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Fms‐like tyrosine kinase 3 (FLT3) internal tandem duplication (ITD) mutations, common in pediatric acute myeloid leukemia (AML), associate with early relapse and poor prognosis. Past studies have suggested additional cooperative mutations are required for leukemogenesis in FLT3‐ITD+ AML. Using RNA sequencing and a next‐generation targeted gene panel, we broadly characterize the co‐occurring genomic alterations in pediatric cytogenetically normal (CN) FLT3‐ITD+ AML to gain a deeper understanding of the clonal patterns and heterogeneity at diagnosis and relapse. We show that chimeric transcripts were present in 21 of 34 (62%) of de novo samples, 2 (6%) of these samples included a rare reoccurring fusion partner BCL11B. At diagnosis, the median number of mutations other than FLT3 per patient was 1 (range 0–3), which involved 8 gene pathways; WT1 and NPM1 mutations were frequently observed (35% and 24%, respectively). Fusion transcripts and high variant allele frequency (VAF) mutants, which included WT1,NPM1,SMARCA2,RAD21, and TYK2, were retained from diagnosis to relapse. We did observe reduction in VAF of simple or single mutation clones, but VAFs were preserved or expanded in more complex clones with multiple mutations. Our data provide the first insight into the genomic complexity of pediatric CN FLT3‐ITD+ AML and could help stratify future targeted treatment strategies. [ABSTRACT FROM AUTHOR]

Despite initial remission, ∼60–70% of adult and 30% of pediatric patients experience relapse or refractory AML. Studies so far have identified base line gene expression profiles of pathogenic and prognostic significance in AML; however, the extent of change in gene expression post-initiation of treatment has not been investigated. Exposure of leukemic cells to chemotherapeutic agents such as cytarabine, a mainstay of AML chemotherapy, can trigger adaptive response by influencing leukemic cell transcriptome and, hence, development of resistance or refractory disease. It is, however, challenging to perform such a study due to lack of availability of specimens post-drug treatment. The primary objective of this study was to identifyin vivocytarabine-induced changes in leukemia cell transcriptome and to evaluate their impact on clinical outcome. The results highlight genes relevant to cytarabine resistance and support the concept of targeting cytarabine-induced genes as a means of improving response. [ABSTRACT FROM AUTHOR]

Motivation: Normalization is critical in DNA copy number analysis. We propose a new method to correctly identify two-copy probes from the genome to obtain representative references for normalization in single nucleotide polymorphism arrays. The method is based on a two-state Hidden Markov Model. Unlike most currently available methods in the literature, the proposed method does not need to assume that the percentage of two-copy state probes is dominant in the genome, as long as there do exist two-copy probes.Results: The real data analysis and simulation study show that the proposed algorithm is successful in that (i) it performs as well as the current methods (e.g. CGHnormaliter and popLowess) for samples with dominant two-copy states and outperforms these methods for samples with less dominant two-copy states; (ii) it can identify the copy-neutral loss of heterozygosity; and (iii) it is efficient in terms of the computational time used.Availability: R scripts are available at http://publichealth.lsuhsc.edu/PAIR.html.Contact: zfang@lsuhsc.eduSupplementary information: Supplementary data are available at Bioinformatics online. [ABSTRACT FROM PUBLISHER]

BACKGROUND. Older age has historically been an adverse prognostic factor in pediatric acute myeloid leukemia (AML). To the authors' knowledge, the impact of age relative to that of other prognostic factors on the outcome of patients treated in recent trials is unknown. METHODS. Clinical outcome and causes of treatment failure of 351 patients enrolled on 3 consecutive protocols for childhood AML between 1991 and 2008 were analyzed according to age and protocol. RESULTS. The more recent protocol (AML02) produced improved outcomes for patients aged 10 years to 21 years compared with 2 earlier studies (AML91 and AML97), with 3-year rates of event-free survival (EFS), overall survival (OS), and cumulative incidence of refractory leukemia or recurrence (CIR) for this group being similar to those of patients aged birth to 9 years: EFS: 58.3% ± 5.4% versus 66.6% ± 4.9% ( P = .20); OS: 68.9% ± 5.1% versus 75.1% ± 4.5% ( P = .36); and CIR: 21.9% ± 4.4% versus 25.3% ± 4.2% ( P = .59). The EFS and OS estimates for patients aged 10 to 15 years overlapped those for patients aged 16 to 21 years. However, the cumulative incidence of toxic death was significantly higher for patients aged 10 to 21 years compared with younger patients (13.2% ± 3.6% vs 4.5% ± 2.0%; P = .028). CONCLUSIONS. The survival rate for older children with AML has improved on the results of a recent trial and is now similar to that of younger patients. However, deaths from toxicity remain a significant problem for patients in the older age group. Future trials should focus on improving supportive care while striving to develop more effective antileukemic therapy. Cancer 2012. © 2012 American Cancer Society. [ABSTRACT FROM AUTHOR]

Differential expression analysis of sequence-count expression data involves performing a large number of hypothesis tests that compare the expression count data of each gene or transcript across two or more biological conditions. The assumptions of any specific hypothesis-testing method will probably not be valid for each of a very large number of genes. Thus, computational evaluation of assumptions should be incorporated into the analysis to select an appropriate hypothesis-testing method for each gene. Here, we generalize earlier work to introduce two novel procedures that use estimates of the empirical Bayesian probability (EBP) of overdispersion to select or combine results of a standard Poisson likelihood ratio test and a quasi-likelihood test for each gene. These EBP-based procedures simultaneously evaluate the Poisson-distribution assumption and account for multiple testing. With adequate power to detect overdispersion, the new procedures select the standard likelihood test for each gene with Poisson-distributed counts and the quasi-likelihood test for each gene with overdispersed counts. The new procedures outperformed previously published methods in many simulation studies. We also present a real-data analysis example and discuss how the framework used to develop the new procedures may be generalized to further enhance performance. An R code library that implements the methods is freely available at www.stjuderesearch.org/depts/biostats/software. [ABSTRACT FROM AUTHOR]

The analysis of microarray data often involves performing a large number of statistical tests, usually at least one test per queried gene. Each test has a certain probability of reaching an incorrect inference; therefore, it is crucial to estimate or control error rates that measure the occurrence of erroneous conclusions in reporting and interpreting the results of a microarray study. In recent years, many innovative statistical methods have been developed to estimate or control various error rates for microarray studies. Researchers need guidance choosing the appropriate statistical methods for analysing these types of data sets. This review describes a family of methods that use a set of P-values to estimate or control the false discovery rate and similar error rates. Finally, these methods are classified in a manner that suggests the appropriate method for specific applications and diagnostic procedures that can identify problems in the analysis are described. [ABSTRACT FROM AUTHOR]

The methodological advancement in microarray data analysis on the basis of false discovery rate (FDR) control, such as the q-value plots, allows the investigator to examinethe FDR from several perspectives. However, when FDR control at the 'customary' levels 0.01, 0.05, or 0.1 does not provide fruitful findings, there is little guidance for making thetrade off between the significance threshold and the FDR level by sound statistical or biologicalconsiderations. Thus, meaningful statistical significance criteria that complementthe existing FDR methods for large-scale multiple tests are desirable. Three statistical significance criteria, the profile information criterion, the total error proportion, and the guide-gene driven selection, are developed in this research. The first two are general significance threshold criteria for large-scale multiple tests; the profile information criterion is related to the recent theoretical studies of the connection between FDR control andminimax estimation, and the total error proportion is closely related to the asymptoticproperties of FDR control in terms of the total error risk. The guide-gene driven selection is an approach to combining statistical significance and the existing biological knowledge of the study at hand. Error properties of these criteria are investigated theoretically and by simulation. The proposed methods are illustrated and compared using an example of genomic screening for novel Arf gene targets. Operating characteristics of q-value andthe proposed significance threshold criteria are investigated and compared in a simulationstudy that employs a model mimicking a gene regulatory pathway. A guideline for usingthese criteria is provided. Splus/R code is available from the corresponding author uponrequest. [ABSTRACT FROM AUTHOR]

