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PDF file - 129K, Supplementary Table 1. Clinical characteristics of discovery cohort. Characteristics of 10 patients from experimental arm of ICON7 trial used in MS discovery experiment; Supplementary Table 2. Number of samples in each treatment arm at each time-point; Supplementary Table 3 Associations between the baseline concentrations (time-point 1) of the serum biomarkers under investigation and the clinical characteristics in the validation study in validation cohort II; Supplementary Table 4 Associations between the baseline concentrations (time-point 1) of the serum biomarkers under investigation and the clinical characteristics in the validation study in both validation cohorts (validation cohort I and validation cohort II); Supplementary Table 5. Estimates from Cox proportional hazards regression for biomarker index (signature negative index < 3 and positive index ≥3), trial arm (standard or experimental) and signature/trial arm interaction in each validation cohort; Supplementary Table 6. Estimates from Cox proportional hazards regression for individual elements of biomarker index (score=1 if marker concentration is on the side of the cut-point favouring bevacizumab in terms of PFS in the optimal model for that marker, or a score=0 otherwise), trial arm (standard or experimental) and index element/trial arm interaction; Supplementary Table 7. Estimates from Cox proportional hazards regression for all possible combinations of the elements of the biomarker index (score=1 if the sum of the combination of elements ≥optimal split, or a score=0 otherwise, where the optimal split is recalculated for each model combination), trial arm (standard or experimental) and model combination score/trial arm interaction; Supplementary Table 8. Estimates from Cox proportional hazards regression for individual elements of biomarker index treated as continuous, trial arm (standard or experimental) and index element/trial arm interaction

Purpose: The aim of this study was to identify and validate novel predictive and/or prognostic serum proteomic biomarkers in patients with epithelial ovarian cancer (EOC) treated as part of the phase III international ICON7 clinical trial.Experimental Design: ICON7 was a phase III international trial in EOC which showed a modest but statistically significant benefit in progression-free survival (PFS) with the addition of bevacizumab to standard chemotherapy. Serum samples from 10 patients who received bevacizumab (five responders and five nonresponders) were analyzed by mass spectrometry to identify candidate biomarkers. Initial validation and exploration by immunoassay was undertaken in an independent cohort of 92 patients, followed by a second independent cohort of 115 patients (taken from across both arms of the trial).Results: Three candidate biomarkers were identified: mesothelin, fms-like tyrosine kinase-4 (FLT4), and α1-acid glycoprotein (AGP). Each showed evidence of independent prognostic potential when adjusting for high-risk status in initial (P < 0.02) and combined (P < 0.01) validation cohorts. In cohort I, individual biomarkers were not predictive of bevacizumab benefit; however, when combined with CA-125, a signature was developed that was predictive of bevacizumab response and discriminated benefit attributable to bevacizumab better than clinical characteristics. The signature showed weaker evidence of predictive ability in validation cohort II, but was still strongly predictive considering all samples (P = 0.001), with an improvement in median PFS of 5.5 months in signature-positive patients in the experimental arm compared with standard arm.Conclusions: This study shows a discriminatory signature comprising mesothelin, FLT4, AGP, and CA-125 as potentially identifying those patients with EOC more likely to benefit from bevacizumab. These results require validation in further patient cohorts. Clin Cancer Res; 19(18); 5227–39. ©2013 AACR.

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PDF file - 269K, Supplementary Figure 1: Differences in AGP, mesothelin and FLT4 concentrations between responders and non responders in the discovery sample set (n=10); Supplementary Figure 2: Progression-free survival in validation cohort I; Supplementary Figure 3: Baseline concentrations of mesothelin, FLT4, AGP and CA125 in: a). validation cohort I, b). validation cohort II, and c). both validation cohorts combined.

