APRIL limits atherosclerosis by binding to heparan sulfate proteoglycans.

Atherosclerotic cardiovascular disease causes heart attacks and strokes, which are the leading causes of mortality worldwide1. The formation of atherosclerotic plaques is initiated when low-density lipoproteins bind to heparan-sulfate proteoglycans (HSPGs)2 and become trapped in the subendothelial space of large and medium size arteries, which leads to chronic inflammation and remodelling of the artery wall2. A proliferation-inducing ligand (APRIL) is a cytokine that binds to HSPGs3, but the physiology of this interaction is largely unknown. Here we show that genetic ablation or antibody-mediated depletion of APRIL aggravates atherosclerosis in mice. Mechanistically, we demonstrate that APRIL confers atheroprotection by binding to heparan sulfate chains of heparan-sulfate proteoglycan 2 (HSPG2), which limits the retention of low-density lipoproteins, accumulation of macrophages and formation of necrotic cores. Indeed, antibody-mediated depletion of APRIL in mice expressing heparan sulfate-deficient HSPG2 had no effect on the development of atherosclerosis. Treatment with a specific anti-APRIL antibody that promotes the binding of APRIL to HSPGs reduced experimental atherosclerosis. Furthermore, the serum levels of a form of human APRIL protein that binds to HSPGs, which we termed non-canonical APRIL (nc-APRIL), are associated independently of traditional risk factors with long-term cardiovascular mortality in patients with atherosclerosis. Our data reveal properties of APRIL that have broad pathophysiological implications for vascular homeostasis.

High Rate of Passenger Lymphocyte Syndrome after ABO Minor Incompatible Lung Transplantation.

Rationale: Passenger lymphocyte syndrome (PLS) may complicate minor ABO mismatched lung transplantation (LuTX) via donor-derived red cell antibody-induced hemolysis.Objectives: To ascertain the incidence and specificity of PLS-relevant antibodies among the study population as well as the dynamics of hemolysis parameters and the transfusion requirement of patients with or without PLS.Methods: In this cohort study, 1,011 patients who received LuTX between January 2010 and June 2019 were studied retrospectively. Prospectively, 87 LuTX (July 2019 to June 2021) were analyzed. Postoperative ABO antibody and hemolytic marker determinations, transfusion requirement, and duration of postoperative hospital care were analyzed. Retrospectively, blood group A recipients of O grafts with PLS were compared with those without.Measurements and Main Results: PLS affected 18.18% (retrospective) and 30.77% (prospective) of A recipients receiving O grafts, 5.13% of B recipients of O grafts, and 20% of AB patients receiving O transplants. Anti-A and anti-A1 were the predominant PLS-inducing antibodies, followed by anti-B and anti-A,B. Significantly lower hemoglobin values (median, 7.4 vs. 8.3 g/dl; P = 0.0063) and an approximately twice as high percentage of patients requiring blood transfusions were seen in PLS. No significant differences in other laboratory markers, duration of hospital stay, or other complications after LuTX were registered.Conclusions: Minor ABO incompatible LuTX recipients are at considerable risk of developing clinically significant PLS. Post-transplant monitoring combining red cell serology and hemolysis marker determination appears advisable so as not to overlook hemolytic episodes that necessitate antigen-negative transfusion therapy.

A neutral comparison of algorithms to minimize L0 penalties for high-dimensional variable selection.

Variable selection methods based on L0 penalties have excellent theoretical properties to select sparse models in a high-dimensional setting. There exist modifications of the Bayesian Information Criterion (BIC) which either control the familywise error rate (mBIC) or the false discovery rate (mBIC2) in terms of which regressors are selected to enter a model. However, the minimization of L0 penalties comprises a mixed-integer problem which is known to be NP-hard and therefore becomes computationally challenging with increasing numbers of regressor variables. This is one reason why alternatives like the LASSO have become so popular, which involve convex optimization problems that are easier to solve. The last few years have seen some real progress in developing new algorithms to minimize L0 penalties. The aim of this article is to compare the performance of these algorithms in terms of minimizing L0 -based selection criteria. Simulation studies covering a wide range of scenarios that are inspired by genetic association studies are used to compare the values of selection criteria obtained with different algorithms. In addition, some statistical characteristics of the selected models and the runtime of algorithms are compared. Finally, the performance of the algorithms is illustrated in a real data example concerned with expression quantitative trait loci (eQTL) mapping.