The role of somatic mitochondrial DNA (mtDNA) mutations in leukemogenesis remains poorly characterized. To determine the impact of somatic mtDNA mutations on this process, we assessed the leukemogenic potential of hematopoietic progenitor cells (HPCs) from mtDNA mutator mice (Polg D257A) with or without NMyc overexpression. We observed a higher incidence of spontaneous leukemogenesis in recipients transplanted with heterozygous Polg HPCs and a lower incidence of NMyc-driven leukemia in those with homozygous Polg HPCs compared to controls. Although mtDNA mutations in heterozygous and homozygous HPCs caused similar baseline impairments in mitochondrial function, only heterozygous HPCs responded to and supported altered metabolic demands associated with NMyc overexpression. Homozygous HPCs showed altered glucose utilization with pyruvate dehydrogenase inhibition due to increased phosphorylation, exacerbated by NMyc overexpression. The impaired growth of NMyc-expressing homozygous HPCs was partially rescued by inhibiting pyruvate dehydrogenase kinase, highlighting a relationship between mtDNA mutation burden and metabolic plasticity in leukemogenesis. [ABSTRACT FROM AUTHOR]

Reduced expression of the uptake transporter, OCTN1 (SLC22A4), has been reported as a strong predictor of poor event‐free and overall survival in multiple cohorts of patients with acute myeloid leukemia (AML) receiving the cytidine nucleoside analog, cytarabine (Ara‐C). To further understand the mechanistic basis of interindividual variability in the functional expression of OCTN1 in AML, we hypothesized a mechanistic connection to DNA methylation‐based epigenetic repression of SLC22A4. We found increased basal SLC22A4 methylation was associated with decreased Ara‐C uptake in AML cell lines. Pre‐treatment with hypomethylating agents, 5‐azacytidine, or decitabine, restored SLC22A4 mRNA expression, increased cellular uptake of Ara‐C, and was associated with increased cellular sensitivity to Ara‐C compared with vehicle‐treated cells. Additionally, lower SLC22A4 methylation status was associated with distinct clinical advantages in both adult and pediatric patients with AML. These findings suggest a regulatory mechanism is involved in the interindividual variability in response to Ara‐C, and provides a basis for the integration of hypomethylating agents into Ara‐C‐based treatment regimens. [ABSTRACT FROM AUTHOR]

The false discovery rate (FDR) is a widely used metric of statistical significance for genomic data analyses that involve multiple hypothesis testing. Power and sample size considerations are important in planning studies that perform these types of genomic data analyses. Here, we propose a three-rectangle approximation of a p-value histogram to derive a formula to compute the statistical power and sample size for analyses that involve the FDR. We also introduce the R package FDRsamplesize2, which incorporates these and other power calculation formulas to compute power for a broad variety of studies not covered by other FDR power calculation software. A few illustrative examples are provided. The FDRsamplesize2 package is available on CRAN. [ABSTRACT FROM AUTHOR]

Consolidation therapies for children with intermediate- or high-risk acute myeloid leukemia (AML) are urgently needed to achieve higher cure rates while limiting therapy-related toxicities. We determined if adoptive transfer of natural killer (NK) cells from haploidentical killer immunoglobulin--like receptor (KIR)--human leukocyte antigen (HLA)-mismatched donors may prolong event-free survival in children with intermediate-risk AML who were in first complete remission after chemotherapy. Patients received cyclophosphamide (Day -- 7), fludarabine (Days -- 6 through -- 2), and subcutaneous interleukin-2 (Days -- 1, 1, 3, 5, 7, and 9). Purified, unmanipulated NK cells were infused on Day 0, and NK cell chimerism and phenotyping from peripheral blood were performed on Days 7, 14, 21, and 28. As primary endpoint, the event-free survival was compared to a cohort of 55 patients who completed chemotherapy and were in first complete remission but did not receive NK cells. Donor NK cell kinetics were determined as secondary endpoints. Twenty-one patients (median age at diagnosis, 6.0 years [range, 0.1--15.3 years]) received a median of 12.5 x 106 NK cells/kg (range, 3.6--62.2 x 106 cells/kg) without major side effects. All but 3 demonstrated transient engraftment with donor NK cells (median peak donor chimerism, 4% [range, 0-43%]). KIR--HLA-mismatched NK cells expanded in 17 patients (81%) and contracted in 4 (19%). However, adoptive transfer of NK cells did not decrease the cumulative incidence of relapse (0.393 [95% confidence interval: 0.182-0.599] vs. 0. 35 [0.209--0.495]; P = .556) and did not improve event-free (60.7 ± 10.9% vs. 69.1 ± 6.8%; P = .553) or overall survival (84.2 ± 8.5% vs. 79.1 ± 6.6%; P = .663) over chemotherapy alone. The lack of benefit may result from insufficient numbers and limited persistence of alloreactive donor NK cells but does not preclude its potential usefulness during other phases of therapy, or in combination with other immunotherapeutic agents. [ABSTRACT FROM AUTHOR]

Genome-wide association studies have discovered many biologically important associations of genes with phenotypes. Typically, genome-wide association analyses formally test the association of each genetic feature (SNP, CNV, etc) with the phenotype of interest and summarize the results with multiplicity-adjusted p-values. However, very small p-values only provide evidence against the null hypothesis of no association without indicating which biological model best explains the observed data. Correctly identifying a specific biological model may improve the scientific interpretation and can be used to more effectively select and design a follow-up validation study. Thus, statistical methodology to identify the correct biological model for a particular genotype-phenotype association can be very useful to investigators. Here, we propose a general statistical method to summarize how accurately each of five biological models (null, additive, dominant, recessive, co-dominant) represents the data observed for each variant in a GWAS study. We show that the new method stringently controls the false discovery rate and asymptotically selects the correct biological model. Simulations of two-stage discovery-validation studies show that the new method has these properties and that its validation power is similar to or exceeds that of simple methods that use the same statistical model for all SNPs. Example analyses of three data sets also highlight these advantages of the new method. An R package is freely available at www.stjuderesearch.org/site/depts/biostats/maew . [ABSTRACT FROM AUTHOR]

Background: Hyperleukocytosis in patients with acute myeloid leukemia (AML) has been associated with worse outcomes. For cytoreduction, leukapheresis has been used but its clinical utility is unknown, and low‐dose cytarabine (LD‐cytarabine) is used as an alternative method. Methods: Children with newly diagnosed AML treated between 1997 and 2017 in institutional protocols were studied. Hyperleukocytosis was defined as a leukocyte count of ≥100 × 109/L at diagnosis. Clinical characteristics, early complications, survival data, and effects of cytoreductive methods were reviewed. Among 324 children with newly diagnosed AML, 49 (15.1%) presented with hyperleukocytosis. Initial management of hyperleukocytosis included leukapheresis or exchange transfusion (n = 16, considered as one group), LD‐cytarabine (n = 18), hydroxyurea (n = 1), and no leukoreduction (n = 14). Results: Compared with patients who received leukapheresis, the percentage decrease in leukocyte counts following intervention was greater among those who received LD‐cytarabine (48% vs. 75%; p =.02), with longer median time from diagnosis to initiation of protocol therapy (28.1 vs. 95.2 hours; p <.001). The incidence of infection was higher in patients (38%) who had leukapheresis than those who receive LD‐cytarabine (0%) or leukoreduction with protocol therapy (14%) (p =.008). No differences were noted in the outcomes among the intervention groups. Although patients with hyperleukocytosis had higher incidences of pulmonary and metabolic complications than did those without, no early deaths occurred, and the complete remission, event‐free survival, overall survival rates, and outcomes of both groups were similar. Conclusion: LD‐cytarabine treatment appears to be a safe and effective means of cytoreduction for children with AML and hyperleukocytosis. Among 324 children with newly diagnosed acute myeloid leukemia (AML), 49 (15.1%) presented with hyperleukocytosis and were managed with leukapheresis or exchange transfusion (n = 16), low‐dose (LD)‐cytarabine (n = 18), hydroxyurea (n = 1), or no leukoreduction (n = 14). No early deaths were observed and LD‐cytarabine treatment appears to be a safe and effective means of cytoreduction for children with AML and hyperleukocytosis. [ABSTRACT FROM AUTHOR]

