1, and restored BMP signaling pathways in human colorectal cancer (CRC) cell lines. Furthermore, in a human CRC – fibroblast co-culture system where gremlin-1 is produced by the fibroblasts, ginisortamab restored BMP signaling in both the CRC cells and fibroblasts, demonstrating its activity in a relevant human tumor microenvironment model. The safety and efficacy of ginisortamab are currently being evaluated in a Phase 1/2 clinical trial in patients with advanced solid tumors (NCT04393298).

Interleukin-13 (IL-13) is a cytokine involved in T-cell immune responses and is a well validated therapeutic target for the treatment of asthma, along with other allergic and inflammatory diseases. IL-13 signals through a ternary signalling complex formed with the receptors IL-13Rα1 and IL-4Rα. This complex is assembled by IL-13 initially binding IL-13Rα1, followed by association of the binary IL-13:IL-13Rα1 complex with IL-4Rα. The receptors are shared with IL-4, but IL-4 initially binds IL-4Rα. Here we report the identification and characterisation of a diverse panel of single-domain antibodies (VHHs) that bind to IL-13 (KD 40 nM-5.5 μM) and inhibit downstream IL-13 signalling (IC50 0.2-53.8 μM). NMR mapping showed that the VHHs recognise a number of epitopes on IL-13, including previously unknown allosteric sites. Further NMR investigation of VHH204 bound to IL-13 revealed a novel allosteric mechanism of inhibition, with the antibody stabilising IL-13 in a conformation incompatible with receptor binding. This also led to the identification of a conformational equilibrium for free IL-13, providing insights into differing receptor signalling complex assembly seen for IL-13 compared to IL-4, with formation of the IL-13:IL-13Rα1 complex required to stabilise IL-13 in a conformation with high affinity for IL-4Rα. These findings highlight new opportunities for therapeutic targeting of IL-13 and we report a successful 19F fragment screen of the IL-13:VHH204 complex, including binding sites identified for several hits. To our knowledge, these 19F containing fragments represent the first small-molecules shown to bind to IL-13 and could provide starting points for a small-molecule drug discovery programme.

Abstract Background Lynch syndrome (LS) is a cancer predisposition syndrome affecting more than 1 in every 300 individuals worldwide. Clinical genetic testing for LS can be life-saving but is complicated by the heavy burden of variants of uncertain significance (VUS), especially missense changes. Result To address this challenge, we leverage a multiplexed analysis of variant effect (MAVE) map covering >94% of the 17,746 possible missense variants in the key LS gene MSH2. To establish this map’s utility in large-scale variant reclassification, we overlay it on clinical databases of >15,000 individuals with LS gene variants uncovered during clinical genetic testing. We validate these functional measurements in a cohort of individuals with paired tumor-normal test results and find that MAVE-based function scores agree with the clinical interpretation for every one of the MSH2 missense variants with an available classification. We use these scores to attempt reclassification for 682 unique missense VUS, among which 34 scored as deleterious by our function map, in line with previously published rates for other cancer predisposition genes. Combining functional data and other evidence, ten missense VUS are reclassified as pathogenic/likely pathogenic, and another 497 could be moved to benign/likely benign. Finally, we apply these functional scores to paired tumor-normal genetic tests and identify a subset of patients with biallelic somatic loss of function, reflecting a sporadic Lynch-like Syndrome with distinct implications for treatment and relatives’ risk. Conclusion This study demonstrates how high-throughput functional assays can empower scalable VUS resolution and prospectively generate strong evidence for variant classification.

Mutations in the nucleocapsid of SARS-CoV-2 may interfere with antigen detection by diagnostic tests. We used several methods to evaluate the effect of various SARS-CoV-2 nucleocapsid mutations on the performance of the Panbio™ and BinaxNOW™ lateral flow rapid antigen tests and a prototype high-throughput immunoassay that utilizes Panbio antibodies. Variant detection was also evaluated by immunoblot and BIAcore™ assay. A panel of 23 recombinant nucleocapsid antigens (rAgs) were produced that included mutations found in circulating SARS-CoV-2 variants, including variants of concern. All mutant rAgs were detected by all assays, at a sensitivity equivalent to wild-type control (Wuhan strain). Thus, using a rAg approach, we found that the SARS-CoV-2 nucleocapsid mutations examined do not directly impact antigen detection or antigen assay performance.

BackgroundSerum albumin binding is an established mechanism to extend the serum half-life of antibody fragments and peptides. The cysteine rich knob domains, isolated from bovine antibody ultralong CDRH3, are the smallest single chain antibody fragments described to date and versatile tools for protein engineering.MethodsHere, we used phage display of bovine immune material to derive knob domains against human and rodent serum albumins. These were used to engineer bispecific Fab fragments, by using the framework III loop as a site for knob domain insertion. ResultsBy this route, neutralisation of the canonical antigen (TNFα) was retained but extended pharmacokinetics in-vivo were achieved through albumin binding. Structural characterisation revealed correct folding of the knob domain and identified broadly common but non-cross-reactive epitopes. Additionally, we show that these albumin binding knob domains can be chemically synthesised to achieve dual IL-17A neutralisation and albumin binding in a single chemical entity. ConclusionsThis study enables antibody and chemical engineering from bovine immune material, via an accessible discovery platform.

