Phosphorylation of GSK3α/ß correlates with activation of AKT and is prognostic for poor overall survival in acute myeloid leukemia patients.

BACKGROUND: Acute myeloid leukemia (AML) patients with highly active AKT tend to do poorly. Cell cycle arrest and apoptosis are tightly regulated by AKT via phosphorylation of GSK3α and ß isoforms which inactivates these kinases. In the current study we examine the prognostic role of AKT mediated GSK3 phosphorylation in AML. METHODS: We analyzed GSK3α/ß phosphorylation by reverse phase protein analysis (RPPA) in a cohort of 511 acute myeloid leukemia (AML) patients. Levels of phosphorylated GSK3 were correlated with patient characteristics including survival and with expression of other proteins important in AML cell survival. RESULTS: High levels of p-GSK3α/ß correlated with adverse overall survival and a lower incidence of complete remission duration in patients with intermediate cytogenetics, but not in those with unfavorable cytogenetics. Intermediate cytogenetic patients with FLT3 mutation also fared better respectively when p-GSK3α/ß levels were lower. Phosphorylated GSK3α/ß expression was compared and contrasted with that of 229 related cell cycle arrest and/or apoptosis proteins. Consistent with p-GSK3α/ß as an indicator of AKT activation, RPPA revealed that p-GSK3α/ß positively correlated with phosphorylation of AKT, BAD, and P70S6K, and negatively correlated with ß-catenin and FOXO3A. PKCδ also positively correlated with p-GSK3α/ß expression, suggesting crosstalk between the AKT and PKC signaling pathways in AML cells. CONCLUSIONS: These findings suggest that AKT-mediated phosphorylation of GSK3α/ß may be beneficial to AML cell survival, and hence detrimental to the overall survival of AML patients. Intrinsically, p-GSK3α/ß may serve as an important adverse prognostic factor for a subset of AML patients.

Corpus Callosum Area in Children and Adults with Autism.

Despite repeated findings of abnormal corpus callosum structure in autism, the developmental trajectories of corpus callosum growth in the disorder have not yet been reported. In this study, we examined corpus callosum size from a developmental perspective across a 30-year age range in a large cross-sectional sample of individuals with autism compared to a typically developing sample. Midsagittal corpus callosum area and the 7 Witelson subregions were examined in 68 males with autism (mean age 14.1 years; range 3-36 years) and 47 males with typical development (mean age 15.3 years; range 4-29 years). Controlling for total brain volume, increased variability in total corpus callosum area was found in autism. In autism, increased midsagittal areas were associated with reduced severity of autism behaviors, higher intelligence, and faster speed of processing (p=0.003, p=0.011, p=0.013, respectively). A trend toward group differences in isthmus development was found (p=0.029, uncorrected). These results suggest that individuals with autism benefit functionally from increased corpus callosum area. Our cross-sectional examination also shows potential maturational abnormalities in autism, a finding that should be examined further with longitudinal datasets.

Surface Adjustment of Reverse Phase Protein Arrays using Positive Control Spots.

Reverse phase protein arrays (RPPA) measure the relative expression levels of a protein in many samples simultaneously. Observed signal from these arrays is a combination of true signal, additive background, and multiplicative spatial effects. Background subtraction alone is not sufficient to remove all nonbiological trends from the data. We developed a surface adjustment that uses information from positive control spots to correct for spatial trends on the array beyond additive background. This method uses a generalized additive model to estimate a smoothed surface from positive controls. When positive controls are printed in a dilution series, a nested surface adjustment performs an intensity-based correction. When applicable, surface adjustment is able to remove spatial trends and increase within slide replicate agreement better than background subtraction alone as demonstrated on two sets of arrays. This work demonstrates the importance of including positive control spots on the array.

Ebf1 or Pax5 haploinsufficiency synergizes with STAT5 activation to initiate acute lymphoblastic leukemia.

As STAT5 is critical for the differentiation, proliferation, and survival of progenitor B cells, this transcription factor may play a role in acute lymphoblastic leukemia (ALL). Here, we show increased expression of activated signal transducer and activator of transcription 5 (STAT5), which is correlated with poor prognosis, in ALL patient cells. Mutations in EBF1 and PAX5, genes critical for B cell development have also been identified in human ALL. To determine whether mutations in Ebf1 or Pax5 synergize with STAT5 activation to induce ALL, we crossed mice expressing a constitutively active form of STAT5 (Stat5b-CA) with mice heterozygous for Ebf1 or Pax5. Haploinsufficiency of either Pax5 or Ebf1 synergized with Stat5b-CA to rapidly induce ALL in 100% of the mice. The leukemic cells displayed reduced expression of both Pax5 and Ebf1, but this had little effect on most EBF1 or PAX5 target genes. Only a subset of target genes was deregulated; this subset included a large percentage of potential tumor suppressor genes and oncogenes. Further, most of these genes appear to be jointly regulated by both EBF1 and PAX5. Our findings suggest a model whereby small perturbations in a self-reinforcing network of transcription factors critical for B cell development, specifically PAX5 and EBF1, cooperate with STAT5 activation to initiate ALL.