The role of elective neck dissection in T1 and T2 nasal cavity squamous cell carcinomas.

PURPOSE: To evaluate the role of elective neck dissection (END) on oncological outcome in early-stage nasal cavity squamous cell carcinomas (SCCs). METHODS: In total, 87 patients with T1 (n = 59; 67.8%) and T2 (n = 28; 32.2%) SCCs were evaluated regarding performance of END, regional recurrences (RR) and its impact on cancer-specific survival (CSS). We further created a risk score based on T-classification, tumor subsite and grading to identify patients whom may benefit from END and calculated the corresponding numbers needed to treat (NNT) to prevent RR. RESULTS: Nine (10.3%) patients experienced RR of whom 3 (5.1%) were T1 and 6 (21.4%) T2 tumors (p = 0.042). All RR originated from moderately or poorly differentiated (G2-G3) SCCs of the nasal septum or vestibule. END was done in 15 (17.2%) patients and none of those experienced RR (p = 0.121). Onset of RR represented the worst prognostic factor for CSS (HR 23.3; p = 0.007) with a 5y-CSS of 44.4% vs. 97.3% (p < 0.001). RR occurred in none of the patients with no or low risk scores compared to 31.6% (6/19) in patients with high-risk scores (p = 0.006). Accordingly, three high-risk patients would need to undergo END (NNT 2.63) to prevent RR compared to a NNT of 8 for the whole cohort. CONCLUSIONS: Although rare, occurrence of RR significantly deteriorates outcome in early stage nasal cavity SCCs, which could be effectively reduced by performance of END. The importance of END is currently underestimated and our proposed risk score helps identifying those patients who will benefit from END.

A new nomogram to predict oncological outcome in laryngeal and hypopharyngeal carcinoma patients after laryngopharyngectomy.

BACKGROUND: To create nomograms for better prediction of the oncological outcome in advanced laryngeal (LxCAs) or hypopharyngeal (HpxCAs) cancer after laryngopharyngectomy. MATERIALS: 239 patients who underwent total laryngectomy or laryngopharyngectomy due to LxCA (52.7%) or HpxCA (47.3%) were included in this study. Based on clinical risk factors (tumor site, lymph node involvement, salvage setting), we created nomograms for prediction of disease-specific survival (DSS) and disease-free survival (DFS). RESULTS: HpxCAs showed a higher rate of lymph node involvement (p < 0.001), a 2.47-fold higher risk of a 2nd head and neck cancer (p = 0.009) and significantly worse loco-regional control rates (p = 0.003) compared to LxCAs. Positive neck nodes and salvage procedures were associated with significantly worse outcome. Nomograms demonstrated that hypopharyngeal tumors with positive neck nodes in salvage situations had the worst oncological outcome with a 5-year DSS of 15-20%. CONCLUSIONS: The oncological outcome is worse in hypopharyngeal carcinomas and could be easily quantified by our nomograms that are based on tumor site, lymph node involvement and salvage situation.

Multi-omics profiling predicts allograft function after lung transplantation.