PDF file - 963K, Supplementary Figure 4: Correlation between biomarkers investigated in validation cohort I; Supplementary Figure 5: Longitudinal patterns in AGP concentration in validation cohort I; Supplementary Figure 6. Prognostic and predictive potential of candidate biomarkers in validation cohort I; Supplementary Figure 7. Kaplan Meier estimates of the survival function showing prognostic and predictive ability of the biomarker index in validation cohort I; Supplementary Figure 8. Prognostic and predictive potential of individual elements of biomarker index in the combined validation cohort.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the virus that causes coronavirus disease 2019. At the time of writing (October 2020), the number of cases of COVID-19 had been approaching 38 million and more than 1 million deaths were attributable to it. SARS-CoV-2 appears to be highly transmissible and could rapidly spread in hospital wards.The work undertaken aimed to estimate the clinical effectiveness and cost-effectiveness of viral detection point-of-care tests for detecting SARS-CoV-2 compared with laboratory-based tests. A further objective was to assess occupancy levels in hospital areas, such as waiting bays, before allocation to an appropriate bay.The perspective was that of the UK NHS in 2020. The setting was a hypothetical hospital with an accident and emergency department.An individual patient model was constructed that simulated the spread of disease and mortality within the hospital and recorded occupancy levels. Thirty-two strategies involving different hypothetical SARS-CoV-2 tests were modelled. Recently published desirable and acceptable target product profiles for SARS-CoV-2 point-of-care tests were modelled. Incremental analyses were undertaken using both incremental cost-effectiveness ratios and net monetary benefits, and key patient outcomes, such as death and intensive care unit care, caused directly by COVID-19 were recorded.A SARS-CoV-2 point-of-care test with a desirable target product profile appears to have a relatively small number of infections, a low occupancy level within the waiting bays, and a high net monetary benefit. However, if hospital laboratory testing can produce results in 6 hours, then the benefits of point-of-care tests may be reduced. The acceptable target product profiles performed less well and had lower net monetary benefits than both a laboratory-based test with a 24-hour turnaround time and strategies using data from currently available SARS-CoV-2 point-of-care tests. The desirable and acceptable point-of-care test target product profiles had lower requirement for patients to be in waiting bays before being allocated to an appropriate bay than laboratory-based tests, which may be of high importance in some hospitals. Tests that appeared more cost-effective also had better patient outcomes.There is considerable uncertainty in the values for key parameters within the model, although calibration was undertaken in an attempt to mitigate this. The example hospital simulated will also not match those of decision-makers deciding on the clinical effectiveness and cost-effectiveness of introducing SARS-CoV-2 point-of-care tests. Given these limitations, the results should be taken as indicative rather than definitive, particularly cost-effectiveness results when the relative cost per SARS-CoV-2 point-of-care test is uncertain.Should a SARS-CoV-2 point-of-care test with a desirable target product profile become available, this appears promising, particularly when the reduction on the requirements for waiting bays before allocation to a SARS-CoV-2-infected bay, or a non-SARS-CoV-2-infected bay, is considered. The results produced should be informative to decision-makers who can identify the results most pertinent to their specific circumstances.More accurate results could be obtained when there is more certainty on the diagnostic accuracy of, and the reduction in time to test result associated with, SARS-CoV-2 point-of-care tests, and on the impact of these tests on occupancy of waiting bays and isolation bays. These parameters are currently uncertain.This report was commissioned by the National Institute for Health Research (NIHR) Evidence Synthesis programme as project number 132154. This project was funded by the NIHR Health Technology Assessment programme and will be published in full inSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the virus that causes coronavirus disease 2019 (COVID-19). SARS-CoV-2 is highly infectious, and this can cause problems in hospitals, where the virus can spread quickly. Laboratory-based tests can determine whether or not a patient has SARS-CoV-2, but these tests are not perfect and can require a considerable time to provide a result. Point-of-care tests to detect SARS-CoV-2 are being developed that may have much shorter times to a test result, although these are likely to be less accurate than laboratory-based tests. The benefit of quicker tests is that a decision to put a patient in a SARS-CoV-2-infected bay or in a non-SARS-CoV-2-infected bay can be made sooner, limiting contact between patients with SARS-CoV-2 and patients without SARS-CoV-2 and reducing the risk of infection transmission. The disadvantage of reduced accuracy is that some patients may be allocated to the wrong bay, increasing the risk of SARS-CoV-2 infection. A computer model was built to explore the impact of using SARS-CoV-2 point-of-care tests for people admitted to hospital. This model estimated the number of infections and deaths due to COVID-19, the costs of testing, and the number of people waiting to be put in an appropriate bay. Strategies were run using different values, including the time to get a test result, the accuracy of tests and whether or not staff who do not have symptoms should be tested. The results of the model indicated that point-of-care tests could be good if there was a large reduction in the time to get a test result and if accuracy was high. However, it is not certain whether or not such tests will become available. When newer SARS-CoV-2 tests are available, the model will allow an estimate of the clinical effectiveness and cost-effectiveness of the test to be made.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the virus that causes coronavirus disease 2019 (COVID-19), which at the time of writing (January 2021) was responsible for more than 2.25 million deaths worldwide and over 100,000 deaths in the UK. SARS-CoV-2 appears to be highly transmissible and could rapidly spread in residential care homes.The work undertaken aimed to estimate the clinical effectiveness and cost-effectiveness of viral detection point-of-care tests for detecting SARS-CoV-2 compared with laboratory-based tests in the setting of a hypothetical care home facility for elderly residents.The perspective was that of the NHS in 2020. The setting was a hypothetical care home facility for elderly residents. Care homes with en suite rooms and with shared facilities were modelled separately.A discrete event simulation model was constructed to model individual residents and simulate the spread of SARS-CoV-2 once it had entered the residential care facility. The numbers of COVID-19-related deaths and critical cases were recorded in addition to the number of days spent in isolation. Thirteen strategies involving different hypothetical SARS-CoV-2 tests were modelled. Recently published desirable and acceptable target product profiles for SARS-CoV-2 point-of-care tests and for hospital-based SARS-CoV-2 tests were modelled. Scenario analyses modelled early release from isolation based on receipt of a negative SARS-CoV-2 test result and the impact of vaccination. Incremental analyses were undertaken using both incremental cost-effectiveness ratios and net monetary benefits.Cost-effectiveness results depended on the proportion of residential care facilities penetrated by SARS-CoV-2. SARS-CoV-2 point-of-care tests with desirable target product profiles appear to have high net monetary benefit values. In contrast, SARS-CoV-2 point-of-care tests with acceptable target product profiles had low net monetary benefit values because of unnecessary isolations. The benefit of allowing early release from isolation depended on whether or not the facility had en suite rooms. The greater the assumed efficacy of vaccination, the lower the net monetary benefit values associated with SARS-CoV-2 point-of-care tests, when assuming that a vaccine lowers the risk of contracting SARS-CoV-2.There is considerable uncertainty in the values for key parameters within the model, although calibration was undertaken in an attempt to mitigate this. Some degree of Monte Carlo sampling error persists because of the timelines of the project. The example care home simulated will also not match those of decision-makers deciding on the clinical effectiveness and cost-effectiveness of introducing SARS-CoV-2 point-of-care tests. Given these limitations, the results should be taken as indicative rather than definitive, particularly the cost-effectiveness results when the relative cost per SARS-CoV-2 point-of-care test is uncertain.SARS-CoV-2 point-of-care tests have considerable potential for benefit for use in residential care facilities, but whether or not this materialises depends on the diagnostic accuracy and costs of forthcoming SARS-CoV-2 point-of-care tests.More accurate results would be obtained when there is more certainty on the diagnostic accuracy of and the reduction in time to test result associated with SARS-CoV-2 point-of-care tests when used in the context of residential care facilities, the proportion of care home penetrated by SARS-CoV-2 and the levels of immunity once vaccination is administered. These parameters are currently uncertain.This report was commissioned by the National Institute for Health Research (NIHR) Evidence Synthesis programme as project number 132154. This project was funded by the NIHR Health Technology Assessment programme and will be published in full inSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the virus that causes coronavirus disease 2019 (COVID-19). SARS-CoV-2 is highly infectious, and this can cause problems in care homes, where the virus can spread quickly. Laboratory-based tests can determine whether or not someone has SARS-CoV-2, but these tests are not perfect and can take a long time to provide a result. Point-of-care tests that can be performed quickly in the care home to detect SARS-CoV-2 are being developed and they may have much shorter times to get a result than laboratory-based tests, although with worse accuracy. The benefit of quicker tests is that decisions to put residents into or release them from isolation can be made sooner, reducing the risk of spreading SARS-CoV-2 and reducing time in isolation. The disadvantage of reduced accuracy is that wrong decisions could be made, resulting in either unnecessary isolation or increased spread of SARS-CoV-2. A computer model was built to explore the impact of using SARS-CoV-2 point-of-care tests for residents of care homes. The model estimated the number of SARS-CoV-2 infections, deaths due to COVID-19 and days in isolation. Strategies were run using different values, including the time to get a test result back, the accuracy of tests, the proportion of care homes where there is a case of SARS-CoV-2, whether residents were isolated individually or in groups and how well vaccines work. The results of the model indicated that point-of-care tests could be good if there was a large decrease in the time to get a test result back, if accuracy was high and if vaccination protection was moderate. However, the accuracy and speed of future point-of-care tests is uncertain. When newer SARS-CoV-2 tests are available, the model will allow an estimate of the clinical effectiveness and cost-effectiveness of the tests to be made.