Background Differential gene expression analysis typically applies a statistical hypothesis testing procedure to the (log-transformed) data for each gene and then adjusts for multiple testing. However, it is not always clear whether a parametric or non-parametric test of hypothesis is preferred for a specific application. In a less complex application, a statistical test or graphical assessment of the fit of the assumed parametric model (e.g. the normal distribution) could be used to make this determination. However, in the analysis of microarray data it is unclear how to assess assumption adequacy for each gene and use it to guide subsequent analysis. Therefore, we introduce the principle of empirical Bayes assumption adequacy averaging and use it to develop a robust method to perform differential expression analysis. The proposed method combines results from the t-test and nonparametric rank-sum test according to the results of assessing the assumption of normality using the Shapiro-Wilk procedure. Through this averaging process, the proposed method is able to rely more heavily on the statistical test that the data suggests is superior for each individual gene. Subsequently, we observed that the proposed method has desirable power and robustness properties in a series of traditional and bootstrap-based simulation studies. Additionally, the proposed method showed greater concordance in gene selection across two studies of gene expression in acute myeloid leukemia than did the t-test or rank-sum test. Finally, we note that the principle of empirical Bayes assumption adequacy averaging can be used to develop robust procedures for a wide variety of experiments. [ABSTRACT FROM AUTHOR]

The article discusses a study on the transcriptome profiling of pediatric patients with acute myeloid leukemia (AML). The study investigated activating mutations of FLT3 in the patients and the outcome of xenotransplantation of primary AML cells. It examined RNA-seq expression values gained through positive quantile transformation (PQT) of a gene.

SAMD9 and SAMD9L germline mutations have recently emerged as a new class of predispositions to pediatric myeloid neoplasms. Patients commonly have impaired hematopoiesis, hypocellular marrows, and a greater risk of developing clonal chromosome 7 deletions leading to MDS and AML. We recently demonstrated that expressing SAMD9 or SAMD9L mutations in hematopoietic cells suppresses their proliferation and induces cell death. Here, we generated a mouse model that conditionally expresses mutant Samd9l to assess the in vivo impact on hematopoiesis. Using a range of in vivo and ex vivo assays, we showed that cells with heterozygous Samd9l mutations have impaired stemness relative to wild-type counterparts, which was exacerbated by inflammatory stimuli, and ultimately led to bone marrow hypocellularity. Genomic and phenotypic analyses recapitulated many of the hematopoietic cellular phenotypes observed in patients with SAMD9 or SAMD9L mutations, including lymphopenia, and pinpointed TGF-β as a potential targetable pathway. Further, we observed nonrandom genetic deletion of the mutant Samd9l locus on mouse chromosome 6, mimicking chromosome 7 deletions observed in patients. Collectively, our study has enhanced our understanding of mutant Samd9l hematopoietic phenotypes, emphasized the synergistic role of inflammation in exaggerating the associated hematopoietic defects, and provided insights into potential therapeutic options for patients. [ABSTRACT FROM AUTHOR]

BACKGROUND Gemtuzumab ozogamicin (GO) is an active agent for the treatment of CD33-postive acute myeloid leukemia (AML) and may improve the outcome of specific patient subgroups when combined with conventional chemotherapy. However, to the best of the authors' knowledge, the effects of GO on levels of minimal residual disease (MRD) are unknown. METHODS Pediatric patients with AML who received GO, either alone or in combination with chemotherapy on the AML02 multicenter trial, were analyzed to determine the effects of GO on MRD and outcome. RESULTS Among 17 patients who received GO alone because of persistent leukemia, 14 had a reduction in their MRD level and 13 became MRD negative. Of the 29 who received chemotherapy in combination with GO after responding poorly to chemotherapy, 28 demonstrated reduced MRD and 13 became MRD negative. Treatment with GO effectively reduced MRD before hematopoietic stem cell transplantation and was not found to be associated with increased treatment-related mortality after transplantation. CONCLUSIONS GO is effective in reducing MRD levels in pediatric patients with AML and may improve the outcome of those patients at high risk of disease recurrence. Cancer 2013;119:4036-4043. © 2013 American Cancer Society. [ABSTRACT FROM AUTHOR]

A whole-genome approach was used to investigate the genetic determinants of cytarabine-induced cytotoxicity. We periormed a meta-analysis of genome-wide association studies involving 523 lymphoblastoid cell lines (LCLs) from individuals of European, African, Asian, and African American ancestry. Several of the highest-ranked single-nucleotide polymorphisms (SNP5) were within the mutated in colorectal cancers (MCC) gene. MCC expression was induced by cytarabine treatment from 1.7-to 26.6-fold in LCL5. A total of 33 SNPs ranked at the top of the meta-analysis (P< 10-5) were successfully tested in a clinical trial of patients randomized to receive low-dose or high-dose cytarabine plus daunorubicin and etoposide; of these, 18 showed association (P < .05) with either cytarabine 50% inhibitory concentration in leukemia cells or clinical response parameters (minimal residual disease, overall survival (OS), and treatment-related mortality). This count (n = 18) was significantly greater than expected by chance (P = .016). For rs1203633, LCLs with AA genotype were more sensitive to cytarabine-induced cytotoxicity (P = 1.31 × 10-6) and AA (vs GA or GG) genotype was associated with poorer OS (P = .015), likely as a result of greater treatment-related mortality (P = .0037) in patients with acute myeloid leukemia (AML). This multicenter AMLD2 study trial was registered at www.clinicaltrials.gov as #NCTOO1 36084. [ABSTRACT FROM AUTHOR]

BACKGROUND: The effect of body mass index (BMI) on the treatment outcomes of children with acute myeloid leukemia (AML) is unclear and needs further evaluation. METHODS: Children with AML (n = 314) who were enrolled in 4 consecutive St. Jude protocols were grouped according to BMI (underweight, <5th percentile; healthy weight, 5th to 85th percentile; and overweight/obese, ≥85th percentile). RESULTS: Twenty-five patients (8%) were underweight, 86 patients (27.4%) were overweight/obese, and 203 patients (64.6%) had healthy weight. The 5-year overall survival rate of overweight/obese patients (46.5% ± 7.3%) was lower than the rate of patients with healthy weight (67.1% ± 4.3%; P < .001); underweight patients also tended to have lower survival rates (50.6% ± 10.7%; P = .18). In a multivariable analysis that was adjusted for age, leukocyte count, French-American-British classification, and study protocols, patients with healthy weight had the best survival rate among the 3 groups ( P = .01). When BMI was considered as continuous variable, patients with lower or higher BMI percentiles had worse survival ( P = .03). There was no difference in the occurrence of induction failure or relapse among BMI groups, although underweight and overweight/obese patients had a significantly higher cumulative incidence of treatment-related mortality, especially because of infection ( P = .009). CONCLUSIONS: An unhealthy BMI was associated with worse survival and more treatment-related mortality in children with AML. Meticulous supportive care with nutritional support and education, infection prophylaxis, and detailed laboratory and physical examination is required for these patients. These measures, together with pharmacokinetics-guided chemotherapy dosing, may improve outcome. Cancer 2012. © 2012 American Cancer Society. [ABSTRACT FROM AUTHOR]

BACKGROUND: Granulocyte-colony-stimulating factor (G-CSF) is effective in accelerating neutrophil recovery after intensive chemotherapy for acute myeloid leukemia (AML). However, the optimal G-CSF dosage for patients with AML has not been determined. To the authors' knowledge, G-CSF dosages have not been compared in a randomized AML study. METHODS: Patients who were enrolled on the St. Jude AML97 protocol and remained on study after window therapy were eligible to participate. The effect of the dosage of G-CSF given after induction chemotherapy Courses 1 and 2 was analyzed in 46 patients who were assigned randomly in a double-blinded manner to receive either 5 µg/kg daily or 10 µg/kg daily of G-CSF. The number of days of G-CSF treatment, neutropenia (an absolute neutrophil count <0.5 x 109/L), and hospitalization; the number of episodes of febrile neutropenia, grade 2 through 4 infection, and antimicrobial therapy; transfusion requirements; the cost of supportive care; and survival were compared between the 2 study arms. RESULTS: No statistically significant differences were observed between the 2 arms in any of the endpoints measured. CONCLUSIONS: The higher G-CSF dosage (10 µg/kg daily) offered no greater benefit than the lower dosage (5 µg/kg daily) in patients who were receiving intensive chemotherapy for AML. [ABSTRACT FROM AUTHOR]

The article reports on results of a study of clinical and biologic features and treatment outcome of children with newly diagnosed acute myeloid leukemia (AML) and hyperleukocytosis. A description of the experimental set-up and measurement method is presented. The study concluded that early mortality in patients with AML and hyperleukocytosis reduced with improved management, including supportive care.