RATIONALE: Lung transplantation is the ultimate treatment option for patients with end-stage respiratory diseases but bears the highest mortality rate among all solid organ transplantations due to chronic lung allograft dysfunction (CLAD). The mechanisms leading to CLAD remain elusive due to an insufficient understanding of the complex post-transplant adaptation processes. OBJECTIVES: To better understand these lung adaptation processes after transplantation and to investigate their association with future changes in allograft function. METHODS: We performed an exploratory cohort study of bronchoalveolar lavage samples from 78 lung recipients and donors. We analysed the alveolar microbiome using 16S rRNA sequencing, the cellular composition using flow cytometry, as well as metabolome and lipidome profiling. MEASUREMENTS AND MAIN RESULTS: We established distinct temporal dynamics for each of the analysed data sets. Comparing matched donor and recipient samples, we revealed that recipient-specific as well as environmental factors, rather than the donor microbiome, shape the long-term lung microbiome. We further discovered that the abundance of certain bacterial strains correlated with underlying lung diseases even after transplantation. A decline in forced expiratory volume during the first second (FEV1) is a major characteristic of lung allograft dysfunction in transplant recipients. By using a machine learning approach, we could accurately predict future changes in FEV1 from our multi-omics data, whereby microbial profiles showed a particularly high predictive power. CONCLUSION: Bronchoalveolar microbiome, cellular composition, metabolome and lipidome show specific temporal dynamics after lung transplantation. The lung microbiome can predict future changes in lung function with high precision.

Preoperative buccal bone volume predicts long-term graft retention following augmentation in the esthetic zone: A retrospective case series.

OBJECTIVES: Buccal bone augmentation in the esthetic zone is routinely used to achieve optimal clinical outcomes. Nonetheless, long-term data are sparse, and it is unknown how baseline buccal bone volume affects the retention of the augmented volume over time. MATERIAL AND METHODS: This is a long-term follow-up retrospective case series. After a preoperative computed tomography scan, implants were placed in the anterior maxilla following guided bone regeneration, autogenous block grafting, or both. At the follow-up, patients received a computed tomography scan and a clinical examination. Buccal bone volume was the primary outcome. Buccal bone thickness, peri-implant, and esthetic parameters were secondary outcomes. RESULTS: After a median follow-up of 6.7 years (interquartile range: 4.9-9.4), 28 implants in 19 patients (median age at augmentation: 43.3 years, interquartile range: 34.4-56.7, 53% female) were followed up. Preoperative buccal bone volume at baseline (V0 ) showed a moderate correlation to final buccal bone volume (Vt , rs  = .43) but a strong correlation to the absolute volumetric change (ΔV = Vt -V0 , rs  = -.80). A linear mixed model for Vt had a large intercept of 91.39 (p < .001) and a rather small slope of .11 for V0 (p = .11). Observed differences between treatments were not statistically significant in the mixed model. V0 above 105 mm3 predicted a negative volume change (ΔV < 0) with a specificity of 100% and a sensitivity of 96%. CONCLUSIONS: The results suggest higher gains in sites with lower V0 and point to a cutoff V0 above which the augmented volume is not retained long-term.

Optical coherence tomography-measured retinal nerve fiber layer thickness values compensated with a multivariate model and discrimination between stable and progressing glaucoma suspects.

PURPOSE: Our previously introduced multivariate model, compensating for intersubject variability, was applied to circumpapillary retinal nerve fiber layer (RNFL) values measured with optical coherence tomography in glaucoma suspects with or without prior progressive optic disc (OD) change in a series of confocal scanning laser tomography (CSLT, HRT III) measurements. METHODS: In this prospective study, OD change during CSLT follow-up was determined with strict, moderate, and liberal criteria of the topographic change analysis (TCA). Model compensation (MC) as well as age compensation (AC) was applied to RNFL sectors (RNFLMC vs. RNFLAC). Diagnostic performance of RNFLMC vs. RNFLAC was tested with an area under the receiver operating characteristic (AUROC) and was compared between methods. RESULTS: Forty-two glaucoma suspects were included. Patients without prior progressive OD change during the CSLT follow-up (= stable) had thicker RNFL thickness values in most areas and for all progression criteria. RNFLMC AUROC for the global RNFL (0.719) and the inferior quadrant (0.711) performed significantly better compared with RNFLAC AUROC (0.594 and 0.631) to discriminate between stable and progressive glaucoma suspects as defined by the moderate criteria of CSLT progression analysis (p = 0.028; p = 0.024). CONCLUSION: MC showed a slight but significant improvement in detection of subjects with prior progressive OD change among a group of glaucoma suspects, when compared to AC, which is the compensation method commonly used during OCT data evaluation in daily routine. Further studies are warranted to validate the present results.