Despite reported increases in referral numbers, a large proportion of those with prediabetes still decline participation in the NHS England Diabetes Prevention Programme (NDPP). The aim of this study was to explore whether investment in interventions to improve uptake of the programme has the potential to be cost-effective. An early cost-utility analysis was conducted using a Markov model parameterized based on secondary data sources. We explored different uptake scenarios and the impact that this had on the maximum allowable intervention price based on cost-effectiveness at the UK NICE willingness to pay threshold of £20,000 (US$ 25,913). Value of information analyses were conducted to explore the potential value of further research to resolve uncertainty at each uptake level. As uptake levels increase, the maximum allowable intervention price and overall expected value of removing decision uncertainty increases. For 5 percentage and 50 percentage points increase in uptake levels, the maximum allowable intervention price is £41.86 (US$ 54.23) and £418.59 (US$ 542.34) per person, and the overall expected value of removing decision uncertainty are £361,818,839 (US$ 468,786,625) and £1,468,712,316 (US$ 1,902,921,063) respectively. There is headroom for investment in interventions that improve uptake to the NDPP, thereby allowing the programme itself to be delivered in a manner that remains cost-effective.

Target Product Profiles (TPPs) outline the characteristics that new health technologies require to address an unmet clinical need. To date, published TPPs for medical tests have focused on infectious diseases, mostly in the context of low- and middle-income countries. Recently, there have been calls for a broader use of TPPs as a mechanism to ensure that diagnostic innovation is aligned with clinical needs, yet the methodology underpinning TPP development remains suboptimal. Here, we propose that early economic evaluation (EEE) should be integrated within the TPP methodology to create a more rigorous framework for the development of “fit-for-purpose” tests. We discuss the potential benefits that EEE could bring to the core activities underpinning TPP development—scoping, drafting, consensus building, and updating—and argue that using EEE to help inform TPPs provides a more objective, evidence-based, and transparent approach to defining test specifications.

Background A Target Product Profile (TPP) outlines the necessary characteristics of an innovative product to address an unmet clinical need. TPPs could be used to better guide manufacturers in the development of ‘fit for purpose’ tests, thus increasing the likelihood that novel tests will progress from bench to bedside. However, there is currently no guidance on how to produce a TPP specifically for medical tests. Methods A systematic review was conducted to summarise the methods currently used to develop TPPs for medical tests, the sources used to inform these recommendations and the test characteristics for which targets are made. Database and website searches were conducted in November 2018. TPPs written in English for any medical test were included. Based on an existing framework, test characteristics were clustered into commonly recognised themes. Results Forty-four TPPs were identified, all of which focused on diagnostic tests for infectious diseases. Three core decision-making phases for developing TPPs were identified: scoping, drafting and consensus-building. Consultations with experts and the literature mostly informed the scoping and drafting of TPPs. All TPPs provided information on unmet clinical need and desirable analytical performance, and the majority specified clinical validity characteristics. Few TPPs described specifications for clinical utility, and none included cost-effectiveness. Conclusions We have identified a commonly used framework that could be beneficial for anyone interested in drafting a TPP for a medical test. Currently, key outcomes such as utility and cost-effectiveness are largely overlooked within TPPs though and we foresee this as an area for further improvement.

Background: The presence of circulating cell-free DNA from tumours in blood (ctDNA) is of major importance to\ud those interested in early cancer detection, as well as to those wishing to monitor tumour progression or diagnose\ud the presence of activating mutations to guide treatment. In 2014, the UK Early Cancer Detection Consortium\ud undertook a systematic mapping review of the literature to identify blood-based biomarkers with potential for the\ud development of a non-invasive blood test for cancer screening, and which identified this as a major area of interest.\ud This review builds on the mapping review to expand the ctDNA dataset to examine the best options for the\ud detection of multiple cancer types.\ud Methods: The original mapping review was based on comprehensive searches of the electronic databases Medline,\ud Embase, CINAHL, the Cochrane library, and Biosis to obtain relevant literature on blood-based biomarkers for cancer\ud detection in humans (PROSPERO no. CRD42014010827). The abstracts for each paper were reviewed to determine\ud whether validation data were reported, and then examined in full. Publications concentrating on monitoring of\ud disease burden or mutations were excluded.\ud Results: The search identified 94 ctDNA studies meeting the criteria for review. All but 5 studies examined one\ud cancer type, with breast, colorectal and lung cancers representing 60% of studies. The size and design of the\ud studies varied widely. Controls were included in 77% of publications. The largest study included 640 patients, but\ud the median study size was 65 cases and 35 controls, and the bulk of studies (71%) included less than 100 patients.\ud Studies either estimated cfDNA levels non-specifically or tested for cancer-specific mutations or methylation\ud changes (the majority using PCR-based methods).\ud Conclusion: We have systematically reviewed ctDNA blood biomarkers for the early detection of cancer. Preanalytical,\ud analytical, and post-analytical considerations were identified which need to be addressed before such\ud biomarkers enter clinical practice. The value of small studies with no comparison between methods, or even the\ud inclusion of controls is highly questionable, and larger validation studies will be required before such methods can\ud be considered for early cancer detection.\ud

