Mutations inhibiting KDM4B drive ALT activation in ATRX-mutated glioblastomas.

Alternative Lengthening of Telomeres (ALT) is a telomere maintenance pathway utilised in 15% of cancers. ALT cancers are strongly associated with inactivating mutations in ATRX; yet loss of ATRX alone is insufficient to trigger ALT, suggesting that additional cooperating factors are involved. We identify H3.3G34R and IDH1/2 mutations as two such factors in ATRX-mutated glioblastomas. Both mutations are capable of inactivating histone demethylases, and we identify KDM4B as the key demethylase inactivated in ALT. Mouse embryonic stem cells inactivated for ATRX, TP53, TERT and KDM4B (KDM4B knockout or H3.3G34R) show characteristic features of ALT. Conversely, KDM4B over-expression in ALT cancer cells abrogates ALT-associated features. In this work, we demonstrate that inactivation of KDM4B, through H3.3G34R or IDH1/2 mutations, acts in tandem with ATRX mutations to promote ALT in glioblastomas.

Factors associated with ambulatory care sensitive emergency department visits for South Carolina Medicaid members with intellectual disability.

INTRODUCTION: Ambulatory care sensitive conditions (ACSCs) can be seen as failure of access or management in primary care settings. Identifying factors associated with ACSCs for individuals with an Intellectual Disability (ID) provide insight into potential interventions. METHOD: To assess the association between emergency department (ED) ACSC visits and a number of demographic and health characteristics of South Carolina Medicaid members with ID. A retrospective cohort of adults with ID was followed from 2001 to 2011. Using ICD-9-CM codes, four ID subgroups, totalling 14 650 members, were studied. RESULTS: There were 106 919 ED visits, with 21 214 visits (19.8%) classified as ACSC. Of those, 82.9% were treated and released from EDs with costs averaging $578 per visit. People with mild and unspecified ID averaged greater than one ED visit per member year. Those with Down syndrome and other genetic cause ID had the lowest rates of ED visits but the highest percentage of ACSC ED visits that resulted in inpatient hospitalisation (26.6% vs. an average of 16.8% for other subgroups). When compared with other residential types, those residing at home with no health support services had the highest ED visit rate and were most likely to be discharged back to the community following an ED visit (85.2%). Adults residing in a nursing home had lower rates of ED visits but were most likely to be admitted to the hospital (38.9%) following an ED visit. Epilepsy and convulsions were the leading cause (29.6%) of ACSC ED visits across all subgroups and residential settings. CONCLUSION: Prevention of ACSC ED visits may be possible by targeting adults with ID who live at home without health support services.

Lineage-specific RUNX3 hypomethylation marks the preneoplastic immune component of gastric cancer.

Runt domain transcription factor 3 (RUNX3) is widely regarded as a tumour-suppressor gene inactivated by DNA hypermethylation of its canonical CpG (cytidine-phosphate-guanidine) island (CGI) promoter in gastric cancer (GC). Absence of RUNX3 expression from normal gastric epithelial cells (GECs), the progenitors to GC, coupled with frequent RUNX3 overexpression in GC progression, challenge this longstanding paradigm. However, epigenetic models to better describe RUNX3 deregulation in GC have not emerged. Here, we identify lineage-specific DNA methylation at an alternate, non-CGI promoter (P1) as a new mechanism of RUNX3 epigenetic control. In normal GECs, P1 was hypermethylated and repressed, whereas in immune lineages P1 was hypomethylated and widely expressed. In human GC development, we detected aberrant P1 hypomethylation signatures associated with the early inflammatory, preneoplastic and tumour stages. Aberrant P1 hypomethylation was fully recapitulated in mouse models of gastric inflammation and tumorigenesis. Cell sorting showed that P1 hypomethylation reflects altered cell-type composition of the gastric epithelium/tumour microenvironment caused by immune cell recruitment, not methylation loss. Finally, via long-term culture of gastric tumour epithelium, we revealed that de novo methylation of the RUNX3 canonical CGI promoter is a bystander effect of oncogenic immortalization and not likely causal in GC pathogenesis as previously argued. We propose a new model of RUNX3 epigenetic control in cancer, based on immune-specific, non-CGI promoter hypomethylation. This novel epigenetic signature may have utility in early detection of GC and possibly other epithelial cancers with premalignant immune involvement.

Use of state administrative data sources to study adolescents and young adults with rare conditions.