Abstract: We describe a filtration-based culture method to enhance sensitivity of routine culture for recovery of fungi from respiratory specimens. The new method resulted in an 8.3% (P = 0.0039) increase in recovery for all organisms and a 6.4% (P = 0.0391) for Candida species when compared to the conventional culture technique. [Copyright &y& Elsevier]

BACKGROUNDInterpatient differences in the accumulation of methotrexate's active polyglutamylated metabolites (MTXPGs) in leukemia cells influence its antileukemic effects.METHODSTo identify genomic and epigenomic and patient variables determining the intracellular accumulation of MTXPGs, we measured intracellular MTXPG levels in acute lymphoblastic leukemia (ALL) cells from 388 newly diagnosed patients after in vivo high-dose methotrexate (HDMTX) (1 g/m2) treatment, defined ALL subtypes, and assessed genomic and epigenomic variants influencing folate pathway genes (mRNA, miRNA, copy number alterations [CNAs], SNPs, single nucleotide variants [SNVs], CpG methylation).RESULTSWe documented greater than 100-fold differences in MTXPG levels, which influenced its antileukemic effects (P = 4 × 10-5). Three ALL subtypes had lower MTXPG levels (T cell ALL [T-ALL] and B cell ALL [B-ALL] with the TCF3-PBX1 or ETV6-RUNX1 fusions), and 2 subtypes had higher MTXPG levels (hyperdiploid and BCR-ABL like). The folate pathway genes SLC19A1, ABCC1, ABCC4, FPGS, and MTHFD1 significantly influenced intracellular MTXPG levels (P = 2.9 × 10-3 to 3.7 × 10-8). A multivariable model including the ALL subtype (P = 1.1 × 10-14), the SLC19A1/(ABCC1 + ABCC4) transporter ratio (P = 3.6 × 10-4), the MTX infusion time (P = 1.5 × 10-3), FPGS mRNA expression (P = 2.1 × 10-3), and MTX systemic clearance (P = 4.4 × 10-2) explained 42% of the variation in MTXPG accumulation (P = 1.1 × 10-38). Model simulations indicated that a longer infusion time (24 h vs. 4 h) was superior in achieving higher intracellular MTXPG levels across all subtypes if ALL.CONCLUSIONSThese findings provide insights into mechanisms underlying interpatient differences in intracellular accumulation of MTXPG in leukemia cells and its antileukemic effectsFUNDINGTHE National Cancer Institute (NCI) and the Institute of General Medical Sciences of the NIH, the Basque Government Programa Posdoctoral de Perfeccionamiento de Personal Investigador doctor, and the American Lebanese Syrian Associated Charities (ALSAC). [ABSTRACT FROM AUTHOR]

Background: Outcomes for children with relapsed or refractory acute myeloid leukaemia remain poor. The BCL-2 inhibitor, venetoclax, has shown promising activity in combination with hypomethylating agents and low-dose cytarabine in older adults for whom chemotherapy is not suitable with newly diagnosed acute myeloid leukaemia. We aimed to determine the safety and explore the activity of venetoclax in combination with standard and high-dose chemotherapy in paediatric patients with relapsed or refractory acute myeloid leukaemia.Methods: We did a phase 1, dose-escalation study at three research hospitals in the USA. Eligible patients were aged 2-22 years with relapsed or refractory acute myeloid leukaemia or acute leukaemia of ambiguous lineage with adequate organ function and performance status. During dose escalation, participants received venetoclax orally once per day in continuous 28-day cycles at either 240 mg/m2 or 360 mg/m2, in combination with cytarabine received intravenously every 12 h at either 100 mg/m2 for 20 doses or 1000 mg/m2 for eight doses, with or without intravenous idarubicin (12 mg/m2) as a single dose, using a rolling-6 accrual strategy. The primary endpoint was the recommended phase 2 dose of venetoclax plus chemotherapy and the secondary endpoint was the proportion of patients treated at the recommended phase 2 dose who achieved complete remission or complete remission with incomplete haematological recovery. Analyses were done on patients who received combination therapy. The study is registered with ClinicalTrials.gov (NCT03194932) and is now enrolling to address secondary and exploratory objectives.Findings: Between July 1, 2017, and July 2, 2019, 38 patients were enrolled (aged 3-22 years; median 10 [IQR 7-13]), 36 of whom received combination therapy with dose escalation, with a median follow-up of 7·1 months (IQR 5·1-11·2). The recommended phase 2 dose of venetoclax was found to be 360 mg/m2 (maximum 600 mg) combined with cytarabine (1000 mg/m2 per dose for eight doses), with or without idarubicin (12 mg/m2 as a single dose). Overall responses were observed in 24 (69%) of the 35 patients who were evaluable after cycle 1. Among the 20 patients treated at the recommended phase 2 dose, 14 (70%, 95% CI 46-88) showed complete response with or without complete haematological recovery, and two (10%) showed partial response. The most common grade 3-4 adverse events were febrile neutropenia (22 [66%]), bloodstream infections (six [16%]), and invasive fungal infections (six [16%]). Treatment-related death occurred in one patient due to colitis and sepsis.Interpretation: The safety and activity of venetoclax plus chemotherapy in paediatric patients with heavily relapsed and refractory acute myeloid leukaemia suggests that this combination should be tested in newly diagnosed paediatric patients with high-risk acute myeloid leukaemia.Funding: US National Institutes of Health, American Lebanese Syrian Associated Charities, AbbVie, and Gateway for Cancer Research. [ABSTRACT FROM AUTHOR]

Consolidation therapies for children with intermediate- or high-risk acute myeloid leukemia (AML) are urgently needed to achieve higher cure rates while limiting therapy-related toxicities. We determined if adoptive transfer of natural killer (NK) cells from haploidentical killer immunoglobulin–like receptor (KIR)–human leukocyte antigen (HLA)-mismatched donors may prolong event-free survival in children with intermediate-risk AML who were in first complete remission after chemotherapy. Patients received cyclophosphamide (Day − 7), fludarabine (Days − 6 through − 2), and subcutaneous interleukin-2 (Days − 1, 1, 3, 5, 7, and 9). Purified, unmanipulated NK cells were infused on Day 0, and NK cell chimerism and phenotyping from peripheral blood were performed on Days 7, 14, 21, and 28. As primary endpoint, the event-free survival was compared to a cohort of 55 patients who completed chemotherapy and were in first complete remission but did not receive NK cells. Donor NK cell kinetics were determined as secondary endpoints. Twenty-one patients (median age at diagnosis, 6.0 years [range, 0.1–15.3 years]) received a median of 12.5 × 106 NK cells/kg (range, 3.6–62.2 × 106 cells/kg) without major side effects. All but 3 demonstrated transient engraftment with donor NK cells (median peak donor chimerism, 4% [range, 0–43%]). KIR–HLA-mismatched NK cells expanded in 17 patients (81%) and contracted in 4 (19%). However, adoptive transfer of NK cells did not decrease the cumulative incidence of relapse (0.393 [95% confidence interval: 0.182–0.599] vs. 0.35 [0.209–0.495]; P =.556) and did not improve event-free (60.7 ± 10.9% vs. 69.1 ± 6.8%; P =.553) or overall survival (84.2 ± 8.5% vs. 79.1 ± 6.6%; P =.663) over chemotherapy alone. The lack of benefit may result from insufficient numbers and limited persistence of alloreactive donor NK cells but does not preclude its potential usefulness during other phases of therapy, or in combination with other immunotherapeutic agents. Trial registration: www.clinicaltrials.gov, NCT00703820. Registered 24 June 2008. [ABSTRACT FROM AUTHOR]