Objective To examine changes in outcome by the Leibovich score using contemporary and historic cohorts of patients presenting with renal cell carcinoma (RCC) Patients and Methods Prospective observational multicenter cohort study, recruiting patients with suspected newly diagnosed RCC. A historical cohort of patients was examined for comparison. Metastasis-free survival (MFS) formed the primary outcome measure. Model discrimination and calibration were evaluated using Cox proportional hazard regression and the Kaplan-Meier method. Overall performance of the Leibovich model was assessed by estimating explained variation. Results Seven hundred and six patients were recruited between 2011 and 2014 and RCC confirmed in 608 (86%) patients. Application of the Leibovich score to patients with localized clear cell RCC in this contemporary cohort demonstrated good model discrimination (c-index = 0.77) but suboptimal calibration, with improved MFS for intermediate- and high-risk patients (5-year MFS 85% and 50%, respectively) compared to the original Leibovich cohort (74% and 31%) and a historic (1998-2006) UK cohort (76% and 37%). The proportion of variation in outcome explained by the model is low and has declined over time (28% historic vs 22% contemporary UK cohort). Conclusion Prognostic models are widely employed in patients with localized RCC to guide surveillance intensity and clinical trial selection. However, the majority of the variation in outcome remains unexplained by the Leibovich model and, over time, MFS rates among intermediate- and high-risk classified patients have altered. These findings are likely to have implications for all such models used in this setting.

Objectives: To validate enhanced liver fibrosis (ELF) test and its components-amino-terminal propeptide of procollagen type III (PIIINP), tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) and HA-as biomarkers of fibrosis in SSc in an independent, international, multicentre cohort.Methods: Two hundred and fifty-four SSc patients from six Rheumatology Centres were included. Sera were collected and stored according to EUSTAR biobanking recommendations and analysed through automated high throughput diagnostics. Statistical analysis was performed with SPSS software.Results: Two hundred and forty-seven SSc patients (mean age 55.7 ± 13.9 years, 202 F) were analysed. ELF score, TIMP-1 and PIIINP levels were higher in males (P = 0.0197, P = 0.0107, P = 0.0108 respectively) and in dcSSc (P = 0.001, P = 0.0008, P < 0.0001 respectively). ELF score and the single markers significantly correlated with modified Rodnan skin score (r = 0.37, P < 0.0001), disease activity and severity (P < 0.0001 for all markers, except for HA P = 0.0001) and inversely with forced vital capacity, (FVC) % (TIMP-1, r = -0.21, P = 0.0012; PIIINP, r = -0.26, P = 0.0001), TLC% (ELF score, r = -0.20, P = 0.0036; TIMP-1, r = -0.32, P < 0.0001; PIIINP, r = -0.28, P < 0.0001), diffusion capacity of the lung for carbon monoxide (DLCO) % (P < 0.0001 for all markers, except for HA P = 0.0115). Multivariate analysis indicated that age (P < 0.001), modified Rodnan skin score (P < 0.001) and DLCO% (P = 0.005) were independently associated with ELF score.Conclusion: Between the first and this validation studies, the value of the ELF score as independent marker of skin and lung involvement in SSc is confirmed in 457 patients. A longitudinal study is on-going to identify an SSc specific algorithm with predictive value for skin and lung progression.

Introduction Although the risks associated with iodinated contrast administration are acknowledged to be very low, screening of kidney function prior to administration is still standard practice in many hospitals. This study has evaluated the feasibility of implementing a screening form in conjunction with point of care (PoC) creatinine testing as a method to manage the risks of post contrast acute kidney injury (PC-AKI) within the CT imaging pathway. Method Over an eight-week period 300 adult outpatients attending a UK CT department for contrast-enhanced scans were approached. Participants completed a screening questionnaire for co-morbidities linked to kidney dysfunction and consented to have a PoC and laboratory creatinine tests. Comparison was made against with previous baseline blood tests obtained within the preceding 3 months, as required by the study site. Participants were also invited to attend for follow up PoC and laboratory bloods tests at 48–72 h. Results 14 patients (4.7%) had a scan-day eGFR below 45mL/min/1.73m2, all identified through screening. The majority of patients (n=281/300; 93.7%) fell in the same risk category based on previous and scan-day blood results. Six PoC test failures were recorded on the scan day. The constant error between the Abbott i-STAT PoC scan-day measurements and the laboratory scan-day measurements was -3.71 (95% CI: -6.41 to -0.50). Five patients had an elevated creatinine (≥25% from baseline) post contrast administration, but no instances of PC-AKI (≥50% from baseline) were identified. Conclusion PoC creatinine testing is a practical method of ensuring renal function and is feasible in the radiology environment.

Background:Acute kidney injury (AKI) can occur rarely in patients exposed to iodinated contrast and result in contrast-induced AKI (CI-AKI). A key risk factor is the presence of preexisting chronic kidney disease (CKD); therefore, it is important to assess patient risk and obtain kidney function measurement prior to administration. Point-of-care (PoC) testing provides an alternative strategy but there remains uncertainty, with respect to diagnostic accuracy and clinical utility.Methods:A device study compared three PoC analysers (Nova StatSensor, Abbott i-STAT and Radiometer ABL800 FLEX) with a reference laboratory standard (Roche Cobas 8000 series, enzymatic creatinine). Three hundred adult patients attending a UK hospital phlebotomy department were recruited to have additional blood samples for analysis on the PoC devices.Results:The ABL800 FLEX had the strongest concordance with laboratory measured serum creatinine (mean bias=−0.86, 95% limits of agreement=−9.6 to 7.9) followed by the i-STAT (average bias=3.88, 95% limits of agreement=−8.8 to 16.6) and StatSensor (average bias=3.56, 95% limits of agreement=−27.7 to 34.8). In risk classification, the ABL800 FLEX and i-STAT identified all patients with an eGFR≤30, whereas the StatSensor resulted in a small number of missed high-risk cases (n=4/13) and also operated outside of the established performance goals.Conclusions:The screening of patients at risk of CI-AKI may be feasible with PoC technology. However, in this study, it was identified that the analyser concordance with the laboratory reference varies. It is proposed that further research exploring PoC implementation in imaging department pathways is needed.