Engagement in purposeful activities and social interactions amongst persons with dementia in special care units compared to traditional nursing homes: An observational study.

BACKGROUND: Persons with dementia spend most time of their day not engaging in activities or social interactions. A care concept of a dementia special care unit that promotes activities and social interaction through a household-like design and individualised stimuli is studied. AIM: To evaluate the main outcomes of the care concept of a dementia special care unit, namely, engagement in activities and social interactions. METHODS: We conducted an observational study as part of a theory-driven evaluation. The Maastricht Daily Life Observation Tool was used to collect momentary assessments multiple times per participant, including engagement in activity, type of activity, engagement in social interaction, interaction partners, location, agitation and mood. Thirty-three residents of the dementia special care units that implemented the care concept and 54 residents with dementia of two traditional nursing homes participated in the study, resulting in 2989 momentary assessments. RESULTS: Residents of both settings did not engage in activities or social interaction in approximately half of the observations. Compared to residents of traditional nursing homes, residents of special care units had a significantly higher chance for engagement in activities and social interaction in the afternoon but not in the morning or evening. CONCLUSION: The care concept of the special care unit facilitated engagement but had its limitations. Further research is needed to develop and evaluate appropriate interventions to enhance engagement of persons with dementia.

Resonance frequency analysis of implants placed in condensed bone.

OBJECTIVES: Resonance frequency analysis (RFA) is used to monitor implant stability. Its output, the Implant Stability Quotient (ISQ), supposedly correlates with insertion torque, a common measurement of primary stability. However, the reliability of RFA in condensed bone remains unclear. MATERIAL AND METHODS: In this human cadaver study in edentulous jaws and fresh extraction sockets, implants were inserted using a split-mouth approach into condensed or untreated bone. Mean ISQ, peak insertion torque, and pre- and postoperative bone volume fractions (BV/TV) were assessed. RESULTS: In edentulous jaws, insertion torque and ISQ correlated both in untreated (r = 0.63, p = 0.02) and in condensed (r = 0.82, p  < 0.01) bone. In extraction sockets, insertion torque and ISQ only correlated in untreated (r = 0.78, p < 0.01), but not in condensed bone (r = 0.15, p = 0.58). In all edentulous jaws, preoperative BV/TV correlated with insertion torque (r = 0.90, p < 0.0001), ISQ (r = 0.64, p < 0.001), and changes in BV/TV (r = -0.71, p < 0.01). In all extraction sockets, preoperative BV/TV did not correlate with either insertion torque (r = 0.33, p = 0.15), ISQ (r = 0.38, p = 0.09), or changes in BV/TV (r = -0.41, p = 0.09). Joint analysis identified preoperative BV/TV as a predictor of postoperative BV/TV (p < 0.001), insertion torque (p < 0.001), and ISQ (p < 0.001). CONCLUSIONS: RFA is feasible for monitoring stability after late implant placement into condensed bone, but not after immediate placement into condensed fresh extraction sites.

A new nomogram to predict the need for tracheostomy in burned patients.

PURPOSE: To evaluate the impact of tracheostomy on complications, dysphagia and outcome in second and third degree burned patients. METHODS: Inpatient mortality, dysphagia, severity of burn injury (ABSI, TBSA) and complications in tracheotomized burn patients were compared to (I) non-tracheotomized burn patients and (II) matched tracheotomized non-burn patients. RESULTS: 134 (30.9%) out of 433 patients who underwent tracheostomy, had a significantly higher percentage of inhalation injury (26.1% vs. 7.0%; p < 0.001), higher ABSI (8.9 ± 2.1 vs. 6.0 ± 2.7; p < 0.001) and TBSA score (41.4 ± 19.7% vs. 18.6 ± 18.8%; p < 0.001) compared to 299 non-tracheotomized burn patients. However, complications occurred equally in tracheotomized burn patients and matched controls and tracheostomy was neither linked to dysphagia nor to inpatient mortality at multivariate analysis. In particular, dysphagia occurred in 6.2% of cases and was significantly linked to length of ICU stay (OR 6.2; p = 0.021), preexisting neurocognitive impairments (OR 5.2; p = 0.001) and patients' age (OR 3.4; p = 0.046). A nomogram was calculated based on age, TBSA and inhalation injury predicting the need for a tracheostomy in severely burned patients. CONCLUSION: Using the new nomogram we were able to predict with significantly higher accuracy the need for tracheostomy in severely burned patients. Moreover, tracheostomy is safe and is not associated with higher incidenc of complications, dysphagia or worse outcome.

