Neuroblastoma Risk Assessment and Treatment Stratification with Hybrid Capture-Based Panel Sequencing.

For many years, the risk-based therapy stratification of children with neuroblastoma has relied on clinical and molecular covariates. In recent years, genome analysis has revealed further alterations defining risk, tumor biology, and therapeutic targets. The implementation of a robust and scalable method for analyzing traditional and new molecular markers in routine diagnostics is an urgent clinical need. Here, we investigated targeted panel sequencing as a diagnostic approach to analyze all relevant genomic neuroblastoma risk markers in one assay. Our "neuroblastoma hybrid capture sequencing panel" (NB-HCSP) assay employs a technology for the high-coverage sequencing (>1000×) of 55 selected genes and neuroblastoma-relevant genomic regions, which allows for the detection of single nucleotide changes, structural rearrangements, and copy number alterations. We validated our assay by analyzing 15 neuroblastoma cell lines and a cohort of 20 neuroblastomas, for which reference routine diagnostic data and genome sequencing data were available. We observed a high concordance for risk markers identified by the NB-HSCP assay, clinical routine diagnostics, and genome sequencing. Subsequently, we demonstrated clinical applicability of the NB-HCSP assay by analyzing routine clinical samples. We conclude that the NB-HCSP assay may be implemented into routine diagnostics as a single assay that covers all essential covariates for initial neuroblastoma classification, extended risk stratification, and targeted therapy selection.

Utilization of the clinical laboratory for the implementation of concussion biomarkers in collegiate football and the necessity of personalized and predictive athlete specific reference intervals.

BACKGROUND: A continued interest in concussion biomarkers makes the eventual implementation of identified biomarkers into routine concussion assessment an eventual reality. We sought to develop and test an interdisciplinary approach that could be used to integrate blood-based biomarkers into the established concussion management program for a collegiate football team. METHODS: We used a CLIA-certified laboratory for all testing and chose biomarkers where clinically validated testing was available as would be required for results used in clinical decision making. We summarized the existing methods and results for concussion assessment across an entire season to identify and demonstrate the challenges with the eventual integration of a parallel process using blood-based tests for concussion management. We analyzed the results of the biomarkers chosen for trends consistent with the outcome assessments provided from the current concussion management protocols. RESULTS: Baseline samples were collected with three additional post-concussion samples collected at three separate time points from players with a diagnosed concussion (n = 12). A summary of results from currently used concussion assessment tools were compared to the representative biomarkers S100B and NSE results. Nine sport-related concussions occurred during practice and three during play. For S100B, 50 % had follow-up testing results lower than the post-injury result. In contrast, 92 % of NSE follow-up results were lower than post-injury. One hundred percent of the results for S100B and NSE were within the athlete-derived reference intervals upon return-to-play and season end. CONCLUSIONS: The reported workflow provides a framework for the eventual implementation of biomarkers for concussion assessment into existing assessment protocols and strengthens the need for reliance on clinical laboratory testing. Athlete-specific reference intervals will be required to adequately interpret results.

Psychosocial factors in sports injury rehabilitation and return to play.

This article discusses the principles and practices that guide psychological intervention with injury, and encourages a psychological approach to injury for clinicians. Part 1 reviews the research literature, and serves as a foundation for the review of clinical practices in part 2. Examination of the research literature highlights 4 areas: (1) psychological factors influencing rehabilitation, (2) social factors affecting rehabilitation, (3) performance concerns among returning athletes, and (4) tools/inventories for assessing psychological readiness to return. A synopsis of an injury intervention plan is provided, and the influence of pain and fear in the rehabilitation process is described.

A systematic review of the biomarker S100B: implications for sport-related concussion management.