BackgroundProtein biomarkers with associations with the activity and outcomes of diseases are being identified by modern proteomic technologies. They may be simple, accessible, cheap and safe tests that can inform diagnosis, prognosis, treatment selection, monitoring of disease activity and therapy and may substitute for complex, invasive and expensive tests. However, their potential is not yet being realised.Design and methodsThe study consisted of three workstreams to create a framework for research: workstream 1, methodology – to define current practice and explore methodology innovations for biomarkers for monitoring disease; workstream 2, clinical translation – to create a framework of research practice, high-quality samples and related clinical data to evaluate the validity and clinical utility of protein biomarkers; and workstream 3, the ELF to Uncover Cirrhosis as an Indication for Diagnosis and Action for Treatable Event (ELUCIDATE) randomised controlled trial (RCT) – an exemplar RCT of an established test, the ADVIA Centaur® Enhanced Liver Fibrosis (ELF) test (Siemens Healthcare Diagnostics Ltd, Camberley, UK) [consisting of a panel of three markers – (1) serum hyaluronic acid, (2) amino-terminal propeptide of type III procollagen and (3) tissue inhibitor of metalloproteinase 1], for liver cirrhosis to determine its impact on diagnostic timing and the management of cirrhosis and the process of care and improving outcomes.ResultsThe methodology workstream evaluated the quality of recommendations for using prostate-specific antigen to monitor patients, systematically reviewed RCTs of monitoring strategies and reviewed the monitoring biomarker literature and how monitoring can have an impact on outcomes. Simulation studies were conducted to evaluate monitoring and improve the merits of health care. The monitoring biomarker literature is modest and robust conclusions are infrequent. We recommend improvements in research practice. Patients strongly endorsed the need for robust and conclusive research in this area. The clinical translation workstream focused on analytical and clinical validity. Cohorts were established for renal cell carcinoma (RCC) and renal transplantation (RT), with samples and patient data from multiple centres, as a rapid-access resource to evaluate the validity of biomarkers. Candidate biomarkers for RCC and RT were identified from the literature and their quality was evaluated and selected biomarkers were prioritised. The duration of follow-up was a limitation but biomarkers were identified that may be taken forward for clinical utility. In the third workstream, the ELUCIDATE trial registered 1303 patients and randomised 878 patients out of a target of 1000. The trial started late and recruited slowly initially but ultimately recruited with good statistical power to answer the key questions. ELF monitoring altered the patient process of care and may show benefits from the early introduction of interventions with further follow-up. The ELUCIDATE trial was an ‘exemplar’ trial that has demonstrated the challenges of evaluating biomarker strategies in ‘end-to-end’ RCTs and will inform future study designs.ConclusionsThe limitations in the programme were principally that, during the collection and curation of the cohorts of patients with RCC and RT, the pace of discovery of new biomarkers in commercial and non-commercial research was slower than anticipated and so conclusive evaluations using the cohorts are few; however, access to the cohorts will be sustained for future new biomarkers. The ELUCIDATE trial was slow to start and recruit to, with a late surge of recruitment, and so final conclusions about the impact of the ELF test on long-term outcomes await further follow-up. The findings from the three workstreams were used to synthesise a strategy and framework for future biomarker evaluations incorporating innovations in study design, health economics and health informatics.Trial registrationCurrent Controlled Trials ISRCTN74815110, UKCRN ID 9954 and UKCRN ID 11930.FundingThis project was funded by the NIHR Programme Grants for Applied Research programme and will be published in full inProgramme Grants for Applied Research; Vol. 6, No. 3. See the NIHR Journals Library website for further project information.

Introduction Numerous factors contribute to uncertainty in test measurement procedures, and this uncertainty can have a significant impact on the downstream clinical utility and cost-effectiveness of testing strategies. Currently, however, there is no clear guidance concerning if or how such factors should be considered within Health Technology Assessments (HTAs) of tests. Objective The aim was to provide an introduction to key concepts in measurement uncertainty for the HTA community and to explore, via systematic review, current methods utilised within HTAs. Methods HTAs of in vitro tests including a model-based economic evaluation were identified via the Centre for Reviews and Dissemination (CRD) HTA database and key reimbursement authority websites. Data were extracted to explore the specific components of measurement uncertainty assessed and methods utilised. The findings were narratively synthesised. Results Of 107 identified HTAs, 20 (19%) attempted to assess components of measurement uncertainty: 15 did so via some form of pre-model assessment (such as a literature review or laboratory survey); four also included components within the economic model; and one considered measurement uncertainty within the model only. One study quantified the impact of measurement uncertainty on cost-effectiveness and found that this parameter significantly changed the results, but did not impact the overall decision uncertainty. Conclusion A minority of HTAs identified from this review used various approaches to assess and/or incorporate the impact of measurement uncertainty, indicating that these assessments are feasible. Uncertainty remains around best practice methodology for conducting such analyses; further research is required to ensure that future HTAs are fit for purpose. Electronic supplementary material The online version of this article (10.1007/s40273-018-0638-1) contains supplementary material, which is available to authorized users.

Background The Enhanced Liver Fibrosis (ELF) test is a serum test including the serum concentrations of amino-terminal pro-peptide of procollagen type III (PIIINP), tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) and hyaluronic acid (HA). A recent single centre study showed that ELF score and its components are markers of overall fibrosis in systemic sclerosis (SSc) mainly reflecting skin and lung involvement (1). Objectives To determine the value of ELF score and its single analytes in an independent multicentre cohort of SSc patients. Methods 254 SSc patients from 6 European Rheumatology Centres were included in this study. Clinical data were collected at time of sampling. Serum samples were collected and stored according to EUSTAR biobanking recommendations (2). Sera were analysed employing a high-throughput in vitro diagnostic (Siemens Alpha-Centaur). Statistical analysis was performed with SPSS software V.24. Results The 254 SSc patients had a mean age 55.8±13.8 years, and included 209 females and 80 patients with diffuse cutaneous SSc (dcSSc). ELF score was overall higher in males than in females (p=0.0236) as well as in dcSSc compared to limited cutaneous SSc patients (p=0.0015). ELF score and the single markers significantly correlated with the degree of skin involvement (mRSS) and inversely correlated with FVC%, TLC% and DLCO%. Concordantly all markers significantly correlated with skin and lung severity as assessed by the Medsger9s scale (Table 1). TIMP-1 and PIIINP levels were higher in patients with lung fibrosis assessed by chest HRCT scan (p=0.0126 and p=0.0308 respectively). Significant correlation (p Conclusions The value of ELF score as independent marker of skin and lung involvement is confirmed in a second independent multicentre cohort of SSc patients. A longitudinal study paired with analysis of large cohort of healthy controls is currently on going to identify a SSc specific test with the highest predictive value for skin and lung progression independently of age and gender. References Abignano G et al. Ann Rheum Dis. 2014. Beyer C et al. Ann Rheum Dis. 2011. Disclosure of Interest None declared