PURPOSE: Pediatric adrenocortical carcinomas (ACCs) are aggressive; the overall survival of patients with ACCs is 40%-50%. Appropriate staging and histologic classification are crucial because children with incomplete resections, metastases, or relapsed disease have a dismal prognosis. The clinical course of pediatric adrenocortical tumors (ACTs) is difficult to predict using the current classification schemas, which rely on subjective microscopic and gross macroscopic variables. Recent advances in adult ACT studies have revealed distinct DNA methylation patterns with prognostic significance that have not been systematically interrogated in the pediatric population. PATIENTS AND METHODS: We performed DNA methylation analyses on 48 newly diagnosed ACTs from the International Pediatric Adrenocortical Tumor Registry and 12 pediatric adrenal controls to evaluate for distinct methylation groups. Pediatric methylation data were also compared systematically with the adult ACC cohort from The Cancer Genome Atlas (TCGA). RESULTS: Two pediatric ACT methylation groups were identified and showed differences in selected clinicopathologic and outcome characteristics. The A1 group was enriched for CTNNB1 variants and unfavorable outcome. The A2 group was enriched for TP53 germline variants, younger age at onset, and favorable outcome. Pediatric ACT methylation groups were maintained when International Pediatric Adrenocortical Tumor Registry cohort data were combined with TCGA cohort data. The CpG-island hypermethylator phenotype characterizing the TCGA cohort was not identified in the pediatric patients. When methylome findings were combined with independent histopathologic review using the Wieneke criteria, a high-risk population was identified with uniform fatal outcome. CONCLUSION: Our results indicate DNA methylation analysis can enhance current diagnostic algorithms. A combination of methylation and histologic classification produced the strongest prediction model and may prove useful in future risk-adapted therapeutic trials. [ABSTRACT FROM AUTHOR]

Multifocal synchronous or metachronous atypical teratoid rhabdoid tumors (ATRTs) and non-central nervous system malignant rhabdoid tumors (extra-CNS MRTs) are rare cancers. We reviewed the clinical and radiologic characteristics of affected patients seen at our institution. Genotyping and analysis of copy number abnormalities (CNAs) in SMARCB1 were performed in germline and tumor samples. Tumor samples underwent genome-wide DNA methylation and CNA analysis. The median age at diagnosis of 21 patients was 0.6 years. Two-thirds of ATRTs and extra-CNS MRTs were diagnosed synchronously. Although kidney tumors predominated, including two patients with bilateral involvement, at least 30% of cases lacked renal involvement. Histopathologic review confirmed MRTs in all cases and INI1 expression loss in all tumors tested. Fourteen (78%) of 18 patients tested had heterozygous germline SMARCB1 abnormalities. At least one allelic SMARCB1 abnormality was confirmed in 81 and 88% of ATRTs and extra-CNS MRTs, respectively. Unsupervised hierarchical clustering analysis of DNA methylation in 27 tumors and comparison with a reference group of 150 ATRTs classified the CNS tumors (n = 14) as sonic hedgehog (64%), tyrosinase (21%), and MYC (14%). The MYC subgroup accounted for 85% of 13 extra-CNS MRTs. Of 16 paired ATRTs and extra-CNS MRTs, the tumors in seven of eight patients showed a different pattern of genome-wide DNA methylation and/or CNAs suggestive of non-clonal origin. CNS and extra-CNS tumors had an identical SMARCB1 amplification (n = 1) or very similar DNA methylation pattern (n = 1) suggestive of clonal origin. All patients died of tumor progression. The clinical and molecular characteristics of multifocal ATRTs and extra-CNS MRTs are heterogeneous with most patients harboring a cancer predisposition. Although independent tumor origin was confirmed in most cases, metastatic spread was also documented. The recognition of their distinct molecular characteristics is critical in selecting new biologic therapies against these deadly cancers. [ABSTRACT FROM AUTHOR]

Evaluating the differential expression of a set of genes belonging to a common biological process or ontology has proven to be a very useful tool for biological discovery. However, existing gene-set association methods are limited to applications that evaluate differential expression across k ⩾ 2 treatment groups or biological categories. This limitation precludes researchers from most effectively evaluating the association with other phenotypes that may be more clinically meaningful, such as quantitative variables or censored survival time variables. Projection onto the Orthogonal Space Testing (POST) is proposed as a general procedure that can robustly evaluate the association of a gene-set with several different types of phenotypic data (categorical, ordinal, continuous, or censored). For each gene-set, POST transforms the gene profiles into a set of eigenvectors and then uses statistical modeling to compute a set of z-statistics that measure the association of each eigenvector with the phenotype. The overall gene-set statistic is the sum of squared z-statistics weighted by the corresponding eigenvalues. Finally, bootstrapping is used to compute a p -value. POST may evaluate associations with or without adjustment for covariates. In simulation studies, it is shown that the performance of POST in evaluating the association with a categorical phenotype is similar to or exceeds that of existing methods. In evaluating the association of 875 biological processes with the time to relapse of pediatric acute myeloid leukemia, POST identified the well-known oncogenic WNT signaling pathway as its top hit. These results indicate that POST can be a very useful tool for evaluating the association of a gene-set with a variety of different phenotypes. We have developed an R package named POST which is freely available in Bioconductor. [ABSTRACT FROM AUTHOR]

Bithalamic gliomas are rare cancers diagnosed based on poorly defined radiologic criteria. Infiltrative astrocytomas account for most cases. While some previous studies reported dismal outcomes for patients with bithalamic gliomas irrespective of therapy and histologic grade, others described better prognoses even without anticancer therapy. Little is known about their molecular characteristics. We reviewed clinical, radiologic, and histologic features of patients with bithalamic gliomas treated at our institution over 15 years. Targeted sequencing of mutational hotspots in H3F3A, HIST1H3B, IDH1/2, and BRAF, and genome-wide analysis of DNA methylation and copy number abnormalities was performed in available tumors. Eleven patients with bithalamic gliomas were identified. Their median age at diagnosis was 4.8 years (range: 1-15.7). Additional involvement of the brainstem, basal ganglia, and cerebral lobes occurred in 11, 9, and 3 cases, respectively. All patients presented with hydrocephalus. Two-thirds of the patients had a histologic diagnosis of anaplastic astrocytoma. Despite aggressive therapy, our youngest patient, the only one diagnosed before 1 year of age, is the sole long-term survivor. DNA methylation could be performed in seven tumors, all of which clustered with the RTK I 'PDGFRA' subgroup by unsupervised hierarchical analysis of methylation array against a previously published cohort of 59 pediatric high-grade gliomas. Sequencing of hotspots mutations could be done in 10 tumors, none of which harbored H3F3A p.K27 and/or the respective DNA methylation signature, and any other hotspot mutations. Amplification of MDM4 (n = 2), PDGFRA (n = 2), and ID2 combined with MYCN (n = 1) were observed in 7 tumors available for analysis. In comparison with the previously published experience with unilateral high-grade thalamic astrocytomas where H3F3A p.K27 was present in two-thirds of cases, the absence of this molecular subgroup in bithalamic gliomas was striking. This finding suggests that unilateral and bithalamic high-grade gliomas may represent two distinct molecular entities. [ABSTRACT FROM AUTHOR]

Oncogenic addiction to the Fms-like tyrosine kinase 3 (FLT3) is a hallmark of acute myeloid leukemia (AML) that harbors the FLT3-internal tandem duplication (FLT3-ITD) mutation. While FLT3 inhibitors like sorafenib show initial therapeutic efficacy, resistance rapidly develops through mechanisms that are incompletely understood. Here, we used RNA-Seq-based analysis of patient leukemic cells and found that upregulation of the Tec family kinase BMX occurs during sorafenib resistance. This upregulation was recapitulated in an in vivo murine FLT3-ITD-positive (FLT3-ITD+) model of sorafenib resistance. Mechanistically, the antiangiogenic effects of sorafenib led to increased bone marrow hypoxia, which contributed to HIF-dependent BMX upregulation. In in vitro experiments, hypoxia-dependent BMX upregulation was observed in both AML and non-AML cell lines. Functional studies in human FLT3-ITD+ cell lines showed that BMX is part of a compensatory signaling mechanism that promotes AML cell survival during FLT3 inhibition. Taken together, our results demonstrate that hypoxia-dependent upregulation of BMX contributes to therapeutic resistance through a compensatory prosurvival signaling mechanism. These results also reveal the role of off-target drug effects on tumor microenvironment and development of acquired drug resistance. We propose that the bone marrow niche can be altered by anticancer therapeutics, resulting in drug resistance through cell-nonautonomous microenvironment-dependent effects. [ABSTRACT FROM AUTHOR]