A volumetric prediction model for postoperative cyst shrinkage.

OBJECTIVES: With only limited information available on dimensional changes after jaw cyst surgery, postoperative cyst shrinkage remains largely unpredictable. We aimed to propose a model for volumetric shrinkage based on time elapsed since cyst surgery. MATERIAL AND METHODS: We used data from patients that underwent cyst enucleation or decompression between 2007 and 2017 and had at least three computed tomography (CT) scans per patient. We fitted one simple exponential decay model [V(t) = V0 · e-ɑt] and one model with a patient-specific decay rate [Vk(t) = V0 · e-ßt + γkt]. RESULTS: Based on 108 CT scans from 36 patients (median age at surgery: 45.5 years, IQR: 32.3-55.3, 44% female), our simple exponential decay model is V(t) = V0 · e-0.0035t where V(t) is the residual cyst volume after time t elapsed since surgery, V0 is the initial cyst volume, and e is the base of the natural logarithm. Considering a patient-specific decay rate, the model is Vk(t) = V0 · e-0.0049t + γkt where γk is normally distributed, with expectation 0 and standard deviation 0.0041. CONCLUSIONS: Using an exponential regression model, we were able to reliably estimate volumetric shrinkage after jaw cyst surgery. The patient-specific decay rate substantially improved the fit of the model, whereas adding specific covariates as interaction effects to model the decay rate did not provide any significant improvement. CLINICAL RELEVANCE: Estimating postoperative cyst shrinkage is relevant for both treatment planning of jaw cyst surgery as well as evaluating the clinical success of the surgical approach.

Implementation of a nurse-led self-management support intervention for patients with cancer-related pain: a cluster randomized phase-IV study with a stepped wedge design (EvANtiPain).

BACKGROUND: Pain self-management support interventions were effective in controlled clinical trials and meta analyses. However, implementation of these complex interventions may not translate into identical effects. This paper evaluates the implementation of ANtiPain, a cancer pain self-management support intervention in routine clinical practice according to the Reach Efficacy-Adoption Implementation Maintenance framework. METHODS: In this cluster randomized study with a stepped wedge design, N = 153 adult patients with cancer-related pain were recruited from 01/17 to 05/18 on 17 wards of 3 hospitals in Vienna, Austria. ANtiPain entailed a face-to-face in-hospital session by a trained nurse to prepare discharge according to key strategies, information on pain self-management, and skills building. After discharge, cancer-pain self-management was coached via phone calls. Patient-level data were collected at recruitment, and 2, 4 and 8 weeks after discharge via postal or online questionnaire. Primary outcome was pain interference with daily activities. Secondary outcomes included pain intensity, self-efficacy, and patient satisfaction. Organizational-level data (e.g., on implementation procedures) were collected by study or intervention nurses. The mixed model to analyze patient-level data included a random intercept and a random slope for individual and a random intercept for ward. RESULTS: Recruitment was slower than expected and unevenly distributed over wards and hospitals. The face-to-face session was clinically feasible (mean duration = 33 min) as well as the mean amount (n = 2) and duration of phone calls (mean = 17 min). Only 16 (46%) of 35 trained nurses performed the intervention on nine wards. To deal with the loss of power, analyses were adapted. Overall effects on pain interference were not significant. However, effects were significant in sub analyses of the nine wards that recruited patients in the intervention period (p = .009). Regarding secondary outcomes, the group-by-time effect was significant for self-efficacy (p = .033), and patient satisfaction with information on pain-self-management (p = .002) and in-hospital pain management (p = .018). CONCLUSIONS: The implementation of ANtiPain improved meaningful patient outcomes on wards that applied the intervention routinely. Our analyses showed that the implementation benefited from being embedded in larger scale projects to improve cancer pain management and that the selection of wards with a high percentage of oncology patients may be crucial. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02891785 Date of registration: September 8, 2016.