Early identification and prognostic stratification of delayed graft function following renal transplantation has significant potential to improve outcome. Mass spectrometry analysis of serum samples, before and on day 2 post transplant from five patients with delayed graft function and five with an uncomplicated transplant, identified aminoacylase-1 (ACY-1) as a potential outcome biomarker. Following assay development, analysis of longitudinal samples from an initial validation cohort of 55 patients confirmed that the ACY-1 level on day 1 or 2 was a moderate predictor of delayed graft function, similar to serum creatinine, complementing the strongest predictor cystatin C. A further validation cohort of 194 patients confirmed this association with area under ROC curves (95% CI) for day 1 serum (138 patients) of 0.74 (0.67–0.85) for ACY-1, 0.9 (0.84–0.95) for cystatin C, and 0.93 (0.88–0.97) for both combined. Significant differences in serum ACY-1 levels were apparent between delayed, slow, and immediate graft function. Analysis of long-term follow-up for 54 patients with delayed graft function showed a highly significant association between day 1 or 3 serum ACY-1 and dialysis-free survival, mainly associated with the donor–brain–dead transplant type. Thus, proteomic analysis provides novel insights into the potential clinical utility of serum ACY-1 levels immediately post transplantation, enabling subdivision of patients with delayed graft function in terms of long-term outcome. Our study requires independent confirmation.

Background: Objectively measured circulating biomarkers of prognosis complementing existing clinicopathological models are needed in renal cell carcinoma (RCC). Methods: Blood samples collected from 216 RCC patients in Leeds before nephrectomy (median follow-up 7 years) were analysed for C-reactive protein (CRP), osteopontin (OPN) and carbonic anhydrase IX (CA9) and prognostic significance determined. Results: CA9, OPN and CRP were univariately prognostic for overall survival (OS), cancer-specific survival (CSS) and disease-free survival (DFS) with CRP and CA9 being independently prognostic for OS/CSS and OS, respectively. Including CA9, OPN and CRP with other conventional prognostic factors gave a superior predictive capacity when compared with a previously published pre-operative clinical nomogram (Karakiewicz et al, 2009). Osteopontin outperformed this nomogram and the post-operative SSIGN score for OS but not for CSS, being significantly predictive for non-cancer deaths. Osteopontin, CRP and CA9 outperformed stage (c-index 76% compared with 70% for stage) and OPN or CA9 identified several subsets of poor prognosis patients including in T1 patients, who may benefit from adjuvant therapy and increased surveillance. Conclusion: Circulating CA9, OPN and CRP add value to existing clinicopathological prognostic factors/models and support further studies to investigate their potential use in improving the clinical management of RCC.

There has been an immense amount of research activity in the emerging field of regenerative medicine that dates back to the early 80's and beyond and many examples where products have made the difficult transition from the laboratory to commercially available goods. This process, like many product developments, is fraught with issues that range from identifying and realising the market needs to the challenge of raising capital. It can be argued that this transition process is even more challenging for regenerative medicine products since, in many cases the existing infrastructural network i.e. regulations and standards that support medicines and medical devices are inappropriate. Indeed this non-compliance with the existing framework is often cited as a barrier to commercialisation. Here, we review the current regulatory framework and standards portfolio and discuss the role that they have in commercialising potential regenerative medicine products. It does not cover the broader issue of regenerative medicine research itself or the economics of commercialisation.

The following in vitro translational study investigated whether enamel matrix derivative (EMD), an approved biomimetic treatment for periodontal disease (Emdogain®) and hard-to-heal wounds (Xelma®), enhanced synovial cell colonization and protein synthesis around a scaffold used clinically for in situ tissue engineering of the torn anterior cruciate ligament (ACL). Synovial cells were enzymatically extracted from bovine synovium and dynamically seeded onto polyethylene terephthalate (PET) scaffolds. The cells were cultured in low-serum medium (0.5% FBS) for 4 weeks with either a single administration of EMD at the start of the 4 week period or multiple administrations of EMD at regular intervals throughout the 4 weeks. Samples were harvested and evaluated using the Hoechst DNA assay, BCA protein assay, cresolphthalein complexone calcium assay, SDS–PAGE, ELISA and electron microscopy. A significant increase in cell number (DNA) (p < 0.01), protein content (p < 0.01) and TGFβ1 synthesis (p < 0.01) was observed with multiple administrations of EMD. Additionally, SDS–PAGE showed an increase in high molecular weight proteins, characteristic of the fibril-forming collagens. Electron microscopy supported these findings, showing that scaffolds treated with multiple administrations of EMD were heavily coated with cells and extracellular matrix (ECM) that enveloped the fibres. Multiple administrations of EMD to synovial cell-seeded scaffolds enhanced the formation of tissue in vitro. Additionally, it was shown that EMD enhanced TGFβ1 synthesis of synovial cells, suggesting a potential mode of action for EMD's capacity to stimulate tissue regeneration. Copyright © 2009 John Wiley & Sons, Ltd.

Polyester scaffolds have been used as an alternative to autogenous tissues for the reconstruction of the anterior cruciate ligament (ACL). They are biocompatible and encourage tissue infiltration, leading to neoligament formation. However, rupture can occur, caused by abrasion of the scaffold against the bone tunnels through which it is implanted. Good early tissue induction is therefore considered essential to protect the scaffold from this abrasion. Enamel matrix derivative (EMD) is used clinically in the treatment of periodontal disease. It is a complex mix of proteins with growth factor-like activity, which enhances periodontal ligament fibroblast attachment, proliferation, and differentiation, leading to the regeneration of periodontal bone and ligament tissues. We hypothesized that EMD might, in a similar manner, enhance tissue induction around scaffolds used in ACL reconstruction. This preliminary investigation adopted a translational approach, modelling in vitro 3 possible clinical modes of EMD administration, to ascertain the suitability of each protocol for application in an animal model or clinically. Preliminary investigations in monolayer culture indicated that EMD had a significant dose-dependent stimulatory effect (p < 0.05, n = 6) on the proliferation of bovine primary synovial cells. However, pre-treating culture plates with EMD significantly inhibited cell attachment (p < 0.01, n = 6). EMD's effects on synovial cells, seeded onto ligament scaffolds, were then investigated in several in vitro experiments modelling 3 possible modes for clinical EMD administration (pre-, intra-, and post-operative). In the pre-operative model, EMD was adsorbed onto scaffolds before the addition of cells. In the intra-operative model, EMD and cells were added simultaneously to scaffolds in the culture medium. In the post-operative model, cells were pre-seeded onto scaffolds before EMD was administered. EMD significantly inhibited cell adhesion in the pre-operative model (p < 0.05, n = 6) and had no significant benefit in the intra-operative model. In the post-operative model, the addition of EMD to previously cell-seeded scaffolds significantly increased their total deoxyribonucleic acid content (p < 0.01, n = 5). EMD's stimulative effect on cell proliferation in vitro suggests that it may accelerate scaffold colonization by cells (and in turn tissue induction) in situ. However, its inhibitory effect on synovial cell attachment in vitro implies that it may only be suited to post-operative administration.