Background: Outcomes for children with acute myeloid leukemia (AML) have improved over the past 20 years even though the medications used for induction therapy have not changed.Methods: This study analyzed data from patients with AML who were enrolled in successive protocols (AML97 and AML02) to determine the contributors to the improved outcomes of the latter clinical trial.Results: There were significant improvements in 5-year overall survival (48.9% vs 71.2%; P < .0001) and event-free survival (43.5% vs 61.8%; P = .002) from AML97 to AML02. The 5-year cumulative incidence of early death (ED)/treatment-related mortality (TRM) was reduced for patients treated in AML02 (18.5% vs 7.9%; P = .007). Although the overall incidence of refractory disease (6.5% vs 5.6%; P = .736) and relapse (29.3% vs 21.0%; P = .12) did not differ between the 2 studies, patients with low-risk AML who were treated in AML02 had a reduced incidence of relapse (27.3% vs 8.8%; P = .036).Conclusions: The improved outcomes of the AML02 trial resulted from improved disease control for low-risk patients and overall decreased ED/TRM. These results emphasize the importance of supportive-care measures throughout chemotherapy courses and hematopoietic cell transplantation and the value of treatment intensity for patients with low-risk AML while underscoring the need for novel therapy, rather than increased therapy intensity, for children with high-risk AML. Cancer 2017;123:3791-3798. © 2017 American Cancer Society. [ABSTRACT FROM AUTHOR]

Background: The tumor protein p53 (TP53) arginine-to-histidine mutation at codon 337 (R337H) predisposes children to adrenocortical tumors (ACTs) and, rarely, to other childhood tumors, but its impact on adult cancer remains undetermined. The objective of this study was to investigate the frequency and types of cancer in relatives of children with ACT who carry the TP53 R337H mutation.Methods: TP53 R337H testing was offered to relatives of probands with ACT. The parental lineage segregating the R337H mutation was identified in all families. The frequency and distribution of cancer types were compared according to R337H status. The authors' data also were compared with those publicly available for children with TP53 mutations other than R337H.Results: The mean and median follow-up times for the probands with ACT were 11.2 years and 9.7 years (range, 3-32 years), respectively. During this time, cancer was diagnosed in 12 of 81 first-degree relatives (14.8%) carrying the R337H mutation but in only 1 of 94 noncarriers (1.1%; P = .0022). At age 45 years, the cumulative risk of cancer was 21% (95% confidence interval, 5%-33%) in carriers and 2% (95% confidence interval, 0%-4%) in noncarriers (P = .008). The frequency of cancer was higher in the R337H segregating lineages than in the nonsegregating lineages (249 of 1410 vs 66 of 984 individuals; P < .001). Breast and gastric cancer were the most common types.Conclusions: TP53 R337H carriers have a lifelong predisposition to cancer with a bimodal age distribution: 1 peak, represented by ACT, occurs in the first decade of life, and another peak of diverse cancer types occurs in the fifth decade. The current findings have implications for genetic counseling and surveillance of R337H carriers. Cancer 2017;123:3150-58. © 2017 American Cancer Society. [ABSTRACT FROM AUTHOR]

In the H AX1/HtrA2- O MI/ P ARL (HOP) mitochondrial protein complex, anti-apoptotic signals are generated by cleavage and activation of the serine protease HtrA2/OMI by the rhomboid protease PARL upon recruitment of both proteases to inner mitochondrial membrane protein HAX1 (HS1-associated protein X-1). Here we report the negative regulation of the HOP complex by human leukemia-associated myeloid leukemia factor 1 (MLF1). We demonstrate that MLF1 physically and functionally associates with HAX1 and HtrA2. Increased interaction of MLF1 with HAX1 and HtrA2 displaces HtrA2 from the HOP complex and inhibits HtrA2 cleavage and activation, resulting in the apoptotic cell death. Conversely, over-expressed HAX1 neutralizes MLF1's effect and inhibits MLF1-induced apoptosis. Importantly, Mlf1 deletion reverses B- and T-cell lymphopenia and significantly ameliorates the progressive striatal and cerebellar neurodegeneration observed in Hax1 −/− mice, with a doubling of the lifespan of Mlf1 −/− /Hax1 −/− animals compared to Hax1 −/− animals. Collectively, these data indicate that MLF1 serves as a proapoptotic antagonist that interacts with the HOP mitochondrial complex to modulate cell survival. [ABSTRACT FROM AUTHOR]

In many case-control designs of genome-wide association (GWAS) or next generation sequencing (NGS) studies, extensive data on secondary traits that may correlate and share the common genetic variants with the primary disease are available. Investigating these secondary traits can provide critical insights into the disease etiology or pathology, and enhance the GWAS or NGS results. Methods based on logistic regression (LG) were developed for this purpose. However, for the identification of rare variants (RVs), certain inadequacies in the LG models and algorithmic instability can cause severely inflated type I error, and significant loss of power, when the two traits are correlated and the RV is associated with the disease, especially at stringent significance levels. To address this issue, we propose a novel set-valued (SV) method that models a binary trait by dichotomization of an underlying continuous variable, and incorporate this into the genetic association model as a critical component. Extensive simulations and an analysis of seven secondary traits in a GWAS of benign ethnic neutropenia show that the SV method consistently controls type I error well at stringent significance levels, has larger power than the LG-based methods, and is robust in performance to effect pattern of the genetic variant (risk or protective), rare or common variants, rare or common diseases, and trait distributions. Because of the SV method's striking and profound advantage, we strongly recommend the SV method be employed instead of the LG-based methods for secondary traits analyses in case-control sequencing studies. [ABSTRACT FROM AUTHOR]

Background: As new technologies allow investigators to collect multiple forms of molecular data (genomic, epigenomic, transcriptomic, etc) and multiple endpoints on a clinical trial cohort, it will become necessary to effectively integrate all these data in a way that reliably identifies biologically important genes. Methods: We introduce CC-PROMISE as an integrated data analysis method that combines components of canonical correlation (CC) and projection onto the most interesting evidence (PROMISE). For each gene, CC-PROMISE first uses CC to compute scores that represent the association of two forms of molecular data with each other. Next, these scores are substituted into PROMISE to evaluate the statistical evidence that the molecular data show a biologically meaningful relationship with the endpoints. Results: CC-PROMISE shows outstanding performance in simulation studies and an example application involving pediatric leukemia. In simulation studies, CC-PROMISE controls the type I error (misleading significance) rate very near the nominal level across 100 distinct null settings in which no molecular-endpoint association exists. Also, CC-PROMISE has better statistical power than three other methods that control type I error in 396 of 400 (99 %) alternative settings for which a molecular-endpoint association is present; the power advantage of CC-PROMISE exceeds 30 % in 127 of the 400 (32 %) alternative settings. These advantages of CC-PROMISE are also observed in an example application. Conclusion: CC-PROMISE very effectively identifies genes for which some form of molecular data shows a biologically meaningful association with multiple related endpoints. Availability: The R package CCPROMISE is currently available from www.stjuderesearch.org/site/depts/biostats/ software. [ABSTRACT FROM AUTHOR]