BACKGROUND: Effective care of young people with rare conditions requires ongoing coordinated medical treatment as well as educational and social support services. However, information on treatment is often lacking due to limited data. South Carolina has a repository of comprehensive health and human service data with which individuals may be tracked across the data systems of multiple state agencies and organizations. OBJECTIVE: To develop a method for studying health care of young persons with rare conditions using this repository. METHODS: We identified individuals aged 15 to 24 years diagnosed during 2000-2010 with Fragile X syndrome (FXS), spina bifida (SB), or muscular dystrophy (MD) using a series of algorithms. ICD-9-CM codes were used to initially identify the cohort from medical billing data. Demographics, medical care, employment, education, and socioeconomic status data were then extracted from linked administrative sources. RESULTS: We identified 1,040 individuals with these rare conditions: 125 with FXS, 695 with SB, and 220 with MD. The vast majority of the cases (95%) were identified in the Medicaid database. Half of the cohort was male, with a higher percentage in the FXS and MD groups. Sixty-two percent of the cohort was enrolled in the last year of high school. Over half of the cohort received support services from the state's disability and special-needs agency; 16% received food assistance. Thirty-eight percent were employed at some point during the study period. Forty-nine individuals with SB and 56 with MD died during the study period. CONCLUSIONS: We used a linked statewide data system to study rare conditions. Strengths include the diversity of information, rigorous identification strategies, and access to longitudinal data. Despite limitations inherent to administrative data, we found that linked state data systems are valuable resources for investigating important public health questions on rare conditions.

Pre-pregnancy body mass index, weight change during pregnancy, and risk of intellectual disability in children.

OBJECTIVE: This study investigated pre-pregnancy body mass index (BMI) and weight change in pregnancy as potential risk factors for intellectual disability (ID) in children. DESIGN: Retrospective cohort study. SETTING: South Carolina, USA. POPULATION: A total of 78,675 mother-child pairs, insured by the South Carolina Medicaid programme, born in the period 2004-2007. METHODS: We analysed South Carolina Medicaid data, linked to data from both the South Carolina Department of Education (DOE) and the South Carolina Department of Disabilities and Special Needs (DDSN). Maternal pre-pregnancy BMI and weight change during pregnancy were obtained from birth certificates. ID cases were identified from the three sources listed above. We used generalised estimating equation logistic regression models to model the odds of ID in children. MAIN OUTCOME MEASURES: Identified as having ID in special education, DDSN, or Medicaid billing records. RESULTS: The risk of ID was greater in children of women with pre-pregnancy obesity, and the risk was greatest in children born to women with morbid obesity (OR 1.52, 95% CI 1.30-1.77 for ID of any severity; OR 1.73, 95% CI 1.23-2.45 for severe ID). Gestational weight change (gain or loss) was not significantly associated with odds of ID. CONCLUSIONS: Pre-pregnancy obesity may be a modifiable risk factor for ID in children, although further study is needed to evaluate whether the association meets criteria for causation.

Religiosity, spirituality and antenatal anxiety in Southern U.S. women.

INTRODUCTION: We investigated the association between religiosity, spirituality, and anxiety in pregnant women, taking into account potential confounders. MATERIALS AND METHODS: From September 2005 through March 2006, pregnant women in three obstetrics practices in the American South were included in a cross-sectional study. The anxiety subscale of the Hospital Anxiety and Depression Scale (HADS) was used to measure anxiety. RESULTS: Of the 344 participating women, 23 screened positive for moderate to severe anxiety (HADS [anxiety] score greater than 10). Overall religiosity or spirituality (odds ratio [OR], 0.53; p=0.006) and social support (OR, 0.42; p<0.0001) were significantly associated with significantly lower odds of a positive anxiety screen. Among the specific measures of religiosity or spirituality, self-rated religiosity, self-rated spirituality, and participation in nonorganizational religious activities were significantly associated with lower odds of moderate to severe anxiety symptoms. DISCUSSION AND CONCLUSIONS: Religiosity and spirituality are associated with reduced anxiety in pregnant women. Additional study is needed to evaluate whether the association is causal.

Is postal follow-up of childhood cancer survivors worthwhile?