Lack of reproducibility is an ongoing problem in some areas of the biomedical sciences. Poor experimental design and a failure to engage with experienced statisticians at key stages in the design and analysis of experiments are two factors that contribute to this problem. The RIPOSTE (Reducing IrreProducibility in labOratory STudiEs) framework has been developed to support early and regular discussions between scientists and statisticians in order to improve the design, conduct and analysis of laboratory studies and, therefore, to reduce irreproducibility. This framework is intended for use during the early stages of a research project, when specific questions or hypotheses are proposed. The essential points within the framework are explained and illustrated using three examples (a medical equipment test, a macrophage study and a gene expression study). Sound study design minimises the possibility of bias being introduced into experiments and leads to higher quality research with more reproducible results. DOI: http://dx.doi.org/10.7554/eLife.05519.001

Although successful and persistent colonization of the gastric mucosa depends on the ability to respond to changing environmental conditions and co-ordinate the expression of virulence factors during the course of infection, Helicobacter pylori possesses relatively few transcriptional regulators. We therefore investigated the contribution of the regulatory protein CsrA to global gene regulation in this important human pathogen. CsrA was necessary for full motility and survival of H. pylori under conditions of oxidative stress. Loss of csrA expression deregulated the oxidant-induced transcriptional responses of napA and ahpC, the acid induction of napA, cagA, vacA, the urease operon, and fur, as well as the heat shock responses of napA, groESL and hspR. Although the level of napA transcript was higher in the csrA mutant, its stability was similar in the wild-type and mutant strains, and less NapA protein was produced in the mutant strain. Finally, H. pylori strains deficient in the production of CsrA were significantly attenuated for virulence in a mouse model of infection. This work provides evidence that CsrA has a broad role in regulating the physiology of H. pylori in response to environmental stimuli, and may be important in facilitating adaptation to the different environments encountered during colonization of the gastric mucosa. Furthermore, CsrA appears to mediate its effects in H. pylori at the post-transcriptional level by influencing the processing and translation of target transcripts, with minimal effect on the stability of the target mRNAs.

Purpose: The aim of this study was to identify and validate novel predictive and/or prognostic serum proteomic biomarkers in patients with epithelial ovarian cancer (EOC) treated as part of the phase III international ICON7 clinical trial. Experimental Design: ICON7 was a phase III international trial in EOC which showed a modest but statistically significant benefit in progression-free survival (PFS) with the addition of bevacizumab to standard chemotherapy. Serum samples from 10 patients who received bevacizumab (five responders and five nonresponders) were analyzed by mass spectrometry to identify candidate biomarkers. Initial validation and exploration by immunoassay was undertaken in an independent cohort of 92 patients, followed by a second independent cohort of 115 patients (taken from across both arms of the trial). Results: Three candidate biomarkers were identified: mesothelin, fms-like tyrosine kinase-4 (FLT4), and α1-acid glycoprotein (AGP). Each showed evidence of independent prognostic potential when adjusting for high-risk status in initial (P < 0.02) and combined (P < 0.01) validation cohorts. In cohort I, individual biomarkers were not predictive of bevacizumab benefit; however, when combined with CA-125, a signature was developed that was predictive of bevacizumab response and discriminated benefit attributable to bevacizumab better than clinical characteristics. The signature showed weaker evidence of predictive ability in validation cohort II, but was still strongly predictive considering all samples (P = 0.001), with an improvement in median PFS of 5.5 months in signature-positive patients in the experimental arm compared with standard arm. Conclusions: This study shows a discriminatory signature comprising mesothelin, FLT4, AGP, and CA-125 as potentially identifying those patients with EOC more likely to benefit from bevacizumab. These results require validation in further patient cohorts. Clin Cancer Res; 19(18); 5227–39. ©2013 AACR.

Background Neutrophil gelatinase-associated lipocalin (NGAL) is a promising biomarker for acute kidney injury that is beginning to be used in clinical practice in addition to research studies. The current study describes an independent validation and comparison of five commercially available NGAL assays, focusing on urine samples. This is an essential step in the translation of this marker to clinical use in terms of allowing valid inter-study comparison and generation of robust results. Methods Two CE (Conformité Européenne)-marked assays, the NGAL Test (BioPorto) on Siemens ADVIA® 1800 and the ARCHITECT Urine NGAL assay on i2000SR (Abbott Laboratories), and three research-use-only (RUO) ELISAs (R&D Systems, Hycult and BioPorto) were evaluated. Imprecision, parallelism, recovery, selectivity, limit of quantitation (LOQ), vulnerability to interference and hook effect were assessed and inter-assay agreement was determined using 68 urine samples from patients with various renal diseases and healthy controls. Results The Abbott and R&D Systems assays demonstrated satisfactory performance for all parameters tested. However for the other three assays evaluated, problems were identified with LOQ (BioPorto/ADVIA®), parallelism (BioPorto ELISA) or several parameters (Hycult). Between-method agreement varied with the Hycult assay in particular being markedly different and highlighting issues with standardization and form of NGAL measured. Conclusions Variability exists between the five NGAL assays in terms of their performance and this should be taken into account when interpreting results from the various clinical or research studies measuring urinary NGAL.