Objectives Personality differences between doctors and patients can affect treatment outcomes. We examine these trait disparities, as well as differences across medical specialities.Design Retrospective, observational statistical analysis of secondary data.Setting Data from two data sets that are nationally representative of doctors and the general population in Australia.Participants We include 23 358 individuals from a representative survey of the general Australian population (with subgroups of 18 705 patients, 1261 highly educated individuals and 5814 working in caring professions) as well as 19 351 doctors from a representative survey of doctors in Australia (with subgroups of 5844 general practitioners, 1776 person-oriented specialists and 3245 technique-oriented specialists).Main outcome measures Big Five personality traits and locus of control. Measures are standardised by gender, age and being born overseas and weighted to be representative of their population.Results Doctors are significantly more agreeable (a: standardised score −0.12, 95% CIs −0.18 to −0.06), conscientious (c: −0.27 to –0.33 to −0.20), extroverted (e: 0.11, 0.04 to 0.17) and neurotic (n: 0.14, CI 0.08 to 0.20) than the general population (a: −0.38 to –0.42 to −0.34, c: −0.96 to –1.00 to −0.91, e: −0.22 to –0.26 to −0.19, n: −1.01 to –1.03 to −0.98) or patients (a: −0.77 to –0.85 to −0.69, c: −1.27 to –1.36 to −1.19, e: −0.24 to –0.31 to −0.18, n: −0.71 to –0.76 to −0.66). Patients (−0.03 to –0.10 to 0.05) are more open than doctors (−0.30 to –0.36 to −0.23). Doctors have a significantly more external locus of control (0.06, 0.00 to 0.13) than the general population (−0.10 to –0.13 to −0.06) but do not differ from patients (−0.04 to –0.11 to 0.03). There are minor differences in personality traits among doctors with different specialities.Conclusions Several personality traits differ between doctors, the population and patients. Awareness about differences can improve doctor–patient communication and allow patients to understand and comply with treatment recommendations.

Despite their low cost, the use of email invitations to distribute surveys to medical practitioners have been associated with lower response rates. This research compares the difference in response rates from using email approach plus online completion rather than a mailed invitation letter plus a choice of online or paper completion. A parallel randomised controlled trial was conducted during the 11th annual wave of the nationally representative Medicine in Australia: Balancing Employment and Life (MABEL) longitudinal survey of doctors. The control group was invited using a mailed paper letter (including a paper survey plus instructions to complete online) and three mailed paper reminders. The intervention group was approached in the same way apart from the second reminder when they were approached by email only. The primary outcome is the response rate and the statistical analysis was blinded. 18,247 doctors were randomly allocated to the control (9,125) or intervention group (9,127), with 9,108 and 9,107 included in the analysis. Using intention to treat analysis, the response rate in the intervention group was 35.92% compared to 37.59% in the control group, a difference of -1.66 percentage points (95% CI: -3.06 to -0.26). The difference was larger for General Practitioners (-2.76 percentage points, 95% CI: -4.65 to -0.87) compared to other specialists (-0.47 percentage points, 95% CI: -2.53 to 1.60). For those who supplied an email address, the average treatment effect on the treated was higher at -2.63 percentage points (95% CI: -4.50 to -0.75) for all physicians, -3.17 percentage points (95% CI: -5.83 to -0.53) for General Practitioners, and -2.1 percentage points (95% CI: -4.75 to 0.56) for other specialists. For qualified physicians, using email to invite participants to complete a survey leads to lower response rates compared to a mailed letter. Lower response rates need to be traded off with the lower costs of using email rather than mailed letters.

Abstract Purpose This study presents patient-specific quality assurance (QA) results from the first 395 clinical cases for the new helical TomoTherapy® platform (Radixact) coupled with dedicated Precision TPS. Methods The passing rate of the Gamma Index (GP%) of 395 helical QA of patient-specific tomotherapy, acquired with ArcCHECK, is presented, analysed and correlated to various parameters of the plan. Following TG-218 recommendations, the clinic specific action limit (ALcs) and tolerance limit (TLcs) were calculated for our clinic and monitored during the analysed period. Results The mean values ​​(± 1 standard deviation) of GP% (3%/2 mm) (both global and local normalization) are: 97.6% and 90.9%, respectively. The proposed ALcs and TLcs, after a period of two years’ process monitoring are 89.4% and 91.1% respectively. Conclusions The phantom measurements closely match the planned dose distributions, demonstrating that the calculation accuracy of the new Precision TPS and the delivery accuracy of the Radixact unit are adequate, with respect to international guidelines and reports. Furthermore, a first correlation with the planning parameters was made. Action and tolerance limits have been set for the new Radixact Linac.