OBJECTIVE: Elevated levels of the astroglial protein S100B have been shown to predict sport-related concussion. However, S100B levels within an athlete can vary depending on the type of physical activity (PA) engaged in and the methodologic approach used to measure them. Thus, appropriate reference values in the diagnosis of concussed athletes remain undefined. The purpose of our systematic literature review was to provide an overview of the current literature examining S100B measurement in the context of PA. The overall goal is to improve the use of the biomarker S100B in the context of sport-related concussion management. DATA SOURCES: PubMed, SciVerse Scopus, SPORTDiscus, CINAHL, and Cochrane. STUDY SELECTION: We selected articles that contained (1) research studies focusing exclusively on humans in which (2) either PA was used as an intervention or the test participants or athletes were involved in PA and (3) S100B was measured as a dependent variable. DATA EXTRACTION: We identified 24 articles. Study variations included the mode of PA used as an intervention, sample types, sample-processing procedures, and analytic techniques. DATA SYNTHESIS: Given the nonuniformity of the analytical methods used and the data samples collected, as well as differences in the types of PA investigated, we were not able to determine a single consistent reference value of S100B in the context of PA. Thus, a clear distinction between a concussed athlete and a healthy athlete based solely on the existing S100B cutoff value of 0.1 µg/L remains unclear. However, because of its high sensitivity and excellent negative predictive value, S100B measurement seems to have the potential to be a diagnostic adjunct for concussion in sports settings. We recommend that the interpretation of S100B values be based on congruent study designs to ensure measurement reliability and validity.

Catechol-O-methyltransferase (COMT) genotype affects age-related changes in plasticity in working memory: a pilot study.

OBJECTIVES: Recent work suggests that a genetic variation associated with increased dopamine metabolism in the prefrontal cortex (catechol-O-methyltransferase Val158Met; COMT) amplifies age-related changes in working memory performance. Research on younger adults indicates that the influence of dopamine-related genetic polymorphisms on working memory performance increases when testing the cognitive limits through training. To date, this has not been studied in older adults. METHOD: Here we investigate the effect of COMT genotype on plasticity in working memory in a sample of 14 younger (aged 24-30 years) and 25 older (aged 60-75 years) healthy adults. Participants underwent adaptive training in the n-back working memory task over 12 sessions under increasing difficulty conditions. RESULTS: Both younger and older adults exhibited sizeable behavioral plasticity through training (P < .001), which was larger in younger as compared to older adults (P < .001). Age-related differences were qualified by an interaction with COMT genotype (P < .001), and this interaction was due to decreased behavioral plasticity in older adults carrying the Val/Val genotype, while there was no effect of genotype in younger adults. DISCUSSION: Our findings indicate that age-related changes in plasticity in working memory are critically affected by genetic variation in prefrontal dopamine metabolism.

Blood-based biomarkers for traumatic brain injury: evaluation of research approaches, available methods and potential utility from the clinician and clinical laboratory perspectives.

Blood-based biomarkers for traumatic brain injury (TBI) have been investigated and proposed for decades, yet the current clinical assessment of TBI is largely based on clinical symptoms that can vary widely amongst patients, and have significant overlap with unrelated disease states. A careful review of current treatment guidelines for TBI further highlights the potential utility of a blood-based TBI biomarker panel in augmenting clinical decision making. Numerous expert reviews on blood-based TBI biomarkers have been published but a close look at the methods used and the astonishing paucity of validation and quality control data has not been undertaken from the vantage point of the clinical laboratory. Further, the field of blood-based TBI biomarker research has failed to adequately examine sex and gender differences between men and women with respect to the clinical care settings, as well as differences in physiological outcomes of TBI biomarker studies. Discussions of tried-and-true laboratory techniques in addition to a few new ones already operating in the clinical laboratory are summarized with a consideration of their utility in TBI biomarker assessment. In the context of TBI biomarkers, the central concerns discussed in this review are the readiness of the clinical laboratory, the willingness of the research environment and the inherent ability of each to radically affect patient outcomes in TBI.

Working memory training improvements and gains in non-trained cognitive tasks in young and older adults.

Previous studies on working memory training have indicated that transfer to non-trained tasks of other cognitive domains may be possible. The aim of this study is to compare working memory training and transfer effects between younger and older adults (n = 60). A novel approach to adaptive n-back training (12 sessions) was implemented by varying the working memory load and the presentation speed. All participants completed a neuropsychological battery of tests before and after the training. On average, younger training participants achieved difficulty level 12 after training, while older training participants only reached difficulty level 5. In younger participants, transfer to Verbal Fluency and Digit Symbol Substitution test was found. In older participants, we observed a transfer to Digit Span Forward, CERAD Delayed Recall, and Digit Symbol Substitution test. Results suggest that working memory training may be a beneficial intervention for maintaining and improving cognitive functioning in old age.