Background VEDOSS project relied on a specific set of inclusion criteria identifying patients in very early stage of systemic sclerosis (SSc) to be submitted to a tight follow up and unravel factors linked to the progression of the disease. The components of the ELF score (PIIINP, TIMP-1 and HA) have been shown to correlate with severity of skin and lung fibrosis in SSc. Objectives Our aim was to analyze the concentrations of ELF components in a subset of patients enrolled in the VEDOSS study and determine their potential diagnostic value. Methods Sera from 114 patients enrolled into the VEDOSS data base and 67 SSc controls fulfilling ACR/EULAR 2013 criteria (subsets: 33 diffuse; 34 limited) were obtained from 2 centres. Serum concentrations of ELF components were determined on Siemens Advia Centaur platform. Logistic regression analysis of the results was performed employing classification of SSc as state variable. Results Among the 114 patients enrolled from VEDOSS, 30 subjects had primary Raynaud9s phenomenon (PRP); 54 had Raynaud9s phenomenon (RP) and at least one of the following “red flags” (risk factors) for SSc: (Puffy fingers, ANA, SSc specific autoantibodies or SSc-specific capillaroscopic pattern) without fulfilling 2013 ACR/EULAR criteria (score 9) despite having no evidence of skin involvement nor interstitial lung disease at lung high resolution CT or pulmonary arterial hypertension, left ventricular systolic impairment or renal involvement. All serum biomarkers concentrations correlated with age (p Conclusions Our data indicate that the SSc score is a simple serum test based model that can be used in patients with RP to aid in the very early diagnosis of SSc. Furthermore, the identification of VEDOSS patients with a high SSc score test and already classified as SSc even in the absence of internal organ involvement can be used in intervention trials aimed to prevent further disease progression. Disclosure of Interest None declared

Background The ELF test and its components (PIIINP, TIMP-1 and HA) have been shown to correlate with skin, lung and overall fibrosis and not with any vascular manifestation of Systemic Sclerosis (SSc) 1 . On the contrary, NT-proBNP has been suggested to be useful for stratification of SSc patients, especially to identify those at risk of pulmonary hypertension 2 . Objectives Aims of this study were: a) to validate ELF score and its single analytes on an independent cohort of scleroderma patients; b) to evaluate whether NT-proBNP could provide additional value to the development of an SSc-specific algorithm. Methods 250 sera from SSc patients from a single UK centre were analysed employing a high-throughput in vitro diagnostic of a routine NHS pathology lab to measure ELF score, its analytes and NT-proBNP levels. All patients fulfilled 2013 ACR/EULAR classification criteria for SSc. Clinical, laboratory and instrumental data were collected at time of sampling. Statistical analysis was performed using SPSS. P Results Multivariate analysis of ELF score (including the variables found statistically significant in univariate analysis) identified age, mRSS and DLCO% as independently associated with ELF score (p Conclusions Our findings validate the value of ELF score in a second independent cohort of 250 SSc sera and suggest that NT-proBNP has a complementary value correlating with other aspects of the disease such as PAH and heart severity. Longitudinal multicentres studies are warranted to determine the sensitivity to change and the predictive value of these biomarkers in SSc patients and to build up a new combined scleroderma specific algorithm including markers of fibrosis and vasculopathy. References Abignano G et al. Ann Rheum Dis 2014 Avouac J et al. Arthritis Care Res 2015 Disclosure of Interest None declared

The recurrence of a tumour at the resection margins in head and neck squamous cell carcinoma (HNSCC) has profound implications on the morbidity and mortality of the patient. At present HNSCC does not undergo any form of molecular analysis to aid treatment strategy and prognosticate for those individuals at higher risk of recurrence. This article aims to review current research into molecular strategies for tumour evaluation, highlighting conflicting evidence and possible novel concepts for further exploration.

Background There is increasing interest in measuring the soluble forms of carbonic anhydrase IX (CA IX) in blood as a marker of hypoxia for prognostic purposes or for predictive use in therapeutic trials in various cancers. Following our initial observations of marked differences in the measured concentrations of CA IX in EDTA plasma versus serum, we sought to investigate these further in order to determine their effects on results in published studies and to ensure accurate measurement in future studies. Methods Serum and EDTA plasma samples from healthy controls and patients with renal cancer were used in the validation of two commercially available enzyme-linked immunosorbent assays (ELISAs) for CA IX with examination of recovery, parallelism and specificity and comparison of paired plasma and serum. Results Successful validation of one of the ELISAs was not achieved with particular problems with parallelism and marked differences in measured CA IX concentrations between EDTA plasma and serum. This appeared to be due to a metal ion-dependent epitope on CA IX recognized by the detection antibody in this assay. The other commercially available ELISA examined was successfully validated and showed no difference in CA IX between EDTA plasma and serum. Conclusions These results have important consequences for published studies using this assay where the conclusions drawn from the measurements made may be invalid. This study highlights the need for stringent validation of commercially available assays, including examination of various sample types, before use in research studies.

Although successful and persistent colonization of the gastric mucosa depends on the ability to respond to changing environmental conditions and co-ordinate the expression of virulence factors during the course of infection, Helicobacter pylori possesses relatively few transcriptional regulators. We therefore investigated the contribution of the regulatory protein CsrA to global gene regulation in this important human pathogen. CsrA was necessary for full motility and survival of H. pylori under conditions of oxidative stress. Loss of csrA expression deregulated the oxidant-induced transcriptional responses of napA and ahpC, the acid induction of napA, cagA, vacA, the urease operon, and fur, as well as the heat shock responses of napA, groESL and hspR. Although the level of napA transcript was higher in the csrA mutant, its stability was similar in the wild-type and mutant strains, and less NapA protein was produced in the mutant strain. Finally, H. pylori strains deficient in the production of CsrA were significantly attenuated for virulence in a mouse model of infection. This work provides evidence that CsrA has a broad role in regulating the physiology of H. pylori in response to environmental stimuli, and may be important in facilitating adaptation to the different environments encountered during colonization of the gastric mucosa. Furthermore, CsrA appears to mediate its effects in H. pylori at the post-transcriptional level by influencing the processing and translation of target transcripts, with minimal effect on the stability of the target mRNAs.

We wish to thank Cremolini and colleagues for their interesting letter that highlights their previous finding that VEGFR-2 may be a predictive marker for benefit from bevacizumab therapy. Their data are derived from the setting of postprogression bevacizumab in advanced colorectal cancer, and there