Month-of-birth-effect in multiple sclerosis in Austria.

BACKGROUND: The month-of-birth-effect (MoBE) describes the finding that multiple sclerosis (MS) patients seem to have been born significantly more frequently in spring, with a rise in May, and significantly less often in autumn and winter with the fewest births in November. OBJECTIVES: To analyse if the MoBE can also be found in the Austrian MS population, and if so, whether the pattern is similar to the reported pattern in Canada, United Kingdom, and some Scandinavian countries. METHODS: The data of 7886 MS patients in Austria were compared to all live births in Austria from 1940 to 2010, that is, 7.256545 data entries of the Austrian birth registry and analysed in detail. RESULTS: Patterns observed in our MS cohort were not different from patterns in the general population, even when stratifying for gender. However, the noticeable and partly significant ups and downs over the examined years did not follow the distinct specific pattern with highest birth rates in spring and lowest birth rates in autumn that has been described previously for countries above the 49th latitude. CONCLUSION: After correcting for month-of-birth patterns in the general Austrian population, there is no evidence for the previously described MoBE in Austrian MS patients.

Efficacy and safety of 4 months of sublingual immunotherapy with recombinant Mal d 1 and Bet v 1 in patients with birch pollen-related apple allergy.

BACKGROUND: Birch pollen-related apple allergy is among the most prevalent food allergies in adolescent/adult subjects and mainly results from sensitization to the major birch pollen allergen Bet v 1 and subsequent cross-reaction with the apple protein Mal d 1. However, specific immunotherapy with birch pollen has inconsistent effects on apple allergy. OBJECTIVE: We sought to compare the safety and efficacy of sublingual immunotherapy (SLIT) with 2 formulations containing either rMal d 1 or rBet v 1 on birch pollen-related apple allergy. METHODS: Sixty participants with birch pollen-related apple allergy were randomized to daily sublingual application of placebo (n = 20) or 25 µg of rMal d 1 (n = 20) or rBet v 1 (n = 20) for 16 weeks. Adverse events were regularly recorded. Sublingual challenges with standardized doses of rMal d 1, skin prick tests with recombinant allergens, and measurements of allergen-specific IgE and IgG4 antibodies were performed before and after treatment. RESULTS: Both formulations caused comparable, mainly local adverse events. No systemic reactions occurred. Compared with the placebo and rBet v 1-treated groups, SLIT with rMal d 1 reduced rMal d 1-induced oral symptoms (P = .001 and P = .038) accompanied by longitudinally reduced rMal d 1-specific cutaneous reactions (P = .022) and enhanced IgG4/IgE ratios (P = .012). SLIT with rBet v 1 neither improved the clinical reactivity to rMal d 1 nor enhanced rMal d 1-specific IgG4/IgE ratios. Participants receiving placebo showed no allergen-specific changes. CONCLUSION: Sublingual treatment with a recombinant food allergen was safe and clinically effective, as determined by using standardized challenges. We present a promising approach for the effective treatment of birch pollen-related apple allergy.

Joint genotype- and ancestry-based genome-wide association studies in admixed populations.