BACKGROUND: Childhood cancer survival has increased over the last 30 years, but long-term effects necessitate continued monitoring of survivors. Since not all of them attend follow-up clinics, this study assesses the efficacy of obtaining information from general practitioners (GPs) through a 5-year rolling postal program. PROCEDURE: Survivors were included who had been diagnosed with a malignancy in the West Midlands since 1957 and were not attending central long-term follow-up clinics. RESULTS: One thousand twenty-seven patients were followed up between 1993 and 2004. Replies were received on 903 (88% response). There were 44 subsequent malignancies and 42 deaths. No medical problems were reported in 341/935 patients (36.5%); in the other 594 endocrine effects were the most common, with visual effects the biggest single problem. Brain tumor survivors had the largest proportion of problems. CONCLUSIONS: The response rate and information quality achieved show that this method of follow-up is feasible, in cases of discharged or defaulting patients. These data will complement those derived from hospital-based follow-up studies, to give a broader understanding of the spectrum of late effects experienced by survivors and may inform the development of specific long-term follow-up protocols.

Down-regulation of ATM protein in HRS cells of nodular sclerosis Hodgkin's lymphoma in children occurs in the absence of ATM gene inactivation.

The tumour component of classical Hodgkin's lymphoma (cHL), Hodgkin Reed-Sternberg (HRS) cells, are believed to be derived from germinal centre (GC) B cells but intriguingly display a characteristic loss of B cell receptor (BCR) expression. The precise mechanisms by which BCR-negative HRS cell progenitors survive negative selection during the GC reaction remain obscure. Individuals with ataxia telangiectasia, caused by biallelic inactivation of the DNA damage response gene, ataxia telangiectasia mutated (ATM), have a higher risk of cHL development. Here we show that, in contrast to normal GC B cells that expressed low but detectable ATM protein, ATM protein was not detected in HRS cells of 17/18 cases of paediatric cHL, all but one with nodular sclerosis (NS) subtype. A comprehensive analysis of the ATM gene in microdissected HRS cells of nine representative tumours showed no evidence of either loss of heterozygosity or consistent pathogenic mutations. Furthermore, bisulphite sequencing of the ATM promoter from HRS cells of five tumours also revealed the absence of hypermethylation. Since our microarray data suggested significantly reduced ATM transcription in HRS cells compared to GC B cells, we conclude that loss of ATM expression could be the result of alterations in upstream regulators of ATM transcription. Importantly, ATM loss in paediatric cHLs has clinical implications and could be potentially exploited to guide future, less toxic, tumour-specific treatments.

Congenital abnormalities and clinical features associated with Wilms' tumour: a comprehensive study from a centre serving a large population.

Altogether 156 children treated for Wilms' tumour (WT) between 1970 and 1998 were studied. Sixty-six children, selected only by their attendance at clinic, were carefully examined and the findings compared to those from a case note review of 90 children. Congenital abnormalities were present in 45% of the examined cohort, in 19% of the case notes review group and in 30% overall. Novel findings included the association of WT with Marshall Smith syndrome, developmental delay in 3 of 4 cases of WT (one bilateral) and 1 sibling from consanguineous Pakistani families and another sibling also had leukaemia. The possibility of rare DNA repair or cancer predisposing disorders among these 4 families requires further study. Careful examination and history taking of an unselected patient cohort revealed a higher than expected incidence of clinical abnormalities which may be overlooked if not specifically sought.

Mutagenesis in mice of nuclear hormone receptor binding sites in the Igf2/H19 imprinting control region.

The H19/Igf2 imprinting control region (ICR) is a DNA methylation-dependent chromatin insulator in somatic cells. The hypomethylated maternally inherited ICR binds the insulator protein CTCF at four sites, and blocks activity of the proximal Igf2 promoter by insulating it from the shared distal enhancers. The hypermethylated paternally inherited ICR lacks CTCF binding and insulator activity, but induces methylation-silencing of the paternal H19 promoter. The paternal-specific methylation of the ICR is established in the male germ cells, while the ICR emerges from the female germ line in an unmethylated form. Despite several attempts to find cis-regulatory elements, it is still unknown what determines these male and female germ cell-specific epigenetic modifications. We recently proposed that five in vivo footprints spanning fifteen half nuclear hormone receptor (NHR) binding sites within the ICR might be involved, and here we report on the effects of mutagenizing all of these half sites in mice. No effect was obtained--in the female and male germ lines the mutant ICR remained hypomethylated and hypermethylated, respectively. The ICR imprinting mechanism remains undefined.

Caring for survivors of childhood cancers: the size of the problem.

