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Alternative combination models according for the relative impact of LDH, REC, RLC and the pattern of visceral metastasis. A nomogram was developed using the nomogram function of R (A). For each patient a linear predictor score (risk score) considering the relative impact of four single factors was calculated as follows: LDH-ratio >2.5 - 100 points; presence of visceral metastasis other than lung - 96 points; REC

Calibration of combination models. Calibration was calculated after 12 and 24 months using the calibrate function in the rms package of R for combination model 1 (A) and combination model 2 (B). Bootstrapping (1000 repeats) was performed to obtain bias-corrected estimates of predicted vs. observed values. Non-convergence reduced the number of included bootstrapping steps for combination model 2 to 981 or 990 after 12 or 24 months, respectively. "Predicted" survival probabilities at 12 or 24 months are those predicted by the Cox model, and "observed" refers to the corresponding Kaplan-Meier survival estimate at the given time-point. Mean absolute error in predictions, the mean squared error, and the 0.9 quantile of the absolute error is reported. "Error" refers to the difference between the predicted values and the corresponding bias-corrected calibrated values. Mean error was

Supplementary Figures S1-S2 from Homeostatic Proliferation Plus Regulatory T-Cell Depletion Promotes Potent Rejection of B16 Melanoma

Multivariate Models including only patients receiving 3 mg/kg ipilimumab. Lactate dehydrogenase (LDH), regulatory T cells (Tregs), myeloid-derived suppressor cells (MDSCs), hazard ratio (HR). * Relative lymphocyte count, absolute monocyte count, absolute and relative eosinophil count

Overview of factors, cut-offs, and analysis of overall survival according to cohorts

Discriminatory ability of combination models. The concordance index (c-index, y-axis) was calculated for the combination of factors with independent impact according to Cox regression analysis (model 6.1) and 13 alternative combination models considering 5, 7, or 8 factors (A). The numbers refer to the rows in A. The c-indices are presented according to the number of combined factors (B). The combination model with highest discriminatory ability (7.4), which considered regulatory T cells in addition to the 6 factors with independent impact according to Cox regression analysis was chosen as combination model 1. No further increase of the c-index compared to combination model 1 was observed if one of the 3 remaining factors was additionally considered (models 8.1, 8.2, 8.3). C-indices were calculated for different combination models accounting for the number of unfavorable values of all factors considered in the given model (C). All possible models derived from combinations of the five routine factors were considered. The c-indices are presented according to the number of considered factors (D). The model with highest discriminatory ability (4.1) was selected.

Overall survival (OS) following pembrolizumab according to the count of favorable baseline factors considering death from any reason as "event". In total, 194 patients with complete data of the four baseline markers as considered in the combination model died during follow up. The reasons of death were progressive melanoma (n=189), voluntary physician-assisted ending of life (n=2), or cerebral insult (n=3). In contrast to all other analyses throughout the manuscript, Kaplan-Meier curves of OS according to the number of favorable baseline factors are presented considering death from any reason as "event". The discriminatory ability was marginally higher compared to the analysis of melanoma-specific OS (c-index 0.783 vs. 0.782), and differences in OS remained significant in all pairwise comparisons between the five categories (log rank p=0.046 regarding the difference between patients with 4 or 3 favorable factors, p=0.009 regarding the difference between patients with 1 and 0 favorable factors, p

Spectrum of factors, cut-offs, and differences in overall survival according to biomarkers in the identification and the confirmation cohort. Absolute (Abs.), Relative (Rel.), Lactate dehydrogenase (LDH), regulatory T cells (Tregs), myeloid-derived suppressor cells (MDSCs). * Green characters indicate the category associated with better survival in the identification cohort. ** Green cells indicate significant differences in overall survival (p < 0.05). Red cells indicate non-significant findings.

Correlations between adverse events and overall survival, clinical response, or biomarker categories. Overall survival was not different between patients stratified according to the occurrence of adverse events (AEs) in general (A) or immune-related AEs (irAEs). (B). Kaplan-Meier analysis is presented and censoring is indicated by vertical lines; p-values were calculated by log rank statistics (A&B). No correlations were observed between the occurrence of irAEs during ipilimumab treatment and the best tumor response (C, D) nor with the proposed combination groups of baseline biomarkers according to the combination model 1 (E) or combination model 2 (F). The best overall tumor response according to immune-related response criteria (irRC) was analyzed either as the rate of patients with an irRC response (sum of those with complete or partial responses) or irRC benefit (sum of those with complete responses, partial responses and stable disease). Differences were not statistically significant.