Membrane-less organelles in cells are large, dynamic protein/protein or protein/ RNA assemblies that have been reported in some cases to have liquid droplet properties. However, the molecular interactions underlying the recruitment of components are not well understood. Herein, we study how the ability to form higher-order assemblies influences the recruitment of the speckle-type POZ protein ( SPOP) to nuclear speckles. SPOP, a cullin-3- RING ubiquitin ligase ( CRL3) substrate adaptor, self-associates into higher-order oligomers; that is, the number of monomers in an oligomer is broadly distributed and can be large. While wild-type SPOP localizes to liquid nuclear speckles, self-association-deficient SPOP mutants have a diffuse distribution in the nucleus. SPOP oligomerizes through its BTB and BACK domains. We show that BTB-mediated SPOP dimers form linear oligomers via BACK domain dimerization, and we determine the concentration-dependent populations of the resulting oligomeric species. Higher-order oligomerization of SPOP stimulates CRL3 SPOP ubiquitination efficiency for its physiological substrate Gli3, suggesting that nuclear speckles are hotspots of ubiquitination. Dynamic, higher-order protein self-association may be a general mechanism to concentrate functional components in membrane-less cellular bodies. [ABSTRACT FROM AUTHOR]

Gliomatosis cerebri (GC), a rare and deadly CNS neoplasm characterized by involvement of at least three cerebral lobes, predominantly affects adults. While a few small series have reported its occurrence in children, little is known about the molecular characteristics of pediatric GC. We reviewed clinical, radiological, and histological features of pediatric patients with primary GC treated at our institution over 15 years. Targeted sequencing of mutational hotspots in H3F3A, IDH1/2, and BRAF, and genome-wide analysis of DNA methylation and copy number abnormalities was performed in available tumors. Thirty-two patients [23 (72 %) with type 1 and 9 (28 %) with type 2 GC] were identified. Median age at diagnosis was 10.2 years (range 1.5-19.1). A median of 4 cerebral lobes (range 3-8) was affected at diagnosis. In addition, symmetrical bithalamic involvement was observed in 9 (28 %) patients. Twenty-two patients (69 %) had an anaplastic astrocytoma. Despite aggressive therapy, only two patients younger than 3 years at diagnosis are long-term survivors. Clustering analysis of methylation array data from 18 cases classified tumors as IDH ( n = 3, 17 %), G34 ( n = 4, 22 %), mesenchymal ( n = 3, 17 %), and RTK I 'PDGFRA' ( n = 8, 44 %). No tumors were classified as K27 subgroup. PDGFRA was the most commonly amplified oncogene in 4 of 22 tumors (18 %). H3F3A p.G34 occurred in all cases classified as G34. Two of 3 cases in the IDH subgroup had IDH1 p.R132H. No H3F3A p.K27 M, IDH2 p.R172, or BRAF p.V600E mutations were observed. There was a trend towards improved survival in the IDH subgroup ( P = 0.056). Patients with bithalamic involvement had worse outcomes ( P = 0.019). Despite some overlap, the molecular features of pediatric GC are distinct from its adult counterpart. Like in adults, the similarity of genetic and epigenetic characteristics with other infiltrative high-grade gliomas suggests that pediatric GC does not represent a distinct molecular entity. [ABSTRACT FROM AUTHOR]

In genetic association studies of an ordered categorical phenotype, it is usual to either regroup multiple categories of the phenotype into two categories and then apply the logistic regression (LG), or apply ordered logistic (oLG), or ordered probit (oPRB) regression, which accounts for the ordinal nature of the phenotype. However, they may lose statistical power or may not control type I error due to their model assumption and/or instable parameter estimation algorithm when the genetic variant is rare or sample size is limited. To solve this problem, we propose a set-valued (SV) system model to identify genetic variants associated with an ordinal categorical phenotype. We couple this model with a SV system identification algorithm to identify all the key system parameters. Simulations and two real data analyses show that SV and LG accurately controlled the Type I error rate even at a significance level of 10−6 but not oLG and oPRB in some cases. LG had significantly less power than the other three methods due to disregarding of the ordinal nature of the phenotype, and SV had similar or greater power than oLG and oPRB. We argue that SV should be employed in genetic association studies for ordered categorical phenotype. [ABSTRACT FROM AUTHOR]

We developed Copy Number Segmentation by Regression Tree in Next Generation Sequencing (CONSERTING), an algorithm for detecting somatic copy-number alteration (CNA) using whole-genome sequencing (WGS) data. CONSERTING performs iterative analysis of segmentation on the basis of changes in read depth and the detection of localized structural variations, with high accuracy and sensitivity. Analysis of 43 cancer genomes from both pediatric and adult patients revealed novel oncogenic CNAs, complex rearrangements and subclonal CNAs missed by alternative approaches. [ABSTRACT FROM AUTHOR]

Recently, there has been a great interest in the analysis of longitudinal data in which the observation process is related to the longitudinal process. In literature, the observation process was commonly regarded as a recurrent event process. Sometimes some observation duration may occur and this process is referred to as a recurrent episode process. The medical cost related to hospitalization is an example. We propose a conditional modeling approach that takes into account both informative observation process and observation duration. We conducted simulation studies to assess the performance of the method and applied it to a dataset of medical costs. [ABSTRACT FROM PUBLISHER]

BACKGROUND A better understanding of when cure can be declared in childhood acute myeloid leukemia (AML) would reduce anxiety and improve quality of life of AML survivors. The authors determined the likelihood that patients with AML would maintain long-term remission after the completion of therapy. METHODS The cumulative risk of relapse, the time to relapse, event-free survival, and overall survival were analyzed for 604 patients with AML who were enrolled in 7 successive clinical trials divided into 3 treatment eras (1976-1991, 1991-1997, and 2002-2008). RESULTS The median time to relapse did not change over time (0.93 years vs 0.76 years vs 0.8 years, respectively, for each consecutive era; P = .22), but the risk of relapse decreased significantly (5-year cumulative incidence of relapse: 52.6% ± 3.1% vs 31.5% ± 3.9% vs 22% ± 3%, respectively, for each consecutive era; P < .001). Among patients who were in remission 4 years from diagnosis, the probabilities of relapse were 1.7%, 2.9%, and 0.9%, respectively, for each consecutive era. In the most recent era, all but 1 of 44 relapses occurred within 4 years of diagnosis. CONCLUSIONS Children with AML who receive treatment with contemporary therapy and remain in remission 4 years from diagnosis probably are cured. Although late relapses and late deaths from other causes are rare, long-term follow-up of survivors is necessary for the timely management of late adverse effects. Cancer 2014;120:2490-2496. © 2014 American Cancer Society. [ABSTRACT FROM AUTHOR]

BACKGROUND Intensive chemotherapy for pediatric acute myeloid leukemia incurs the risk of infectious complications, but the benefits of antibiotic prophylaxis remain unclear. METHODS In the current study, among 103 children treated on the AML02 protocol between October 2002 and October 2008 at St. Jude Children's Research Hospital, the authors retrospectively assessed the effect of antibiotic prophylaxis on the frequency of febrile neutropenia, clinically or microbiologically confirmed infections (including bacteremia), and antibiotic resistance, as well as on the results of nasal and rectal surveillance cultures. Initially, patients received no prophylaxis or oral cephalosporin (group A). The protocol was then amended to administer intravenous cefepime alone or intravenous vancomycin plus either oral cephalosporin, oral ciprofloxacin, or intravenous cefepime (group B). RESULTS There were 334 infectious episodes. Patients in group A had a significantly greater frequency of documented infections and bacteremia (both P < .0001) (including gram-positive and gram-negative bacteremia; P = .0003 and .001, respectively) compared with patients in group B, especially viridans streptococcal bacteremia ( P = .001). The incidence of febrile neutropenia without documented infection was not found to be different between the 2 groups. Five cases of bacteremia with vancomycin-resistant enterococci (VRE) occurred in group B (vs none in group A), without related mortality. Two of these cases were preceded by positive VRE rectal surveillance cultures. CONCLUSIONS Outpatient intravenous antibiotic prophylaxis is feasible in children with acute myeloid leukemia and reduces the frequency of documented infection but not of febrile neutropenia. Despite the emergence of VRE bacteremia, the benefits favor antibiotic prophylaxis. Creative approaches to shorten the duration of prophylaxis and thereby minimize resistance should be explored. Cancer 2014;120:1985-1992. © 2014 American Cancer Society. [ABSTRACT FROM AUTHOR]