Many survivors of childhood cancer have significant health problems due to their illness or treatment. This population-based study examines the number of long-term survivors, their disabilities and consequent long-term care needs. Survival rates for children diagnosed with cancer between 1960 and 1999 in the West Midlands, United Kingdom (UK), were used to estimate future long-term survivor numbers. Treatment and late effects data on a cohort of patients surviving for more than 5 years were used to consider continuing care needs. Between the 1960s and 1990s, 5-year survival increased from 23% to 70%. There were 98 5-year survivors in 1970, and numbers may exceed 2,100 by the end of 2005. Most (at least 61%) survivors in the West Midlands Region have one or more chronic medical problems and may require multidisciplinary care. We conclude that, in order to determine how to provide cost-effective care for this increasing population, protocol delivered management with audit is needed.

Seasonal differences in the onset of the EBV-positive and -negative forms of paediatric Hodgkin's lymphoma.

In this study, we have shown that there are seasonal differences in the onset of the (Epstein-Barr virus) EBV-positive and -negative forms of paediatric Hodgkin's lymphoma (HL). This suggests aetiological differences between the two forms of this disease. EBV-positive HL might be a rare consequence of primary EBV infection.

Sacrococcygeal teratomas: the UK Children's Cancer Study Group's experience. I. Neonatal.

The aim of this study was to review the United Kingdom Children's Cancer Study Group (UKCCSG) experience of sacrococcygeal teratomas (SCT) including histological presentation, response to surgery and chemotherapy, and long term effects of the tumour and treatment. This paper presents the results for those children diagnosed during the neonatal period. Children aged up to 4 weeks with biopsy proven localised or metastatic sacrococcygeal germ cell tumours were eligible. From 1st January 1989 to 31st December 1997 (9 years), 15 UKCCSG centres registered 51 neonates with SCT into GC 8901. Surgery alone was performed in all and the prognosis was good - except for 1 baby who died from massive haemorrhage at the initial operation and 1 who died from the complications of prematurity. Seven of the 51 children (14%) who had teratomas in the neonatal period (5 mature, two immature) had yolk sac tumour (YST) recurrence at: 4, 12, 15, 20, 20, 28 and 32 months of age. These children received chemotherapy in the form of etoposide/bleomycin/carboplatin (JEB) and are alive and well at review. These results emphasise the need for oncological follow-up of SCT and the good response to JEB chemotherapy of malignant teratomas and YST.

Molecular analysis of single colonies reveals a diverse origin of initial clonal proliferation in B-precursor acute lymphoblastic leukemia that can precede the t(12;21) translocation.

The pathogenesis of pediatric B-precursor acute lymphoblastic leukemia is largely unknown, and even with nonrandom chromosomal translocations present, the precise order of clonal molecular events is undefined. We developed an in vitro system using cytokines interleukin (IL)-3, IL-7, IL-10, and FMS-like tyrosine kinase 3 ligand with CD40 ligand-expressing fibroblasts to obtain single blast colonies from which clonal immunoglobulin heavy chain (IgH), T-cell receptor delta gene rearrangements, and, in t(12;21)-positive cases, TEL-AML1 fusion transcripts could be simultaneously PCR amplified. The proliferation of early tumor progenitors increased subclone detection enabling us, in seven diagnostic samples, to determine the stage of differentiation at which each leukemia occurred. Four were derived from the stage before initiation of IgH rearrangement, one during recombination of variable, joining, and diversity segments of the heavy chain gene VDJ(H), and two after completion of IgH rearrangement. Furthermore, analysis of a t(12;21)-positive leukemia with unusually late onset, identified both TEL-AML1-positive and -negative colonies carrying a clonal T-cell receptor delta rearrangement, inferring the presence of clonal expansion before the occurrence of the t(12;21). In contrast, in a typical, early onset t(12;21)-positive leukemia, the t(12;21) appeared to be the first clonal event. In both leukemias, the t(12;21) occurred before recombination of variable, joining and diversity segments of the heavy chain gene VDJ.

Neuroblastomas with chromosome 11q loss and single copy MYCN comprise a biologically distinct group of tumours with adverse prognosis.

Neuroblastoma is a heterogeneous tumour and its effective clinical management is dependent on accurate prognostic evaluation. In approximately 25% of patients amplification of the MYCN oncogene is known to be associated with a poor outcome. In order to identify additional molecular markers with prognostic potential in non-MYCN-amplified neuroblastomas, we looked for a correlation between clinical outcome and loss of heterozygosity (LOH) on four chromosomes that frequently show alteration in neuroblastoma (chromosomes 3, 4, 11 and 14). Chromosome 11q loss (with frequent parallel loss of chromosomes 3p, 4p and/or 14q) was found exclusively in tumours without MYCN amplification and was significantly associated with poor event-free survival. The 2-year event-free survival rate for 11q LOH cases was 30%, compared to 34% for MYCN-amplified cases and 100% for cases without these abnormalities. While 11q LOH was associated predominantly with advanced-stage disease, 2 cases with low-stage disease and 11q LOH both suffered relapses. We conclude that chromosome 11q loss defines a biologically distinct group of tumours without MYCN amplification that appear to have potential for aggressive metastatic growth. Thus this genetic alteration may be an important new prognostic marker in neuroblastoma.