Objective To examine the association between financial incentives from entry into a vaccine competition with the probability of vaccination for COVID-19.Design A cross-sectional study with adjustment for covariates using logistic regression.Setting October and November 2021, Australia.Participants 2375 respondents of the Taking the Pulse of the Nation survey.Primary and secondary outcome measures The proportion of respondents who had any vaccination, a first dose only, or second dose after the competition opened.Results Those who entered the competition were 2.27 (95% CI 1.73 to 2.99) times more likely to be vaccinated after the competition opened on 1 October than those who did not enter—an increase in the probability of having any dose of 0.16 (95 % CI 0.10 to 0.21) percentage points. This increase was mostly driven by those receiving second doses. Entrants were 2.39 (95% CI 1.80 to 3.17) times more likely to receive their second dose after the competition opened.Conclusions Those who entered the Million Dollar Vax competition were more likely to have a vaccination after the competition opened compared with those who did not enter the competition, with this effect dominated by those receiving second doses.

Abstract Background This study investigated if the evidence on the success of the Pay for Performance (P4P) schemes in healthcare is changing as the schemes continue to evolve by updating a previous systematic review. Methods A meta-regression analysis using 116 studies evaluating P4P schemes published between January 2010 to February 2018. The effects of the research design, incentive schemes, use of incentives, and the size of the payment to revenue ratio on the proportion of statically significant effects in each study were examined. Results There was evidence of an increase in the range of countries adopting P4P schemes and weak evidence that the proportion of studies with statistically significant effects have increased. Factors hypothesized to influence the success of schemes have not changed. Studies evaluating P4P schemes which made payments for improvement over time, were associated with a lower proportion of statistically significant effects. There was weak evidence of a positive association between the incentives’ size and the proportion of statistically significant effects. Conclusion The evidence on the effectiveness of P4P schemes is evolving slowly, with little evidence that lessons are being learned concerning the design and evaluation of P4P schemes.

TNF can be inhibited by small molecules that stabilize the TNF trimer in an asymmetric conformation. Here, the authors develop a monoclonal antibody that selectively binds this inactive form of TNF, enabling both target engagement assessment and structural characterization of TNF binding to TNF receptor 1.

Abstract Background The public-private mix of healthcare remains controversial. This paper examines physicians’ preferences for public sector work in the context of dual practice, whilst accounting for other differences in the characteristics of jobs. Methods A discrete choice experiment is conducted with data from 3422 non-GP specialists from the Medicine in Australia: Balancing Employment and Life (MABEL) panel survey of physicians. Results Physicians prefer to work in the public sector, though the value of working in the public sector is very small at 0.14% of their annual earnings to work an additional hour per week. These preferences are heterogeneous. Contrary to other studies that show risk averse individuals prefer public sector work, for physicians, we find that those averse to taking career or clinical risks prefer to work in the private sector. Those with relatively low earnings prefer public sector work and those with high earnings prefer private sector work, though these effects are small. Conclusions Other job characteristics are more important than the sector of work, suggesting that these should be the focus of policy to influence specialist’s allocation of time between sectors.

Abstract Background The geographic distribution of health workers is a pervasive policy concern. Many governments are responding by introducing financial incentives to attract health care workers to locate in areas that are underserved. However, clear evidence of the effectiveness of such financial incentives is lacking. Methods This paper examines General Practitioners’ (GPs) relocation choices in Australia and proposes a dynamic location choice model accounting for both source and destination factors associated with a choice to relocate, thereby accounting for push and pull factors associated with job separation. The model is used to simulate financial incentive policies and assess potential for such policies to redistribute GPs. This paper examines the role of financial factors in relocating established GPs into neighbourhoods with relatively low socioeconomic status. The paper uses a discrete choice model and panel data on GPs’ actual changes in location from one year to the next. Results This paper finds that established GPs are not very mobile, even when a financial incentive is offered. Policy simulation predicts that 93.2% of GPs would remain at their current practice and that an additional 0.8% would be retained or would relocate in a low-socioeconomic status (SES) neighbourhood in response to a hypothetical financial incentive of a 10% increase in the earnings of all metropolitan GPs practising in low-SES neighbourhoods. Conclusion With current evidence on the effectiveness of redistribution programmes limited to newly entering GPs, the policy simulations in this paper provide an insight into the potential effectiveness of financial incentives as a redistribution policy targeting the entire GP population. Overall, the results suggest that financial considerations are part of many factors influencing the location choice of GPs. For instance, GP practice ownership played almost as important a role in mobility as earnings.

Abstract Background We investigate factors affecting Australian general practitioners’ decisions to adopt novel oral anticoagulants (NOACs) for the prevention of stroke/systemic embolism among patients with atrial fibrillation. Australia has a national homogeneous review and coverage system, which enables us to distinguish physician level factors while maintaining system level factors and patient coverage information constant. Methods We conduct a cohort analyses by using longitudinal physician survey data from the Medicine in Australia: Balancing Employment and Life panel survey of Australian physicians (MABEL). MABEL data contain rich physician-level information such as age, gender, education, risk preferences, personality, physicians’ communications with other medical professionals, and other practice characteristics. Importantly, the survey data were linked, with physician’s consent, to actual utilization data from the Australian Pharmaceutical Benefits Scheme and the Medicare Benefits Schedule between January 1, 2012 and December 31, 2015. We measure speed (days until first time prescribing) of adopting NOACs. We estimate a Cox proportional hazard model to estimate factors affecting the adoption speed. Results Several factors predict earlier adoption of NOACs: being male, more likely to take clinical risk, higher prescribing volume, being a principal or partner in the practice instead of an employee, spending less time in a typical consultation, and practicing in more affluent areas or areas with a higher proportion of older patients. GPs in Queensland are more likely to adopt NOACs and more likely to be extensive early adopters compared to other GPs. Other characteristics including physician personality, family circumstances, their involvement with public hospitals and teaching activities, and the distance between physician practice location to other clinics in the area are not statistically associated with earlier adoption. Conclusions Our paper is one of the first to study the relationship between GPs’ risk preferences, personality and their decisions to adopt new prescription drugs. Because NOACs are commonly prescribed and considered more cost-effective than their older counterpart, understanding factors affecting physicians’ decisions to adopt NOACs has direct policy implications. Our results also highlight that even with universal coverage for prescription drugs, access to new drugs is different among patients, partially because who their doctors are and where they practice.