Cerebral volume loss, cognitive deficit, and neuropsychological performance: comparative measures of brain atrophy: II. Traumatic brain injury.

Traumatic brain injury (TBI) results in a variable degree of cerebral atrophy that is not always related to cognitive measures across studies. However, the use of different methods for examining atrophy may be a reason why differences exist. The purpose of this manuscript was to examine the predictive utility of seven magnetic resonance imaging (MRI)-derived brain volume or indices of atrophy for a large cohort of TBI patients (n = 65). The seven quantitative MRI (qMRI) measures included uncorrected whole brain volume, brain volume corrected by total intracranial volume, brain volume corrected by the ratio of the individual TICV by group TICV, a ventricle to brain ratio, total ventricular volume, ventricular volume corrected by TICV, and a direct measure of parenchymal volume loss. Results demonstrated that the various qMRI measures were highly interrelated and that corrected measures proved to be the most robust measures related to neuropsychological performance. Similar to an earlier study that examined cerebral atrophy in aging and dementia, these results suggest that a single corrected brain volume measure is all that is necessary in studies examining global MRI indicators of cerebral atrophy in relationship to cognitive function making additional measures of global atrophy redundant and unnecessary.

Postoperative radiotherapy use and patterns of care analysis for node positive or parametria positive cervical cancer.

OBJECTIVE: The objective of this study is to perform a patterns of care analysis for post-hysterectomy patients with node positive (LN+) or parametria positive cervical cancer. METHODS: A retrospective analysis was conducted utilizing the Surveillance, Epidemiology and End Results (SEER) Program from 1973 to 2006. We identified 2247 women with cervical carcinoma who had undergone hysterectomy and were found to have LN+ and/or positive parametria. RESULTS: Of the 2247 identified, 80.1% (1800) received postoperative radiotherapy (RT) while 19.9% (447) did not. Of those receiving RT, a significantly greater proportion had worse risk factors including higher clinical stage, and nodal bulk yet cause-specific survival (CSS) was equivalent between the two groups. RT utilization has increased over time from 67.2% in the cohort analyzed from 1973 to 1982 to 81.8% in the cohort analyzed from 2004 to 2006 (p = 0.0003). Blacks had worse CSS than whites (HR 1.35, 95% CI [1.05, 1.75]; p = 0.02). The proportion of those receiving RT was lower in blacks than whites (74.7% vs. 80.5%; p = 0.0358). From 1973 to 1982, 87.5% of blacks received RT while 62% of whites received RT (p = 0.0463). From 2004 to 2006, 64.4% of blacks received RT while 83.0% of whites received RT (p = 0.0024). CONCLUSIONS: Despite randomized data supporting the use of postoperative concurrent chemoradiotherapy for LN+ or parametria positive cervical cancer, the proportion of blacks not receiving RT is increasing over time. This is the largest patterns of care analysis to date of RT in patients with LN+ and/or parametria positive cervical cancer.

A population-based study of subsequent primary malignancies after endometrial cancer: genetic, environmental, and treatment-related associations.

PURPOSE: To examine the risk of subsequent primary malignancies (SPMs) in women diagnosed with endometrial cancer. METHODS AND MATERIALS: The National Cancer Institute's Survival, Epidemiology, and End Results database was used to determine the risk of SPM after endometrial cancer in 69,739 women diagnosed between 1973 and 2005. Standardized incidence ratios were calculated (observed/expected [O/E]) for SPM sites. RESULTS: Median follow-up was 11.2 years, representing 757,567 person-years of follow-up. The risk of SPM was significantly increased for small intestine (O/E = 1.48; 99% confidence interval [CI], 1.03-2.05), colon (O/E = 1.16; CI, 1.09-1.24), vagina (O/E = 2.71; CI, 1.86-3.8), and urinary bladder (O/E = 1.41; CI, 1.25-1.59) SPMs and decreased for oral cavity and pharynx (O/E = 0.75; CI, 0.6-0.93), lung and bronchus (O/E = 0.78; CI, 0.72-0.84), and esophagus (O/E = 0.58; CI, 0.37-0.86) SPMs. Patients receiving external-beam radiotherapy for endometrial cancer had an increased risk of colon (p < 0.001), bladder (p < 0.001), vagina (p = 0.04), and soft-tissue (p = 0.014) SPMs. Patients treated with brachytherapy had an increased risk of bladder SPM (p = 0.006). A positive bidirectional association with endometrial cancer was observed for colorectal cancer, with a negative bidirectional association for oropharyngeal and lung cancers. CONCLUSIONS: Genetic, environmental, and treatment-related factors influence SPM risk. Genetic factors may contribute to the increased risk of colorectal cancer. Smoking's negative effect on endometrial cancer risk factors might explain the decreased risk of lung and oropharyngeal cancer. Patients treated with radiotherapy likely have a small but significantly increased risk of bladder, vagina, colon, and soft-tissue SPM.