Panels of antibodies used for flow cytometry. * Cells were fixed and permeabilized with FoxP3 buffer (BD). Only frequencies of CD14+ cells from MDSC panel 1 and frequencies of CD4+, CD8+ T cells, as well as their ratio, were included from Treg panel 1. 1 Panel 1 (n=25), 2 Panel 2 (n=184).

Algorithm for cut-off point definition

Purpose: To investigate the antitumor efficacy of T-cell anergy reversal through homeostatic proliferation and regulatory T-cell (Treg) depletion in a clinically relevant murine adoptive immunotherapy model.Experimental Design: B16 melanoma cells were engineered to express the model SIYRYYGL (SIY) antigen to enable immune monitoring. Tumor-specific T cells expanded in tumor-challenged wild-type hosts but became hyporesponsive. To examine whether lymphopenia-induced homeostatic proliferation could reverse tumor-induced T-cell anergy, total splenic T cells were transferred into lymphopenic RAG2−/− mice or control P14/RAG2−/− mice. Tumor growth was measured, and SIY-specific immune responses were monitored using ELISPOT and SIY/Kb tetramers. To determine whether Treg depletion could synergize with homeostatic proliferation, RAG2−/− mice received total or CD25-depleted T cells, followed or preceded by B16.SIY challenge. This approach was further investigated in wild-type mice lymphodepleted with sublethal total body irradiation.Results: Adoptive transfer of total splenic T cells into RAG2−/− mice moderately affected the growth rate of B16.SIY. As Treg expansion occurred in tumor-bearing mice, CD25+ T cells were depleted from total T cells before adoptive transfer. Interestingly, transfer of CD25-depleted T cells into RAG2−/− mice resulted in potent rejection of B16 melanoma in both prophylactic and short-term preimplanted tumor settings and was associated with maintained T-cell effector function. Using a clinically applicable approach, wild-type mice were lymphodepleted using sublethal total body irradiation, which similarly supported tumor rejection upon transfer of CD25-depleted T cells.Conclusions: Our results indicate that combined CD25 depletion and homeostatic proliferation support a potent antitumor immune response—an approach with potential for clinical translation.

Purpose: To identify baseline peripheral blood biomarkers associated with clinical outcome following ipilimumab treatment in advanced melanoma patients.Experimental Design: Frequencies of myeloid-derived suppressor cells (MDSC) and regulatory T cells (Treg), serum lactate dehydrogenase (LDH), routine blood counts, and clinical characteristics were assessed in 209 patients. Endpoints were overall survival (OS) and best overall response. Statistical calculations were done by Kaplan–Meier and Cox regression analysis, including calibration and discrimination by C-statistics.Results: Low baseline LDH, absolute monocyte counts (AMC), Lin−CD14+HLA-DR−/low-MDSC frequencies, and high absolute eosinophil counts (AEC), relative lymphocyte counts (RLC), and CD4+CD25+FoxP3+-Treg frequencies were significantly associated with better survival, and were considered in a combination model. Patients (43.5%) presenting with the best biomarker signature had a 30% response rate and median survival of 16 months. In contrast, patients with the worst biomarkers (27.5%) had only a 3% response rate and median survival of 4 months. The occurrence of adverse events correlated with neither baseline biomarker signatures nor the clinical benefit of ipilimumab. In another model, limited to the routine parameters LDH, AMC, AEC, and RLC, the number of favorable factors (4 vs. 3 vs. 2–0) was also associated with OS (P < 0.001 for all pairwise comparisons) in the main study and additionally in an independent validation cohort.Conclusions: A baseline signature of low LDH, AMC, and MDSCs as well as high AEC, Tregs, and RLC is associated with favorable outcome following ipilimumab. Prospective investigation of the predictive impact of these markers following ipilimumab and other treatments, e.g., PD-1 antibodies, is warranted. Clin Cancer Res; 22(12); 2908–18. ©2016 AACR.