Abstract Background The capacity for high-income countries to supply enough locally trained doctors to minimise their reliance on overseas-trained doctors (OTDs) is important for equitable global workforce distribution. However, the ability to achieve self-sufficiency of individual countries is poorly evaluated. This review draws on a decade of research evidence and applies additional stratified analyses from a unique longitudinal medical workforce research program (the Medicine in Australia: Balancing Employment and Life survey (MABEL)) to explore Australia’s rural medical workforce self-sufficiency and inform rural workforce planning. Australia is a country with a strong medical education system and extensive rural workforce policies, including a requirement that newly arrived OTDs work up to 10 years in underserved, mostly rural, communities to access reimbursement for clinical services through Australia’s universal health insurance scheme, called Medicare. Findings Despite increases in the number of Australian-trained doctors, more than doubling since the late 1990s, recent locally trained graduates are less likely to work either as general practitioners (GPs) or in rural communities compared to local graduates of the 1970s–1980s. The proportion of OTDs among rural GPs and other medical specialists increases for each cohort of doctors entering the medical workforce since the 1970, peaking for entrants in 2005–2009. Rural self-sufficiency will be enhanced with policies of selecting rural-origin students, increasing the balance of generalist doctors, enhancing opportunities for remaining in rural areas for training, ensuring sustainable rural working conditions and using innovative service models. However, these policies need to be strongly integrated across the long medical workforce training pathway for successful rural workforce supply and distribution outcomes by locally trained doctors. Meanwhile, OTDs substantially continue to underpin Australia’s rural medical service capacity. The training pathways and social support for OTDs in rural areas is critical given their ongoing contribution to Australia’s rural medical workforce. Conclusion It is essential for Australia to monitor its ongoing reliance on OTDs in rural areas and be considerate of the potential impact on global workforce distribution.

Objective To investigate factors associated with the use of telehealth by general practitioners (GPs) during COVID-19.Design A nationally representative longitudinal survey study of Australian doctors analysed using regression analysis.Setting General practice in Australia during the COVID-19 pandemic.Participants 448 GPs who completed both the 11th wave (2018–2019) of the Medicine in Australia: Balancing Employment and Life (MABEL) Survey and the MABEL COVID-19 Special Online Survey (May 2020).Outcome measures Proportion of all consultations delivered via telephone (audio) or video (audiovisual); proportion of telehealth consultations delivered via video.Results 46.1% of GP services were provided using telehealth in early May 2020, with 6.4% of all telehealth consultations delivered via video. Higher proportions of telehealth consultations were observed in GPs in larger practices compared with solo GPs: between +0.21 (95% CI +0.07 to +0.35) and +0.28 (95% CI +0.13 to +0.44). Greater proportions of telehealth consultations were delivered through video for GPs with appropriate infrastructure and for GPs with more complex patients: +0.10 (95% CI +0.04 to +0.16) and +0.04 (95% CI +0.00 to +0.08), respectively. Lower proportions of telehealth consultations were delivered via video for GPs over 55 years old compared with GPs under 35 years old: between −0.08 (95% CI −0.02 to −0.15) and −0.15 (95% CI −0.07 to −0.22), and for GPs in postcodes with a higher proportion of patients over 65 years old: −0.005 (95% CI −0.001 to -0.008) for each percentage point increase in the population over 65 years old.Conclusions GP characteristics are strongly associated with patterns of telehealth use in clinical work. Infrastructure support and relative pricing of different consultation modes may be useful policy instruments to encourage GPs to deliver care by the most appropriate method.