Pediatric high-grade glioma (HGG) is a devastating disease with a less than 20% survival rate 2 years after diagnosis. We analyzed 127 pediatric HGGs, including diffuse intrinsic pontine gliomas (DIPGs) and non-brainstem HGGs (NBS-HGGs), by whole-genome, whole-exome and/or transcriptome sequencing. We identified recurrent somatic mutations in ACVR1 exclusively in DIPGs (32%), in addition to previously reported frequent somatic mutations in histone H3 genes, TP53 and ATRX, in both DIPGs and NBS-HGGs. Structural variants generating fusion genes were found in 47% of DIPGs and NBS-HGGs, with recurrent fusions involving the neurotrophin receptor genes NTRK1, NTRK2 and NTRK3 in 40% of NBS-HGGs in infants. Mutations targeting receptor tyrosine kinase-RAS-PI3K signaling, histone modification or chromatin remodeling, and cell cycle regulation were found in 68%, 73% and 59% of pediatric HGGs, respectively, including in DIPGs and NBS-HGGs. This comprehensive analysis provides insights into the unique and shared pathways driving pediatric HGG within and outside the brainstem. [ABSTRACT FROM AUTHOR]

Background: Several analyte specific reagents (ASRs) are available for the detection and differentiation of HSV-1, HSV-2, and VZV in clinical specimens. However, there is limited data on the test performance of these reagents used in multiplexed PCR assays. Objective: This study compared the performance of two multiplexed ASR sets for detection of HSV-1, HSV-2, and VZV in dermal specimens. Study design: Two commercially available ASRs were combined to produce multiplexed PCR assays for simultaneous detection of HSV-1, HSV-2, and VZV. Seeded samples were used to determine the limit of detection (LOD) for each assay. Patient samples (n =156) were tested in duplicate and results for each method compared to the reference standard of culture. Both extracted and unextracted specimens were used in the study. Results: Both multiplexed PCR assays showed similar test performance, with minimal LOD differences observed. The LOD was 103 copies/mL for HSV-1 and HSV-2 using the Focus assay compared to 5×103 copies/mL and 2×104 copies/mL, respectively for the EraGen assay. Both assays showed equal performance for VZV with a LOD of 5×103 copies/mL. Analytical specificity testing showed no cross reactivity with other selected DNA viruses. Both assays showed similar performance when clinical samples were tested using both extracted and unextracted specimens. Conclusion: Commercially available ASRs can be successfully multiplexed for the PCR detection of HSV-1, HSV-2, and VZV using dermal specimens. Either direct testing or nucleic acid extracted specimens can be used with similar performance in these assays. [ABSTRACT FROM AUTHOR]

Motivation: There is a substantial body of works in the biology literature that seeks to characterize the cyclic behavior of genes during cell division. Gene expression microarrays made it possible to measure the expression profiles of thousands of genes simultaneously in time-course experiments to assess changes in the expression levels of genes over time. In this context, the commonly used procedures for testing include the permutation test by de Lichtenberg et al. and the Fisher’s G-test, both of which are designed to evaluate periodicity against noise. However, it is possible that a gene of interest may have expression that is neither cyclic nor just noise. Thus, there is a need for a new test for periodicity that can identify cyclic patterns against not only noise but also other non-cyclic patterns such as linear, quadratic or higher order polynomial patterns.Results: To address this weakness, we have introduced an empirical Bayes approach to test for periodicity and compare its performance in terms of sensitivity and specificity with that of the permutation test and Fisher’s G-test through extensive simulations and by application to a set of time-course experiments on the Schizosaccharomyces pombe cell-cycle gene expression. We use ‘conserved’ and ‘cycling’ genes by Lu et al. to assess the sensitivity and CESR genes by Chenet al. to assess the specificity of our new empirical Bayes method.Availability and implementation: The SAS Macro for our empirical Bayes test for periodicity is included in the supplementary materials along with a sample run of the MACRO program.Contact: mkocak1@uthsc.eduSupplementary information: Supplementary data are available at Bioinformatics online. [ABSTRACT FROM AUTHOR]

To identify genomic alterations in chronic lymphocytic leukemia (CLL), we performed single-nucleotide polymorphism-array analysis using Affymetrix Version 6,0 on 353 samples from untreated patients entered in the CLL8 treatment trial. Based on paired-sample analysis (n = 144), a mean of 1.8 copy number alterations per patient were identified; approximately 60% of patients carried no copy number alterations other than those detected by fluorescence in situ hybridization analysis. Copy-neutral loss-of-heterozygosity was detected in 6% of CLL patients and was lound most frequently on 13q, 17p, and 11q. Minimally deleted regions were refined on 13q14 (deleted in 61% of patients) to the DLEU1 and DLEU2 genes, on 11q22.3 (27% of patients) to ATM, on 2p16.1-2p15 (gained in 7% of patients) to a 1.9-Mb fragment containing 9 genes, and on 8q24.21 (5% of patients) to a segment 486 kb proximal to the MYC locus. 13q deletions exhibited proximal and distal breakpoint cluster regions. Among the most common novel lesions were deletions at 15q15.1 (4% of patients), with the smallest deletion (70.48 kb) found in the MGA locus. Sequence analysis of MGA in 59 samples revealed a truncating mutation in one CLL patient lacking a 15q deletion. MNTat 17p13.3, which in addition to MGA and MYC encodes for the network of MAX-interacting proteins, was also deleted recurrently. [ABSTRACT FROM AUTHOR]

Summary: To define the mutation spectrum in non-Down syndrome acute megakaryoblastic leukemia (non-DS-AMKL), we performed transcriptome sequencing on diagnostic blasts from 14 pediatric patients and validated our findings in a recurrency/validation cohort consisting of 34 pediatric and 28 adult AMKL samples. Our analysis identified a cryptic chromosome 16 inversion (inv(16)(p13.3q24.3)) in 27% of pediatric cases, which encodes a CBFA2T3-GLIS2 fusion protein. Expression of CBFA2T3-GLIS2 in Drosophila and murine hematopoietic cells induced bone morphogenic protein (BMP) signaling and resulted in a marked increase in the self-renewal capacity of hematopoietic progenitors. These data suggest that expression of CBFA2T3-GLIS2 directly contributes to leukemogenesis. [ABSTRACT FROM AUTHOR]

Retinoblastoma is a childhood cancer of the developing retina that begins in utero and is diagnosed in the first years of life. Biallelic RB1 gene inactivation is the initiating genetic lesion in retinoblastoma. The p53 gene is intact in human retinoblastoma but the pathway is believed to be suppressed by increased expression of MDM4 (MDMX) and MDM2. Here we quantify the expression of MDM4 and MDM2 mRNA and protein in human fetal retinae, primary retinoblastomas, retinoblastoma cell lines and several independent orthotopic retinoblastoma xenografts. We found that MDM4 is the major p53 antagonist expressed in retinoblastoma and in the developing human retina. We also discovered that MDM4 protein steady state levels are much higher in retinoblastoma than in human fetal retinae. This increase would not have been predicted based on the mRNA levels. We explored several possible post-transcriptional mechanisms that may contribute to the elevated levels of MDM4 protein. A proportion of MDM4 transcripts are alternatively spliced to produce protein products that are reported to be more stable and oncogenic. We also discovered that a microRNA predicted to target MDM4 (miR191) was downregulated in retinoblastoma relative to human fetal retinae and a subset of samples had somatic mutations that eliminated the miR-191 binding site in the MDM4 mRNA. Taken together, these data suggest that post-transcriptional mechanisms may contribute to stabilization of the MDM4 protein in retinoblastoma. [ABSTRACT FROM AUTHOR]

Summary: Medulloblastomas that display a large cell/anaplastic morphology and overexpress the cellular c-MYC gene are highly aggressive and carry a very poor prognosis. This so-called MYC-subgroup differs in its histopathology, gene expression profile, and clinical behavior from other forms of medulloblastoma. We generated a mouse model of MYC-subgroup medulloblastoma by transducing Trp53-null cerebellar progenitor cells with Myc. The cardinal features of these mouse medulloblastomas closely mimic those of human MYC-subgroup tumors and significantly differ from mouse models of the Sonic-Hedgehog- and WNT-disease subgroups. This mouse model should significantly accelerate understanding and treatment of the most aggressive form of medulloblastoma and infers distinct roles for MYC and MYCN in tumorigenesis. [Copyright &y& Elsevier]