Variable slope normalization of reverse phase protein arrays.

MOTIVATION: Reverse phase protein arrays (RPPA) measure the relative expression levels of a protein in many samples simultaneously. A set of identically spotted arrays can be used to measure the levels of more than one protein. Protein expression within each sample on an array is estimated by borrowing strength across all the samples, but using only within array information. When comparing across slides, it is essential to account for sample loading, the total amount of protein printed per sample. Currently, total protein is estimated using either a housekeeping protein or the sample median across all slides. When the variability in sample loading is large, these methods are suboptimal because they do not account for the fact that the protein expression for each slide is estimated separately. RESULTS: We propose a new normalization method for RPPA data, called variable slope (VS) normalization, that takes into account that quantification of RPPA slides is performed separately. This method is better able to remove loading bias and recover true correlation structures between proteins. AVAILABILITY: Code to implement the method in the statistical package R and anonymized data are available at (http://bioinformatics.mdanderson.org/supplements.html).

Overexpression of ZNF342 by juxtaposition with MPO promoter/enhancer in the novel translocation t(17;19)(q23;q13.32) in pediatric acute myeloid leukemia and analysis of ZNF342 expression in leukemia.

We report a novel translocation t(17;19)(q22;q13.32) found in 100% of blast cells from a pediatric acute myeloid leukemia (AML) patient. Fluorescence in situ hybridization and vectorette polymerase chain reaction were used to precisely map the chromosomal breakpoint located on the derivative chromosome 17 at 352 bp 5' of MPO, encoding myeloperoxidase a highly expressed protein in myeloid cells, and 2,085 bp 5' of ZNF342 on 19q, encoding a transcription factor expressed in human stem cells and previously implicated in mouse models of leukemia. Analysis of RNA levels from the patient sample revealed significant overexpression of ZNF342, potentially contributing to AML formation. This is the first report of a translocation in myeloid leukemia occurring only in the promoter/enhancer regions of the two genes involved, similar to translocations commonly found in lymphoid malignancies. Analysis of ZNF342 protein levels in a large dataset of leukemia samples by reverse phase protein array showed that higher levels of ZNF342 expression in acute lymphoblastic leukemia was associated with poorer outcome (P = 0.033). In the myeloid leukemia samples with the highest ZNF342 expression, there was overrepresentation of FLT3 internal tandem duplication (P = 0.0016) and AML subtype M7 (P = 0.0002). Thus, overexpression of ZNF342 by translocation or other mechanisms contributes to leukemia biology in multiple hematopoietic compartments.

Superior temporal gyrus, language function, and autism.

Deficits in language are a core feature of autism. The superior temporal gyrus (STG) is involved in auditory processing, including language, but also has been implicated as a critical structure in social cognition. It was hypothesized that subjects with autism would display different size-function relationships between the STG and intellectual-language-based abilities when compared to controls. Intellectual ability was assessed by either the Wechsler Intelligence Scale for Children-Third Edition (WISC-III) or Wechsler Adult Intelligence Scale-Third Edition (WAIS-III), where three intellectual quotients (IQ) were computed: verbal (VIQ), performance (PIQ), and full-scale (FSIQ). Language ability was assessed by the Clinical Evaluation of Language Fundamentals-Third Edition (CELF-3), also divided into three index scores: receptive, expressive, and total. Seven to 19-year-old rigorously diagnosed subjects with autism (n = 30) were compared to controls (n = 39; 13 of whom had a deficit in reading) of similar age who were matched on education, PIQ, and head circumference. STG volumes were computed based on 1.5 Tesla magnetic resonance imaging (MRI). IQ and CELF-3 performance were highly interrelated regardless of whether subjects had autism or were controls. Both IQ and CELF-3 ability were positively correlated with STG in controls, but a different pattern was observed in subjects with autism. In controls, left STG gray matter was significantly (r = .42, p < or = .05) related to receptive language on the CELF-3; in contrast, a zero order correlation was found with autism. When plotted by age, potential differences in growth trajectories related to language development associated with STG were observed between controls and those subjects with autism. Taken together, these findings suggest a possible failure in left hemisphere lateralization of language function involving the STG in autism.