Therapeutic antibodies must have “drug-like” properties. These include high affinity and specificity for the intended target, biological activity, and additional characteristics now known as “developability properties”: long-term stability and resistance to aggregation when in solution, thermodynamic stability to prevent unfolding, high expression yields to facilitate manufacturing, low self-interaction, among others. Sequence-based liabilities may affect one or more of these characteristics. Improving the stability and developability of a lead antibody is typically achieved by modifying its sequence, a time-consuming process that often results in reduced affinity. Here we present a new antibody library format that yields high-affinity binders with drug-like developability properties directly from initial selections, reducing the need for further engineering or affinity maturation. The innovative semi-synthetic design involves grafting natural complementarity-determining regions (CDRs) from human antibodies into scaffolds based on well-behaved clinical antibodies. HCDR3s were amplified directly from B cells, while the remaining CDRs, from which all sequence liabilities had been purged, were replicated from a large next-generation sequencing dataset. By combining two in vitro display techniques, phage and yeast display, we were able to routinely recover a large number of unique, highly developable antibodies against clinically relevant targets with affinities in the subnanomolar to low nanomolar range. We anticipate that the designs and approaches presented here will accelerate the drug development process by reducing the failure rate of leads due to poor antibody affinities and developability.Abbreviations: AC-SINS: affinity-capture self-interaction nanoparticle spectroscopy; CDR: complementarity-determining region; CQA: critical quality attribute; ELISA: enzyme-linked immunoassay; FACS: fluorescence-activated cell sorting; Fv: fragment variable; GM-CSF: granulocyte-macrophage colony-stimulating factor; HCDR3: heavy chain CDR3; IFN2a: interferon α-2; IL6: interleukin-6; MACS: magnetic-activated cell sorting; NGS: next generation sequencing; PCR: polymerase chain reaction; SEC: size-exclusion chromatography; SPR: surface plasmon resonance; TGFβ-R2: transforming growth factor β-R2; VH: variable heavy; VK: variable kappa; VL: variable light; Vl: variable lambda;

As a novel alternative to established surface display or combinatorial chemistry approaches for the discovery of therapeutic peptides, we present a method for the isolation of small, cysteine-rich domains from bovine antibody ultralong complementarity-determining regions (CDRs). We show for the first time that isolated bovine antibody knob domains can function as autonomous entities by binding antigen outside the confines of the antibody scaffold. This yields antibody fragments so small as to be considered peptides, each stabilised by an intricate, bespoke arrangement of disulphide bonds. For drug discovery, cow immunisations harness the immune system to generate knob domains with affinities in the picomolar to low nanomolar range, orders of magnitude higher than unoptimized peptides from naïve library screening. Using this approach, knob domain peptides that tightly bound Complement component C5 were obtained, at scale, using conventional antibody discovery and peptide purification techniques.

Primary health care has been recognized as a critical strategy for improving population health in developing countries. This paper investigates the effect of primary care physicians on the infant mortality rate in Brazil using a dynamic panel data approach. This method accounts for the endogeneity problem and the persistence of infant mortality over time. The empirical analysis uses an eight-year panel of municipalities between 2005 and 2012. The results indicate that primary care physician supply contributed to the decline of infant mortality in Brazil. An increase of one primary care physician per 10,000 population was associated with 7.08 fewer infant deaths per 10,000 live births. This suggests that, in addition to other determinants, primary care physicians can play an important role in accounting for the reduction of infant mortality rates.

Objectives: Networks of clinical experts are being established internationally to help embed evidence based care in health systems. There is emerging evidence that these clinical networks can drive quality improvement programs, but the features that distinguish successful networks are largely unknown. We examined the factors that make clinical networks effective at improving quality of care and facilitating system-wide changes. Methods: We conducted a retrospective cross-sectional study of 19 state-wide clinical networks that reflected a range of medical and surgical specialty care and were in operation from 2006 to 2008 in New South Wales, Australia. We conducted qualitative interviews with network leaders to characterise potential impacts, and conducted internet surveys of network members to evaluate external support and the organisational and program characteristics of their respective networks. The main outcome measures were median ratings of individual network impacts on quality of care and system-wide changes, determined through independent assessment of documented evidence by an expert panel. Results: We interviewed 19 network managers and 32 network co-chairs; 592 network members completed internet surveys. Three networks were rated as having had high impact on quality of care, and seven as having had high impact on system-wide change. Better-perceived strategic and operational network management was significantly associated with higher ratings of impact on quality of care (coefficient estimate 0.86; 95% confidence interval [CI] 0.02, 1.69). Better-perceived leadership of the network manager (coefficient estimate 0.47; 95% CI 0.10, 0.85) and strategic and operational network management (coefficient estimate 0.23; 95% CI 0.06, 0.41) were associated with higher ratings of impact on system-wide change. Conclusions: This study represents the largest study of clinical networks undertaken to date. The results suggest that clinical networks that span the health system can improve quality of care and facilitate system-wide change. Network management and leadership, encompassing both strategic and operational elements at the organisational level, appear to be the primary influences on network success. These findings can guide future organisational and system-wide change programs and the development or strengthening of clinical networks to help implement evidence based care to improve service delivery and outcomes.

Every human possesses millions of distinct antibodies. It is now possible to analyze this diversity via next-generation sequencing of immunoglobulin genes (Ig-seq). This technique produces large volume sequence snapshots of B-cell receptors that are indicative of the antibody repertoire. In this paper, we enrich these large-scale sequence datasets with structural information. Enriching a sequence with its structural data allows better approximation of many vital features, such as its binding site and specificity. Here, we describe the structural annotation of antibodies pipeline that maps the outputs of large Ig-seq experiments to known antibody structures. We demonstrate the viability of our protocol on five separate Ig-seq datasets covering ca. 35 m unique amino acid sequences from ca. 600 individuals. Despite the great theoretical diversity of antibodies, we find that the majority of sequences coming from such studies can be reliably mapped to an existing structure.