Expression of NTRK1/TrkA affects immunogenicity of neuroblastoma cells.

High levels of the NTRK1/TrkA receptor are expressed in low-stage neuroblastomas, which are characterized by a good patient prognosis and often undergo spontaneous regression. In addition to apoptosis, tumor-immune responses might contribute to this regression. We hypothesized that TrkA expression might enhance the immune response to neuroblastomas. Immunohistochemistry on neuroblastoma tissue microarrays confirmed significantly higher lymphocyte infiltration in low-stage compared with high-stage tumors. Flow cytometry of human SH-SY5Y cells stably transfected with NTRK1/TrkA cDNA revealed significant upregulation of major histocompatibility complex (MHC) class I complexes on TrkA-expressing cells. Corresponding to this upregulation, T cell activity and cytoxicity was enhanced in the presence of SY5Y-TrkA cells or by medium conditioned by them, suggesting the existence of additional soluble factors stimulating the T cell response. Activation of natural killer (NK) cells was only increased in the presence of SY5Y-TrkA conditioned medium (CM) and not in co-culture assays, suggesting a dominant inhibitory effect of upregulated MHC class I as the primary NK cell escape mechanism of TrkA-expressing neuroblastomas. We reanalyzed gene expression data obtained from the cell culture model to identify additional genes involved in the TrkA-mediated modulation of immune responses. Upregulation of selected target genes in SY5Y-TrkA cells was confirmed on transcript and protein levels. However, none of the proteins were detected in medium conditioned by SY5Y-TrkA cells, arguing against these factors as soluble mediators of the TrkA-induced immune response. We here provide evidence that TrkA expression in neuroblastoma leads to an increased immunogenicity that may contribute to a less malignant phenotype and/or spontaneous regression of neuroblastoma cells.

The impact of increased blood lactate on serum S100B and prolactin concentrations in male adult athletes.

S100B is an astroglial protein that is increased in the peripheral bloodstream after traumatic brain injury (TBI). Elevated serum levels of S100B have been shown to be predictive of mild TBI. Furthermore, physical activity (PA) can affect S100B levels. Interestingly, increased serum S100B concentrations have been detected in athletes without apparent TBI. Such increases could be attributed to tissue hypoperfusion reflected by blood lactate concentrations [BLa(-)] and/or increased serotonergic activity reflected by prolactin (PRL). The impact of increased blood lactates on peripheral S100B levels per se are yet unknown. The purpose of our study was to investigate if increased blood lactate induced by sodium lactate infusion, without the "side effects" of PA, resulted in changes in serum S100B and PRL. Twelve male adults were given a sodium lactate infusion for a period of 24 min by a perfusor with an infusion rate of 0.01 mL kg(-1) min(-1), increased every 3 min. The main outcome measures showed no increase in serum S100B (p > 0.05). Prolactin increased significantly (p < 0.05) after [BLa(-)] exceeded a concentration of 4 mmol L(-1). Furthermore, the expected values of blood lactate achieved peak values ranging from 11 to 15 mmol L(-1). We conclude that neither increased blood lactate nor serum PRL play an exclusive role in the regulation of S100B. Nevertheless, PA should be surveyed in medical history and critically assessed in determining the severity of TBI, especially in sports. Further studies are needed to clarify the impact of PA on the biomarker S100B.

Targeted expression of mutated ALK induces neuroblastoma in transgenic mice.