Purpose: Biomarkers for outcome after immune-checkpoint blockade are strongly needed as these may influence individual treatment selection or sequence. We aimed to identify baseline factors associated with overall survival (OS) after pembrolizumab treatment in melanoma patients.Experimental Design: Serum lactate dehydrogenase (LDH), routine blood count parameters, and clinical characteristics were investigated in 616 patients. Endpoints were OS and best overall response following pembrolizumab treatment. Kaplan–Meier analysis and Cox regression were applied for survival analysis.Results: Relative eosinophil count (REC) ≥1.5%, relative lymphocyte count (RLC) ≥17.5%, ≤2.5-fold elevation of LDH, and the absence of metastasis other than soft-tissue/lung were associated with favorable OS in the discovery (n = 177) and the confirmation (n = 182) cohort and had independent positive impact (all P < 0.001). Their independent role was subsequently confirmed in the validation cohort (n = 257; all P < 0.01). The number of favorable factors was strongly associated with prognosis. One-year OS probabilities of 83.9% versus 14.7% and response rates of 58.3% versus 3.3% were observed in patients with four of four compared to those with none of four favorable baseline factors present, respectively.Conclusions: High REC and RLC, low LDH, and absence of metastasis other than soft-tissue/lung are independent baseline characteristics associated with favorable OS of patients with melanoma treated with pembrolizumab. Presence of four favorable factors in combination identifies a subgroup with excellent prognosis. In contrast, patients with no favorable factors present have a poor prognosis, despite pembrolizumab, and additional treatment advances are still needed. A potential predictive impact needs to be further investigated. Clin Cancer Res; 22(22); 5487–96. ©2016 AACR.

Patients treated with pembrolizumab according to clinical site

Overall survival according to the relative lymphocyte and eosinophil counts in 3 subgroups of patients defined by LDH and the pattern of visceral metastasis. Kaplan Meier curves for OS are presented according to relative lymphocyte count (left), relative eosinophil counts (middle) and the count of favorable pre-treatment values considering both factors (right). In patients 132 patients with LDH-ratio {less than or equal to}2.5 AND no visceral metastasis other than lung (A) OS was significantly longer in patients with RLC {greater than or equal to}17.5% (p=0.003) and in those with 2 vs 0 favorable pretreatment values (p=0.022). In 310 patients with either LDH-ratio {less than or equal to}2.5 OR no visceral metastasis other than lung (B) OS was significantly in all pairwise comparisons of the three biomarker categories (all log rank p2.5 AND visceral metastasis other than lung (C) the differences in OS between patients with 2 vs 0 favorable pretreatment factors was not statistically significant (log rank p=0.108).

Relative proportion and survival analyses of all possible combination groups accounting for LDH, pattern of visceral metastasis, RLC and REC

Detailed gating strategy for quantification of subsets of monocytes and myeloid-derived suppressor cells (MDSCs), T cells and regulatory T cells (Tregs). Total cells were selected by gating on Time vs. SSC-A. Duplicates were removed via progressive gating on FSC-H vs. FSC-A and SSC-H vs. SSC-A. Dead cells were excluded by considering only EMA-negative cells. (A) A lineage cocktail (CD3, CD19, CD56) was used to avoid cross-contamination. Previously described MDSC populations were identified as Lin-CD14+HLA-DRlow and Lin-CD14-CD15+CD11b+ within the all-cell gate. Overall monocytes were defined as CD14+, while subsets were separated into classical monocytes (Lin-CD14+CD16-HLA-DR+), non-classical monocytes (Lin-CD14-CD16+HLA-DR+) and Lin- CD14-CD16dimHLA-DR+ monocytes within the all-cell gate. (B) A morphological gate was used to identify the population of lymphocytes. Next, CD3+ cells were selected and further separated into CD4+ and CD8+ cells. Ki67 expression was investigated on CD4+ and CD8+ cells. CD8+ T cells with suppressive potential were defined as CD103+. Previously described phenotypes of Tregs were defined as CD4+CD25+FoxP3+ and CD4+CD127lowCD25+FoxP3+. These were further subdivided into proliferating (Ki67+CD45RA-) and non-proliferating Tregs (Ki67-CD45RA+).