Purpose: This study proposes to map the barriers to scaling the microfinance industry in the U.S., as it pertains to home maintenance and improvement for low-income households. The selected context of analysis is the American city of Baltimore, due to the city’s high need for housing repair and large percentage of residents with limited access to finance. Originality/Value: Most research has discarded microfinance as a viable option for a housing market solution in the U.S. This paper discusses how the market of microfinance for housing repair could improve its financial sustainability seizing the smaller dollar value of repair loans, relative to housing purchase, and the high and recurring need for repair. Design/methodology/approach: Qualitative research was conducted on how microfinance for housing repair works in Baltimore City, leveraging secondary government and private research, along with interviews with lenders and borrowers. Data were analyzed through PESTEL framework, describing the macro-environmental context. Findings: The market for Housing Microfinance (HM) loan products in Baltimore matches the academic literature. Similar market demands exist as they relate to an increasingly aging housing stock. Lender supply of financing seems “healthy”, but it is mostly from philanthropic or government sources favoring “affordability” over financial sustainability.

We tested the hypothesis that: basal cardiac autonomic modulation (i.e. heart rate variability, HRV) was associated with the parasympathetic withdrawal during, and overall Yo-Yo IR1 performance; and therefore HRV could be used as a simple tool for specific fitness assessment. Nine sub-elite soccer referees performed the Yo-Yo IR1 until exhaustion with simultaneous recording of heart rate with the autonomic response during exercise calculated as the area under the curve (AUC). In the same week but with at least 48 hrs between days, a night-time HR recording was obtained from participants for determination of basal HRV. Correlation analysis demonstrated strong (r>0.83) relationships among Yo-Yo IR1 performance, AUC and basal HRV. These results indicate that both nocturnal HRV and AUC can be utilised as simple and valid monitoring tools of soccer referees' fitness.Nós testamos a hipótese de: que a modulação autonômica cardíaca (i.e. variabilidade da freqüência cardíaca, VFC) está associada com a retirada parassimpática durante a execução do Yo-Yo IR1; e que, portanto, a VFC poderia ser usada como uma ferramenta simples para a avaliação do desempenho físico específico. Nove árbitros sub-elite de futebol executaram o Yo-Yo IR1 até a exaustão com registro simultâneo da freqüência cardíaca e VFC para o cálculo da área sob a curva (ASC). Na mesma semana, mas com mínimo de 48 horas entre dias, obteve-se um registro noturno da FC dos participantes para a determinação da VFC basal. As análises de correlação demonstraram elevados coeficientes (r>0,83) entre o desempenho no Yo-Yo IR1, a ASC e a VFC basal. Estes resultados indicam que a VFC noturna e a ASC podem ser empregadas como ferramentas simples e válidas para o monitoramento do desempenho de árbitros de futebol.

Condensed Abstract: The prevalence and prognostic importance of CM occurring as a consequence of AF is poorly defined. This study investigated the incidence of CM in patients with AF, its clinical features and long-term outcomes. We demonstrated that CM is common in patients presenting acutely with newly diagnosed rapid AF, and carries a worse long-term prognosis. Systolic dysfunction was reversible in an important proportion of patients, suggesting a greater prevalence of rate-related CM in AF than has previously been postulated. This underscores the importance of appropriate rhythm management strategies and repeat imaging studies. Summary Background: Atrial fibrillation (AF) may precipitate LV dysfunction, potentially leading to cardiomyopathy (CM). The prevalence and prognostic importance of CM occurring as a consequence of AF is poorly defined. We investigated the incidence of CM in patients with AF, its clinical features and long-term outcomes. Methods: We reviewed 292 consecutive patients (average age 72 ± 13yrs) presenting acutely with AF and tachycardia over a 3 year period from June 2004. Clinical details were obtained from medical records. CM was defined as ejection fraction (EF) ≤ 50% on index admission. Results: Echo was performed 93% of patients at index admission, and 69 (24%) had CM (average EF% = 37 ± 11), 60 of which were newly diagnosed. Patients with CM had significantly higher presenting heart rate (141 ± 19 vs. 132 ± 23 bpm), larger end-diastolic (5.7 vs. 5.2 cm) and end-systolic (4.5 vs. 3.2 cm) dimensions, and larger left atrial size (4.6 vs. 4.3 cm) (P < 0.05 for all). They were also statistically more likely (P < 0.05) to be male, present with breathlessness, have a history of coronary disease, and be treated with digoxin and warfarin. Follow-up echo between 6 and 12 months was performed in 46% of patients with new CM, and average EF rose to 53 ± 12%. At an average follow-up of 2.5 years, there was a significant increase in mortality in CM patients (16% vs. 9.5%, P < 0.05). Conclusion: CM is common in patients presenting acutely with newly diagnosed rapid AF, and carries a worse long-term prognosis. Systolic dysfunction was reversible in an important proportion of patients, suggesting a greater prevalence of rate-related CM in AF than has previously been postulated. This underscores the importance of appropriate rhythm management strategies and repeat imaging studies.