Activating anaplastic lymphoma kinase (ALK) mutations were recently detected in most familial and 10% of sporadic neuroblastomas. However, the role of mutated ALK in tumorigenesis remains elusive. We demonstrate that targeted expression of the most frequent and aggressive variant, ALK(F1174L), is tumorigenic in mice. Tumors resembled human neuroblastomas in morphology, metastasis pattern, gene expression, and the presence of neurosecretory vesicles as well as synaptic structures. This ALK-driven neuroblastoma mouse model precisely recapitulated the genetic spectrum of the disease. Chromosomal aberrations were syntenic to those in human neuroblastoma, including 17q gain and MYCN oncogene amplification. Targeted ALK(F1174L) and MYCN coexpression revealed a strong synergism in inducing neuroblastoma with minimal chromosomal aberrations, suggesting that fewer secondary hits are required for tumor induction if both oncoproteins are targeted. Treatment of ALK(F1174L) transgenic mice with the ALK inhibitor TAE-684 induced complete tumor regression, indicating that tumor cells were addicted to ALK(F1174L) activity. We conclude that an activating mutation within the ALK kinase domain is sufficient to induce neuroblastoma development, and ALK inhibitors show promise for treating human neuroblastomas harboring ALK mutations.

Serum Concentrations of S100B are not Affected by Cycling to Exhaustion With or Without Vibration.

The calcium-binding protein S100B is produced primarily by astrocytes and exerts concentration-dependent paracrine and autocrine effects on neurons and glia. The numerous findings of a correlation between S100B and traumatic brain injury (TBI) have resulted in the employment of this protein as a clinical biomarker for such injury. Our present aim was to determine whether cycling with (V) or without (NV) vibration alters serum concentrations of S100B. Twelve healthy, male non-smokers (age: 25.3±1.6 yrs, body mass: 74.2±5.9 kg, body height: 181.0±3.7 cm, VO2peak: 56.9±5.1 ml·min(-1)·kg(-1) (means ± SD)) completed in random order two separate trials to exhaustion on a vibrating bicycle (amplitude 4 mm and frequency 20 Hz) connected to an ergometer. The initial workload of 100 W was elevated by 50 W every 5 min and the mean maximal period of exercise was 25:27±1:30 min. The S100B in venous blood taken at rest, immediately after the test, and 30, 60 and 240 min post-exercise exhibited no significant differences (p>0.05), suggesting that cycling with and without vibration does not influence this parameter.

Lactate infusion at rest increases BDNF blood concentration in humans.

Studies in humans use blood lactate to determine the degree of the exercise intensity, suggesting that exercise with elevated blood lactate concentrations results in increased BDNF plasma concentrations. However, it is not clear if lactate per se or rather other mechanisms are responsible for changes in blood BDNF concentrations. The lactate clamp method at rest is an appropriate method to examine physiological responses of lactate on the human organism without the effects of exercise. Eight male sport students placed in a sitting position received intravenous infusions with a 4 molar sodium-lactate solution in an incremental design starting with an infusion rate of 0.01ml/kgBW/min for the first three minutes, which was increased every three minutes by 0.01ml/kgBW/min up to 0.08ml/kg/min in the 24th minute. All together each subject received 4.2mmol of infusion. Venous blood samples were taken before and immediately after the infusion as well as in the 24th and the 60th min after the infusion period and analysed for BDNF. Blood gases and capillary blood lactate (La) were analysed before the test, every three minutes directly before increasing the infusion rate, at the end of the infusion and in the post infusions period until the 12th min and after 24 and 60min. BDNF and La increased significantly after the infusion and reached baseline values at the end of the experiment (p<0.05, p<0.01, respectively). pH and hydrogen ions increased from the beginning until the end of the infusion period (p<0.01). This data suggest that blood lactate is involved in the regulation of BDNF blood concentrations.

The miR-17-92 microRNA cluster regulates multiple components of the TGF-ß pathway in neuroblastoma.

The miR-17-92 microRNA cluster is often activated in cancer cells, but the identity of its targets remains elusive. Using SILAC and quantitative mass spectrometry, we examined the effects of activation of the miR-17-92 cluster on global protein expression in neuroblastoma (NB) cells. Our results reveal cooperation between individual miR-17-92 miRNAs and implicate miR-17-92 in multiple hallmarks of cancer, including proliferation and cell adhesion. Most importantly, we show that miR-17-92 is a potent inhibitor of TGF-ß signaling. By functioning both upstream and downstream of pSMAD2, miR-17-92 activation triggers downregulation of multiple key effectors along the TGF-ß signaling cascade as well as direct inhibition of TGF-ß-responsive genes.