Overall survival and distribution after first dose of ipilimumab according to subsequent treatments. Of 209 patients, 71 received at least one additional systemic line of treatment after ipilimumab. 137 individuals did not receive further therapy and data were not available for one patient. 47 (combination model 1) or 67 (combination model 2) of 71 patients had complete data for classification according to biomarker combination models. The representation of PD-1/PD-L1-treated patients in the biomarker groups was shifted towards favorable biomarker combination groups for both combination models compared to those without subsequent PD-1/PD-L1 treatment. Therefore, a confounding effect of subsequent treatment with PD-1/PD-L1 antibodies on the biomarker results of this study cannot be ruled out (A). To investigate the potential confounding impact on OS and biomarker findings, subsequent treatments were categorized into three different groups: BRAF/MEK inhibitors (n=24), PD-1/PD-L1 antibodies (n=28), and chemotherapy/other treatments (n=33) and analyzed by the Kaplan-Meier method (B). Patients treated with PD-1/PD-L1 antibodies had a significant better survival compared to all 71 patients (p=0.006), while no significant difference was observed for the other two groups. Kaplan Meier analysis of overall survival of patients classified according to combination model 1 (C) or combination model 2 (D) is presented after exclusion of individuals who received subsequent treatment with anti-PD-1 or PD-L1 antibodies, as a confounding effect could not be ruled out. However, the prognostic impact of the proposed biomarker combinations at baseline of ipilimumab treatment remained strong (p

Background and ObjectiveThe spectrum of myelin oligodendrocyte glycoprotein (MOG) antibody–associated disorder (MOGAD) comprises monophasic diseases such as acute disseminated encephalomyelitis (ADEM), optic neuritis (ON), and transverse myelitis and relapsing courses of these presentations. Persistently high MOG antibodies (MOG immunoglobulin G [IgG]) are found in patients with a relapsing disease course. Prognostic factors to determine the clinical course of children with a first MOGAD are still lacking. The objective of the study is to assess the clinical and laboratory prognostic parameters for a risk of relapse and the temporal dynamics of MOG‐IgG titers in children with MOGAD in correlation with clinical presentation and disease course.MethodsIn this prospective multicenter hospital-based study, children with a first demyelinating attack and complete data set comprising clinical and radiologic findings, MOG-IgG titer at onset, and clinical and serologic follow-up data were included. Serum samples were analyzed by live cell-based assay, and a titer level of ≥1:160 was classified as MOG-IgG–positive.ResultsOne hundred sixteen children (f:m = 57:59) with MOGAD were included and initially diagnosed with ADEM (n = 59), unilateral ON (n = 12), bilateral ON (n = 16), myelitis (n = 6), neuromyelitis optica spectrum disorder (n = 8) or encephalitis (n = 6). The median follow-up time was 3 years in monophasic and 5 years in relapsing patients. There was no significant association between disease course and MOG-IgG titers at onset, sex, age at presentation, or clinical phenotype. Seroconversion to MOG-IgG–negative within 2 years of the initial event showed a significant risk reduction for a relapsing disease course. Forty-two/one hundred sixteen patients (monophasic n = 26, relapsing n = 16) had serial MOG-IgG testing in years 1 and 2 after the initial event. In contrast to relapsing patients, monophasic patients showed a significant decrease of MOG-IgG titers during the first and second years, often with seroconversion to negative titers. During the follow-up, MOG-IgG titers were persistently higher in relapsing than in monophasic patients. Decrease in MOG-IgG of ≥3 dilution steps after the first and second years was shown to be associated with a decreased risk of relapses. In our cohort, no patient experienced a relapse after seroconversion to MOG-IgG–negative.DiscussionIn this study, patients with declining MOG-IgG titers, particularly those with seroconversion to MOG-IgG–negative, are shown to have a significantly reduced relapse risk.

Despite remarkably improved outcomes with immune checkpoint inhibition, many patients with metastatic melanoma will eventually require further therapy. Chemotherapy has limited activity when used first-line but can alter the tumour microenvironment and does improve efficacy when used in combination with immunotherapy in lung cancer. Whether chemotherapy after checkpoint inhibitor failure has relevant activity in patients with metastatic melanoma is unknown.Patients with metastatic melanoma treated with chemotherapy after progression on immunotherapy with checkpoint inhibitors were identified retrospectively from 24 melanoma centres. Objective response rate (ORR), progression-free survival (PFS), overall survival (OS) and safety were examined.In total, 463 patients were treated between 2007 and 2017. Fifty-six per cent had received PD-1-based therapy before chemotherapy. Chemotherapy regimens included carboplatin + paclitaxel (32%), dacarbazine (25%), temozolomide (15%), taxanes (9%, nab-paclitaxel 4%), fotemustine (6%) and others (13%). Median duration of therapy was 7.9 weeks (0-108). Responses included 0.4% complete response (CR), 12% partial response (PR), 21% stable disease (SD) and 67% progressive disease (PD). Median PFS was 2.6 months (2.2, 3.0), and median PFS in responders was 8.7 months (6.3, 16.3), respectively. Twelve-month PFS was 12% (95% CI 2-15%). In patients who had received anti-PD-1 before chemotherapy, the ORR was 11%, and median PFS was 2.5 months (2.1, 2.8). The highest activity was achieved with single-agent taxanes (N = 40), with ORR 25% and median PFS 3.9 months (2.1, 6.2). Median OS from chemotherapy start was 7.1 months (6.5, 8.0). Subsequent treatment with checkpoint inhibitors achieved a response rate of 16% with a median PFS of 19.1 months (2.0-43.1 months). No unexpected toxicities were observed.Chemotherapy has a low response rate and short PFS in patients with metastatic melanoma who have failed checkpoint inhibitor therapy, although activity varied between regimens. Chemotherapy has a limited role in the management of metastatic melanoma.