PURPOSE: Primary open-angle glaucoma (POAG) is a degenerative eye disease for which early treatment is critical to mitigate visual impairment and irreversible blindness. POAG-associated loci individually confer incremental risk. Genetic risk score(s) (GRS) could enable POAG risk stratification. Despite significantly higher POAG burden among individuals of African ancestry (AFR), GRS are limited in this population. A recent large-scale, multi-ancestry meta-analysis identified 127 POAG-associated loci and calculated cross-ancestry and ancestry-specific effect estimates, including in European ancestry (EUR) and AFR individuals. We assessed the utility of the 127-variant GRS for POAG risk stratification in EUR and AFR Veterans in the Million Veteran Program (MVP). We also explored the association between GRS and documented invasive glaucoma surgery (IGS). DESIGN: Cross-sectional study. PARTICIPANTS: MVP Veterans with imputed genetic data, including 5830 POAG cases (445 with IGS documented in the electronic health record) and 64 476 controls. METHODS: We tested unweighted and weighted GRS of 127 published risk variants in EUR (3382 cases and 58 811 controls) and AFR (2448 cases and 5665 controls) Veterans in the MVP. Weighted GRS were calculated using effect estimates from the most recently published report of cross-ancestry and ancestry-specific meta-analyses. We also evaluated GRS in POAG cases with documented IGS. MAIN OUTCOME MEASURES: Performance of 127-variant GRS in EUR and AFR Veterans for POAG risk stratification and association with documented IGS. RESULTS: GRS were significantly associated with POAG (P < 5 × 10-5) in both groups; a higher proportion of EUR compared with AFR were consistently categorized in the top GRS decile (21.9%-23.6% and 12.9%-14.5%, respectively). Only GRS weighted by ancestry-specific effect estimates were associated with IGS documentation in AFR cases; all GRS types were associated with IGS in EUR cases. CONCLUSIONS: Varied performance of the GRS for POAG risk stratification and documented IGS association in EUR and AFR Veterans highlights (1) the complex risk architecture of POAG, (2) the importance of diverse representation in genomics studies that inform GRS construction and evaluation, and (3) the necessity of expanding diverse POAG-related genomic data so that GRS can equitably aid in screening individuals at high risk of POAG and who may require more aggressive treatment.

Obesity is a risk factor for many human diseases. However, the underlying molecular causes of obesity are not well understood. Here, we report that protein tyrosine phosphatase receptor T (PTPRT) knockout mice are resistant to high-fat diet-induced obesity. Those mice avoid many deleterious side effects of high-fat diet-induced obesity, displaying improved peripheral insulin sensitivity, lower blood glucose and insulin levels. Compared to wild type littermates, PTPRT knockout mice show reduced food intake. Consistently, STAT3 phosphorylation is up-regulated in the hypothalamus of PTPRT knockout mice. These studies implicate PTPRT-modulated STAT3 signaling in the regulation of high-fat diet-induced obesity.

Escherichia coli can experience a multifaceted life, in some cases acting as a commensal while in other cases causing intestinal and/or extraintestinal disease. Several studies suggest enteroaggregative E. coli are the predominant cause of E. coli-mediated diarrhea in the developed world and are second only to Campylobacter sp. as a cause of bacterial-mediated diarrhea. Furthermore, enteroaggregative E. coli are a predominant cause of persistent diarrhea in the developing world where infection has been associated with malnourishment and growth retardation.In this study we determined the complete genomic sequence of E. coli 042, the prototypical member of the enteroaggregative E. coli, which has been shown to cause disease in volunteer studies. We performed genomic and phylogenetic comparisons with other E. coli strains revealing previously uncharacterised virulence factors including a variety of secreted proteins and a capsular polysaccharide biosynthetic locus. In addition, by using Biolog Phenotype Microarrays we have provided a full metabolic profiling of E. coli 042 and the non-pathogenic lab strain E. coli K-12. We have highlighted the genetic basis for many of the metabolic differences between E. coli 042 and E. coli K-12.This study provides a genetic context for the vast amount of experimental and epidemiological data published thus far and provides a template for future diagnostic and intervention strategies.

The prevalence and prognostic importance of CM occurring as a consequence of AF is poorly defined. This study investigated the incidence of CM in patients with AF, its clinical features and long-term outcomes. We demonstrated that CM is common in patients presenting acutely with newly diagnosed rapid AF, and carries a worse long-term prognosis. Systolic dysfunction was reversible in an important proportion of patients, suggesting a greater prevalence of rate-related CM in AF than has previously been postulated. This underscores the importance of appropriate rhythm management strategies and repeat imaging studies. Background Atrial fibrillation (AF) may precipitate LV dysfunction, potentially leading to cardiomyopathy (CM). The prevalence and prognostic importance of CM occurring as a consequence of AF is poorly defined. We investigated the incidence of CM in patients with AF, its clinical features and long-term outcomes. Methods We reviewed 292 consecutive patients (average age 72 ± 13yrs) presenting acutely with AF and tachycardia over a 3 year period from June 2004. Clinical details were obtained from medical records. CM was defined as ejection fraction (EF) ≤ 50% on index admission. Results Echo was performed 93% of patients at index admission, and 69 (24%) had CM (average EF% = 37 ± 11), 60 of which were newly diagnosed. Patients with CM had significantly higher presenting heart rate (141 ± 19 vs. 132 ± 23 bpm), larger end-diastolic (5.7 vs. 5.2 cm) and end-systolic (4.5 vs. 3.2 cm) dimensions, and larger left atrial size (4.6 vs. 4.3 cm) ( P < 0.05 for all). They were also statistically more likely ( P < 0.05) to be male, present with breathlessness, have a history of coronary disease, and be treated with digoxin and warfarin. Follow-up echo between 6 and 12 months was performed in 46% of patients with new CM, and average EF rose to 53 ± 12%. At an average follow-up of 2.5 years, there was a significant increase in mortality in CM patients (16% vs. 9.5%, P < 0.05). Conclusion CM is common in patients presenting acutely with newly diagnosed rapid AF, and carries a worse long-term prognosis. Systolic dysfunction was reversible in an important proportion of patients, suggesting a greater prevalence of rate-related CM in AF than has previously been postulated. This underscores the importance of appropriate rhythm management strategies and repeat imaging studies.