Quantitative temporal lobe differences: autism distinguished from controls using classification and regression tree analysis.

The temporal lobe is thought to be abnormal in autism, yet standard volumetric analyses are often unrevealing when age, sex, IQ, and head size are controlled. Quantification of temporal lobe structures were obtained in male subjects with autism and controls, where subjects with head circumference (HC) defined macrocephaly were excluded, so that volume differences were not just related to the higher prevalence of macrocephaly in autism. Various statistical methods were applied to the analysis including a classification and regression tree (CART) method, a non-parametric technique that helps define patterns of relationships that may be meaningful in distinguishing temporal lobe differences between subjects with autism and age and IQ matched controls. Subjects with autism were also compared to a separate control group with reading disorder (RD), with the prediction that the temporal lobe morphometric analysis of the reading disorder controls would be more similar to that of the autism group. The CART method yielded a high specificity in classifying autism subjects from controls based on the relationship between the volume of the left fusiform gyrus (LFG) gray and white matter, the right temporal stem (RTS) and the right inferior temporal gyrus gray matter (RITG-GM). Reading disordered individuals were more similar to subjects with autism. Simple size differences did not distinguish the groups. These findings demonstrate different relationships within temporal lobe structures that distinguish subjects with autism from controls. Results are discussed in terms of pathological connectivity within the temporal lobe as it relates to autism.

Cerebral volume loss, cognitive deficit and neuropsychological performance: comparative measures of brain atrophy: I. Dementia.

There are several magnetic resonance (MR) imaging methods to measure brain volume and cerebral atrophy; however, the best measure for examining potential relationships between such measures and neuropsychological performance has not been established. Relationships between seven measures of MR derived brain volume or indices of atrophy and neuropsychological performance in the elderly subjects of the population-based Cache County, Utah Study of Aging and Memory (n = 195) were evaluated. The seven MR measures included uncorrected total brain volume (TBV), TBV corrected by total intracranial volume (TICV), TBV corrected by the ratio of the individuals TICV by group TICV (TBVC), a ventricle-to-brain ratio (VBR), total ventricular volume (TVV), TVV corrected by TICV, and a measure of parenchymal volume loss. The cases from the Cache County Study were comprised of elderly individuals classified into one of four subject groups based on a consensus diagnostic process, independent of quantitative MR imaging findings. The groups included subjects with Alzheimer's disease (AD, n = 85), no dementia but mild/ambiguous (M/A) deficits (n = 30), a group of subjects with non-AD dementia or neuropsychiatric disorder including vascular dementia (n = 60), and control subjects (n = 20). Neuropsychological performance was based on the Mini-Mental Status Exam (MMSE) and an expanded neuropsychological test battery (consortium to establish a registry for Alzheimer's disease (CERAD). The results demonstrated that the various quantitative MR measures were highly interrelated and no single measure was statistically superior. However, TBVC, TBV/TICV and VBR consistently exhibited the more robust relationships with neuropsychological performance. These results suggest that a single corrected brain volume measure or index is sufficient in studies examining global MR indicators of cerebral atrophy in relation to cognitive function and recommends use of either TBVC, TBV/TICV, or VBR.

Temporal lobe, autism, and macrocephaly.

BACKGROUND AND PURPOSE: Because of increased prevalence of macrocephaly in autism, head size must be controlled for in studies that examine volumetric findings of the temporal lobe in autistic subjects. We prospectively examined temporal lobe structures in individuals with autism who were normocephalic or macrocephalic (head circumference > 97th percentile) and in control subjects who were normocephalic or macrocephalic or who had a reading disorder (unselected for head size). The rationale for the reading disorder group was to have control subjects with potential temporal lobe anomalies, but who were not autistic. METHODS: In individuals aged 7-31 years, autism was diagnosed on the basis of standardized interview and diagnostic criteria. Control subjects ranged in age from 7 to 22 years. All subjects were male. MR morphometrics of the major temporal lobe structures were based on ANALYZE segmentation routines, in which total brain volume and total intracranial volume (TICV) were calculated. Both group comparisons and developmental analyses were performed. RESULTS: No distinct temporal lobe abnormalities of volume were observed once head size (TICV) was controlled for. In autistic and control subjects, robust growth patterns were observed in white and gray matter that differed little between the groups. Although subtle differences were observed in some structures (ie, less white matter volume in the region of the temporal stem and overall temporal lobe), none was statistically significant. CONCLUSION: No major volumetric anomalies of the temporal lobe were found in cases of autism when IQ, TICV, and age were controlled. Temporal lobe abnormalities that may be associated with autism are likely to be more related to functional organization within the temporal lobe than to any gross volumetric difference.