BackgroundFor effective tumor elimination, cytotoxic CD8+ T cells must recognize tumor-derived antigens presented on class I major histocompatibility complex (MHC-I). Despite a general association between the expression of immunogenic antigens, typically neoantigens, and response to immunotherapy, the majority of patients lack strong endogenous responses to most putative neoantigens due to mechanisms that are not well understood. Cytotoxic CD8+ T-cell responses are induced by dendritic cells (DCs) cross-presenting tumor-derived peptides on MHC-I. We hypothesized that cross presentation may form an unappreciated source of bias in the induction of cytotoxic T-cell responses.MethodsWe used stable isotope labeling of amino acids combined with immunopeptidomics to distinguish cross-presented from endogenous MHC-I peptides on DCs. To test impacts on T-cell activation, we targeted the model antigen SIINFEKL to specific subcellular compartments in tumor cells, which were used as sources for cross presentation to T cells. In vitro observations were validated using DNA and RNA sequencing data from two cohorts of patients with melanoma undergoing checkpoint blockade therapy. We used a novel quantitative mass spectrometry approach to measure the levels of model antigen on cross-presenting DCs following various means of tumor cell death.ResultsDCs exhibited a strong bias for cross-presenting peptides derived from cytoplasmic proteins and against those from plasma membrane proteins, which was confirmed using the model antigen SIINFEKL. In patients with melanoma, the proportion of membrane-derived neoantigens was correlated with reduced survival and failure to respond to therapy. Quantification of cross-presented SIINFEKL revealed that the mode of cell death could overcome DCs’ bias against plasma membrane proteins.ConclusionsCross presentation of cellular antigens by DCs may impose constraints on the range of peptides available to activate CD8+ T cells that have previously gone unappreciated. The share of neoantigens arising from membrane-derived sources may render some tumors less immunogenic due to inefficient cross presentation. These observations carry important implications for the encounter and intracellular processing of cellular antigens by DCs and merit further clinical studies for their therapeutic potential in stratifying patient populations and design of vaccine-based therapies.

The oncogenic BRAF inhibitor vemurafenib improves outcomes for patients with advanced BRAFIn this open-label, multicentre study, patients with previously treated or untreated melanoma and the BRAFBetween March 2011 and January 2013, 3224 patients were enrolled, and 3219 patients received ≥1 dose of vemurafenib (safety population); median follow-up time was 33.4 months. Vemurafenib's long-term benefits were confirmed, and no new safety signals identified. The prognostic index showed between-group differences in OS, with tight, non-overlapping confidence intervals. Validation in a pooled group of 666 vemurafenib-treated clinical trial patients revealed a similar pattern; the pattern was similar in 280 patients treated with vemurafenib plus cobimetinib.Final results from the vemurafenib safety study confirm vemurafenib's tolerability in BRAF

Background In neonates with hypoplastic left heart complex (HLHC), biventricular repair is considered superior to univentricular repair. The Z-scores of the mitral and the aortic valve annulus are primary factors for the choice of repair. Predictive cutoff values for the feasibility and optimal outcome of biventricular repair are unknown. This study assesses the growth of left side heart structures and the midterm outcome after biventricular repair with an interatrial fenestration in our HLHC population. Methods Retrospective study of 19 HLHC patients who underwent biventricular repair in a single tertiary referral center between 2004 and 2013. The cardiac dimensions (mitral and aortic valve annulus, left ventricle inlet length, left ventricular internal diastolic dimension) were measured before and at 6, 12, 24, and 48 months after biventricular repair. Results The follow-up ranged from 2 to 98 months. There was no early mortality, and the midterm survival rate was 95%. One patient died of a noncardiac- and nonintervention-related cause. Seven patients (37%) required a total of 8 reinterventions because of recurring or residual obstructive lesions. After biventricular repair, the left cardiac structures grew significantly. Conclusions Neonatal biventricular repair is successful and safe in HLHC patients, even with preoperative mitral and aortic valve annulus Z-scores of −4.5 and −5.5, respectively. Residual or restenosis required reintervention in 37% of our HLHC population, but was not significantly correlated with the magnitude of the preoperative Z-scores. Within the first 6 months of follow-up, the Z-scores almost normalized.