Objective Patients with newly diagnosed cardiomyopathy frequently undergo coronary angiography to exclude significant coronary artery disease (CAD). Contrast-enhanced cardiac magnetic resonance imaging (CE-CMR) can detect myocardial scar in-vivo, and has the potential to identify patients with cardiomyopathy secondary to prior myocardial infarction. Aim To investigate the accuracy of contrast-enhanced cardiac magnetic resonance imaging (CE-CMR) in differentiating ischemic from non-ischemic cardiomyopathy in patients with new onset heart failure and previously undiagnosed left ventricular systolic dysfunction. Methods Forty eight patients (mean age 54 ± 8) were prospectively identified from symptoms and echocardiography, and underwent both CE-CMR and angiography. Patients with >70% diameter stenosis in > 1 major epicardial vessel on angi-ography were considered to have an ischemic cardiomyopathy. Myocardial scar was assessed using delayed enhancement inversion-recovery imaging after gadolinium administration. Patients with subendocardial enhancement typical of a myocardial infarction were classified as having an ischemic etiology by CE-CMR. Results Sixteen patients were classified as ischemic by angiography, and of these 15 had subendocardial enhancement on CE-CMR. The sensitivity and negative predictive value for CE-MRI were 94% and 97% respectively for detecting an ischemic cause. However 5 patients with subendocardial enhancement by CE-CMR had no obstructive coronary disease, suggesting misclassification by angiography, and reducing the specificity (82%) of CE-CMR. Conclusion The lack of subendocardial scar on CE-CMR identifies patients with new-onset heart failure and cardiomyopathy who may not require angiography.

Purpose: To investigate the set up and operation of a Managed Clinical Network for cardiac services and assess its impact on patient care. Methods: This single case study used process evaluation with observational before and after comparison of indicators of quality of care and costs. The study was conducted in Dumfries and Galloway, Scotland and used a three-level framework. Process evaluation of the network set-up and operation through a documentary review of minutes; guidelines and protocols; transcripts of fourteen semi-structured interviews with health service personnel including senior managers, general practitioners, nurses, cardiologists and members of the public. Outcome evaluation of the impact of the network through interrupted time series analysis of clinical data of 202 patients aged less than 76 years admitted to hospital with a confirmed myocardial infarction one-year pre and one-year post, the establishment of the network. The main outcome measures were differences between indicators of quality of care targeted by network protocols. Economic evaluation of the transaction costs of the set-up and operation of the network and the resource costs of the clinical care of the 202 myocardial infarction patients from the time of hospital admission to 6 months post discharge through interrupted time series analysis. The outcome measure was different in National Health Service resource use. Results: Despite early difficulties, the network was successful in bringing together clinicians, patients and managers to redesign services, exhibiting most features of good network management. The role of the energetic lead clinician was crucial, but the network took time to develop and ‘bed down’. Its primary “modus operand” was the development of a myocardial infarction pathway and associated protocols. Of sixteen clinical care indicators, two improved significantly following the launch of the network and nine showed improvements, which were not statistically significant. There was no difference in resource use. Discussion and conclusions: The Managed Clinical Network made a difference to ways of working, particularly in breaching traditional boundaries and involving the public, and made modest changes in patient care. However, it required a two-year “set-up” period. Managed clinical networks are complex initiatives with an increasing profile in health care policy. This study suggests that they require energetic leadership and improvements are likely to be slow and incremental.

Neurotropic strains of mouse hepatitis virus (MHV), a coronavirus, cause acute and chronic demyelinating encephalomyelitis with similarities to the human disease multiple sclerosis. Here, using a lineage-tracking system, we show that some cells, primarily oligodendrocytes (OLs) and oligodendrocyte precursor cells (OPCs), survive the acute MHV infection, are associated with regions of demyelination, and persist in the central nervous system (CNS) for at least 150 d. These surviving OLs express major histocompatibility complex (MHC) class I and other genes associated with an inflammatory response. Notably, the extent of inflammatory cell infiltration was variable, dependent on anatomic location within the CNS, and without obvious correlation with numbers of surviving cells. We detected more demyelination in regions with larger numbers of T cells and microglia/macrophages compared to those with fewer infiltrating cells. Conversely, in regions with less inflammation, these previously infected OLs more rapidly extended processes, consistent with normal myelinating function. Together, these results show that OLs are inducers as well as targets of the host immune response and demonstrate how a CNS infection, even after resolution, can induce prolonged inflammatory changes with CNS region-dependent impairment in remyelination.