In genome-wide association studies (GWAS) genetic loci that influence complex traits are localized by inspecting associations between genotypes of genetic markers and the values of the trait of interest. On the other hand, admixture mapping, which is performed in case of populations consisting of a recent mix of two ancestral groups, relies on the ancestry information at each locus (locus-specific ancestry). Recently it has been proposed to jointly model genotype and locus-specific ancestry within the framework of single marker tests. Here, we extend this approach for population-based GWAS in the direction of multimarker models. A modified version of the Bayesian information criterion is developed for building a multilocus model that accounts for the differential correlation structure due to linkage disequilibrium (LD) and admixture LD. Simulation studies and a real data example illustrate the advantages of this new approach compared to single-marker analysis or modern model selection strategies based on separately analyzing genotype and ancestry data, as well as to single-marker analysis combining genotypic and ancestry information. Depending on the signal strength, our procedure automatically chooses whether genotypic or locus-specific ancestry markers are added to the model. This results in a good compromise between the power to detect causal mutations and the precision of their localization. The proposed method has been implemented in R and is available at http://www.math.uni.wroc.pl/~mbogdan/admixtures/.

Fallback tests for co-primary endpoints.

When efficacy of a treatment is measured by co-primary endpoints, efficacy is claimed only if for each endpoint an individual statistical test is significant at level α. While such a strategy controls the family-wise type I error rate (FWER), it is often strictly conservative and allows for no inference if not all null hypotheses can be rejected. In this paper, we investigate fallback tests, which are defined as uniform improvements of the classical test for co-primary endpoints. They reject whenever the classical test rejects but allow for inference also in settings where only a subset of endpoints show a significant effect. Similarly to the fallback tests for hierarchical testing procedures, these fallback tests for co-primary endpoints allow one to continue testing even if the primary objective of the trial was not met. We propose examples of fallback tests for two and three co-primary endpoints that control the FWER in the strong sense under the assumption of multivariate normal test statistics with arbitrary correlation matrix and investigate their power in a simulation study. The fallback procedures for co-primary endpoints are illustrated with a clinical trial in a rare disease and a diagnostic trial. © 2016 The Authors. Statistics in Medicine published by John Wiley & Sons Ltd.

Quantitative Analysis of Mutant Subclones in Chronic Myeloid Leukemia: Comparison of Different Methodological Approaches.

Identification and quantitative monitoring of mutant BCR-ABL1 subclones displaying resistance to tyrosine kinase inhibitors (TKIs) have become important tasks in patients with Ph-positive leukemias. Different technologies have been established for patient screening. Various next-generation sequencing (NGS) platforms facilitating sensitive detection and quantitative monitoring of mutations in the ABL1-kinase domain (KD) have been introduced recently, and are expected to become the preferred technology in the future. However, broad clinical implementation of NGS methods has been hampered by the limited accessibility at different centers and the current costs of analysis which may not be regarded as readily affordable for routine diagnostic monitoring. It is therefore of interest to determine whether NGS platforms can be adequately substituted by other methodological approaches. We have tested three different techniques including pyrosequencing, LD (ligation-dependent)-PCR and NGS in a series of peripheral blood specimens from chronic myeloid leukemia (CML) patients carrying single or multiple mutations in the BCR-ABL1 KD. The proliferation kinetics of mutant subclones in serial specimens obtained during the course of TKI-treatment revealed similar profiles via all technical approaches, but individual specimens showed statistically significant differences between NGS and the other methods tested. The observations indicate that different approaches to detection and quantification of mutant subclones may be applicable for the monitoring of clonal kinetics, but careful calibration of each method is required for accurate size assessment of mutant subclones at individual time points.

Half the price, twice the gain: How to simultaneously decrease animal numbers and increase precision with good experimental design.

Animal research often involves experiments in which the effect of several factors on a particular outcome is of scientific interest. Many researchers approach such experiments by varying just one factor at a time. As a consequence, they design and analyze the experiments based on a pairwise comparison between two groups. However, this approach uses unreasonably large numbers of animals and leads to severe limitations in terms of the research questions that can be answered. Factorial designs and analyses offer a more efficient way to perform and assess experiments with multiple factors of interest. We will illustrate the basic principles behind these designs, discussing a simple example with only two factors before suggesting how to design and analyze more complex experiments involving larger numbers of factors based on multiway analysis of variance.