Purpose BECTS (benign childhood epilepsy with centrotemporal spikes) is associated with characteristic EEG findings. This study examines the influence of anti-convulsive treatment on the EEG. Methods In a randomized controlled trial including 43 children with BECTS, EEGs were performed prior to treatment with either Sulthiame or Levetiracetam as well as three times under treatment. Using the spike-wave-index, the degree of EEG pathology was quantified. The EEG before and after initiation of treatment was analyzed. Both treatment arms were compared and the EEG of the children that were to develop recurrent seizures was compared with those that were successfully treated. Results Regardless of the treatment agent, the spike-wave-index was reduced significantly under treatment. There were no differences between the two treatment groups. In an additional analysis, the EEG characteristics of the children with recurrent seizures differed statistically significant from those that did not have any further seizures. Conclusion Both Sulthiame and Levetiracetam influence the EEG of children with BECTS. Persistent EEG pathologies are associated with treatment failures.

Background Hypoplastic left heart complex (HLHC) is characterized by a mitral valve or an aortic valve annular Z score Z scores of the mitral and aortic valve annuli are major determinants of HLHC. Therefore, the algorithm for Z -score calculation is essential for diagnosis. However, no single universal method of calculation is in use. In the scientific literature addressing HLHC, various Z -score calculation methods have been applied. The aim of this study was to evaluate Z scores derived from two-dimensional echocardiographic dimensions in patients with HLHC. Methods To compare the different published methods using two-dimensional echocardiographic measures for Z -score calculation, a cohort of 18 newborns diagnosed with HLHC was retrospectively evaluated. In addition, the methods to determine body surface area in newborns were evaluated. Results Three Z -score calculation methods were included and compared. Using the method of Daubeney et al . to calculate Z scores in our cohort illustrated a lack of correlation beyond a Z score et al . and Pettersen et al . Z scores calculated using Zilberman et al .'s and Pettersen et al .'s methods were fairly consistent. The equations used by Pettersen et al . are based on the largest population of neonates. Conclusion Although the different methods for calculating Z scores for mitral and aortic valve dimensions correspond fairly well in the normal range, Z scores Z -score calculation algorithms used by Pettersen et al . appear to be the most appropriate for use in an evaluation of HLHC. Because these different methods can yield different values, reporting the method as well as the Z score is essential for an accurate diagnosis. Similarly, the method used to determine body surface area should be reported.

Pharmacokinetic monitoring is increasingly becoming an important part of clinical care of tyrosine kinase inhibitor treatment. Vemurafenib is an oral tyrosine kinase inhibitor that inhibits mutated serine/threonine protein kinase B-Raf (BRAF) and is approved for the treatment of adult patients with BRAF V600 mutation-positive unresectable or metastatic melanoma. The aim of this study was to establish the relationship between dried blood spot (DBS) and plasma concentrations of vemurafenib to enable the use of DBS sampling, which is a minimally invasive form of sample collection. In total, 43 paired plasma and DBS samples (in duplicate) were obtained from 8 melanoma patients on vemurafenib therapy and were analyzed using high-performance liquid chromatography-tandem mass spectrometry. Plasma concentrations were predicted from the DBS concentrations using 2 methods: (1) individual hematocrit correction and blood cell-to-plasma partitioning and (2) the calculated slope explaining the relationship between DBS and plasma concentrations (without individual hematocrit correction). Vemurafenib DBS concentrations and plasma concentrations showed a strong correlation (r = 0.964), and the relationship could be described by ([vemurafenib]plasma = [vemurafenib]DBS /0.64). The predicted plasma concentrations were within ±20% of the analyzed plasma concentrations in 97% and 100% of the samples for the methods with and without hematocrit correction, respectively. In conclusion, DBS concentrations and plasma concentrations of vemurafenib are highly correlated. Plasma concentrations can be predicted from DBS concentration using the blood cell-to-plasma partition and the average hematocrit value of this cohort (0.40 L/L). DBS sampling for pharmacokinetic monitoring of vemurafenib treatment can be used in clinical practice.