South Asian ethnicity and material deprivation increase the risk of Epstein-Barr virus infection in childhood Hodgkin's disease.

In order to further define the factors associated with the observed variations in the Epstein-Barr virus-positive rate in childhood Hodgkin's disease, we have studied the effect of material deprivation (measured by the Townsend score) and ethnic origin on the frequency of Epstein-Barr virus-positivity in 55 cases of childhood Hodgkin's disease, diagnosed between 1981 and 1999, from a multi-ethnic region of the United Kingdom. Epstein-Barr virus status was determined by immunohistochemistry for the Epstein-Barr virus-encoded latent membrane protein-1. 62% of cases were Epstein-Barr virus-positive. Ethnic group was the strongest predictor of Epstein-Barr virus-positivity, with South Asians having a more than 20-fold risk of being Epstein-Barr virus-positive compared with non-South Asians. An increased risk was still present after adjusting for deprivation. Townsend scores were significantly higher (indicating more deprivation) in the Epstein-Barr virus-positive group, particularly in males. The relative risk of Epstein-Barr virus-positivity showed a gradient with increasing Townsend score; the risk being 7-times higher in the most deprived quartile compared with the least deprived group. Although the association between Townsend score and Epstein-Barr virus-positivity was reduced after adjusting for ethnic group, the risk of Epstein-Barr virus-positivity was still 3-times higher in the most deprived compared with the least deprived quartile. In addition, cases having 2 or more siblings were 5-times as likely to be Epstein-Barr virus-positive as those from smaller families. These results provide the first evidence of a strong association between Epstein-Barr virus-positive Hodgkin's disease and South Asian children from the United Kingdom. In addition, deprivation may increase the likelihood of Epstein-Barr virus-positive disease independently of ethnicity.

Imprinting in the germ line.

Genomic imprinting is an epigenetic system of gene regulation in mammals. It determines the parent-of-origin-dependent expression of a small number of imprinted genes during development, i.e., the maternal allele is inactive while the paternal is active, or vice versa. Imprinting is imparted in the germ line and involves differential DNA methylation such that particular DNA regions become methylated in one sex of germ line but not in the other. Inheritance of these differential egg and sperm methylation states is then transmitted to somatic cells, where they lead to differential maternal and paternal allelic activity, or monoallelic expression. Increasing evidence indicates that the inherited and stable differential allelic methylation regulates monoallelic expression by influencing the activity of gene regulatory elements-for one allele the element is switched off by methylation, while for the other the element is left potentially active by the lack of methylation. An interesting feature of the germ line is that, despite the presence of genomic imprinting, either as imprints inherited from the zygote or as new imprints imparted according to germ cell sex, imprinted genes are biallelically expressed as if imprints were not present. One explanation for this observation is that imprints have no influence over the germ cell's transcriptional machinery, i.e., imprinting may be neutralized in the germ cell lineage. This phenomenon may have a common basis with other unique features of the germ line, such as totipotency, perhaps in some unique aspect of chromatin structure.

A complex duplication created by gene targeting at the imprinted H19 locus results in two classes of methylation and correlated Igf2 expression phenotypes.

Imprinting of the mouse H19 and Igf2 genes is dependent on the presence of an intervening imprinting control region (ICR) situated 2 kb upstream of H19 and approximately 70 kb downstream of Igf2. Several recent studies have provided substantial evidence that the unmethylated maternal ICR acts as an insulator that prevents activation of Igf2 by a suite of enhancers downstream of the H19 gene. The methylated paternal ICR and H19 promoter have no activity, allowing sole activation of Igf2 expression. We have produced mice in which a duplication of the H19/Igf2 ICR produces, in each generation, two classes of methylation levels that correlated with two Igf2 imprinting phenotypes. One hypermethylated class also shows activation of the normally silent Igf2 gene, whereas the other hypomethylated class shows only slight activation of Igf2, in agreement with methylation's role in ICR function. This study describes a rare, possibly unique type of mutation that induces two distinct phenotypes in each generation.