The aim of the current literature study was to perform a literature review of the factors contributing to exercise limitation and physiological response to exercise in patients with Fontan circulation. In patients with Fontan circulation, peak oxygen uptake ranged from about 14.4 to 32.3 ml/min/kg, and showed a slowed acceleration in the kinetics of oxygen uptake at the onset of exercise. Peak heart rate during exercise was decreased to an average of 153 +/- 10 bpm and arterial oxygen saturation was also decreased at peak exercise, with an average of 89.5 +/- 1.94%. Cardiac output was subnormal, owing to reduced stroke volume, heart rate response and affected pulmonary venous return. Ventilatory anaerobic threshold was below normal values. Moreover, the ventilatory equivalent for carbon dioxide was found to be higher. Patients with Fontan circulation possess a unique physiological response to exercise. Although there is a wide range in exercise capacity among patients, all patients have reduced tolerance to exercise. Cardiac, pulmonary, and muscular factors might play a role in reduced exercise capacity and this distinct response to exercise.

Purpose: To determine exercise response during cardiopulmonary exercise testing in children and adolescents with dystrophinopathies. Methods: Exercise response on the cardiopulmonary exercise test (CPET) was compared with a standard care test protocol. Results: Nine boys (aged 10.8 +/- 4.7 years) with Becker muscular dystrophy (n = 6) and Duchenne muscular dystrophy (n = 3) were included. The feasibility of the CPET was similar to a standard care test protocol, and no serious adverse events occurred. In 67% of the subjects with normal or only mildly impaired functional capacity, the CPET could be used to detect moderate to severe cardiopulmonary exercise limitations. Conclusions: The CPET seems to be a promising outcome measure for cardiopulmonary exercise limitations in youth with mild functional limitations. Further research with larger samples is warranted to confirm current findings and investigate the additional value of the CPET to longitudinal follow-up of cardiomyopathy and the development of safe exercise programs for youth with dystrophinopathies.

We describe the case of a 6-month-old boy with psychomotor retardation, craniofacial dysmorphism, cleft lip and palate, as well as hearing and visual impairment. Analysis of G-banded chromosomes of the propositus showed a de novo interstitial deletion of the short arm of chromosome 12, del(12)(p12.1p12.3). Molecular cytogenetic analysis with bacterial artificial chromosomes (BAC) clones was used to refine the extent of the deletion. The deleted segment encompasses about 12.5 Mb between markers D12S1832 and G62375. The phenotypic consequences of the deletion are discussed and compared with other cases of interstitial deletions of proximal chromosome 12p.

Contains fulltext : 107818.pdf (Publisher’s version ) (Open Access) The management of patients with isolated congenital complete atrioventricular block (CCAVB) has changed during the last decades. The current policy is to pace the majority of patients based on a variety of criteria, among which is limited exercise capacity. Data regarding exercise capacity in this population stems from previous publications reporting small case series of unpaced patients. Therefore, we have investigated the exercise capacity of a group of contemporary children with CCAVB. Sixteen children (mean age 11.5 +/- 4; seven boys, nine girls) with CCAVB were tested. In 13 patients, a median number of three pacemakers were implanted, whereas in three patients no pacemaker was given. All patients had an echocardiogram and completed a cardiopulmonary cycle exercise test. Exercise parameters were determined and compared with reference values obtained from healthy Dutch peers. The peak oxygen uptake/body mass was reduced to 34.4 +/- 9.5 ml kg(-1) min(-1) (79 +/- 24% of predicted) and the ventilatory threshold was reduced to 52 +/- 17% of peak oxygen uptake (78 +/- 21% of predicted), whereas the peak work load/body mass was 2.8 +/- 0.6 W/kg (91 +/- 24% of predicted), which was similar to controls. Importantly, 25% of the paced patients showed upper rate restriction by the pacemaker. In conclusion, children with CCAVB show a reduced peak oxygen uptake and ventilatory threshold, whereas they show normal peak work rates. This indicates that they generate more energy during exercise from anaerobic energy sources. Paced children with CCAVB do not perform better than unpaced children. 01 april 2012