DIAPH3 predicts survival of patients with MGMT-methylated glioblastoma.

Background: Glioblastoma is one of the most aggressive primary brain tumors, with a poor outcome despite multimodal treatment. Methylation of the MGMT promoter, which predicts the response to temozolomide, is a well-established prognostic marker for glioblastoma. However, a difference in survival can still be detected within the MGMT methylated group, with some patients exhibiting a shorter survival than others, emphasizing the need for additional predictive factors. Methods: We analyzed DIAPH3 expression in glioblastoma samples from the cancer genome atlas (TCGA). We also retrospectively analyzed one hundred seventeen histological glioblastomas from patients operated on at Saint-Luc University Hospital between May 2013 and August 2019. We analyzed the DIAPH3 expression, explored the relationship between mRNA levels and Patient's survival after the surgical resection. Finally, we assessed the methylation pattern of the DIAPH3 promoter using a targeted deep bisulfite sequencing approach. Results: We found that 36% and 1% of the TCGA glioblastoma samples exhibit copy number alterations and mutations in DIAPH3, respectively. We scrutinized the expression of DIAPH3 at single cell level and detected an overlap with MKI67 expression in glioblastoma proliferating cells, including neural progenitor-like, oligodendrocyte progenitor-like and astrocyte-like states. We quantitatively analyzed DIAPH3 expression in our cohort and uncovered a positive correlation between DIAPH3 mRNA level and patient's survival. The effect of DIAPH3 was prominent in MGMT-methylated glioblastoma. Finally, we report that the expression of DIAPH3 is at least partially regulated by the methylation of three CpG sites in the promoter region. Conclusion: We propose that combining the DIAPH3 expression with MGMT methylation could offer a better prediction of survival and more adapted postsurgical treatment for patients with MGMT-methylated glioblastoma.

Impact of SNP microarray analysis of compromised DNA on kinship classification success in the context of investigative genetic genealogy.

Single nucleotide polymorphism (SNP) data generated with microarray technologies have been used to solve murder cases via investigative leads obtained from identifying relatives of the unknown perpetrator included in accessible genomic databases, an approach referred to as investigative genetic genealogy (IGG). However, SNP microarrays were developed for relatively high input DNA quantity and quality, while DNA typically obtainable from crime scene stains is of low DNA quantity and quality, and SNP microarray data obtained from compromised DNA are largely missing. By applying the Illumina Global Screening Array (GSA) to 264 DNA samples with systematically altered quantity and quality, we empirically tested the impact of SNP microarray analysis of compromised DNA on kinship classification success, as relevant in IGG. Reference data from manufacturer-recommended input DNA quality and quantity were used to estimate genotype accuracy in the compromised DNA samples and for simulating data of different degree relatives. Although stepwise decrease of input DNA amount from 200 ng to 6.25 pg led to decreased SNP call rates and increased genotyping errors, kinship classification success did not decrease down to 250 pg for siblings and 1st cousins, 1 ng for 2nd cousins, while at 25 pg and below kinship classification success was zero. Stepwise decrease of input DNA quality via increased DNA fragmentation resulted in the decrease of genotyping accuracy as well as kinship classification success, which went down to zero at the average DNA fragment size of 150 base pairs. Combining decreased DNA quantity and quality in mock casework and skeletal samples further highlighted possibilities and limitations. Overall, GSA analysis achieved maximal kinship classification success from 800 to 200 times lower input DNA quantities than manufacturer-recommended, although DNA quality plays a key role too, while compromised DNA produced false negative kinship classifications rather than false positive ones.

Exploring Differentially Methylated Genes in Vulvar Squamous Cell Carcinoma.

DNA methylation is the most widely studied mechanism of epigenetic modification, which can influence gene expression without alterations in DNA sequences. Aberrations in DNA methylation are known to play a role in carcinogenesis, and methylation profiling has enabled the identification of biomarkers of potential clinical interest for several cancers. For vulvar squamous cell carcinoma (VSCC), however, methylation profiling remains an under-studied area. We sought to identify differentially methylated genes (DMGs) in VSCC, by performing Infinium MethylationEPIC BeadChip (Illumina) array sequencing, on a set of primary VSCC (n = 18), and normal vulvar tissue from women with no history of vulvar (pre)malignancies (n = 6). Using a false-discovery rate of 0.05, beta-difference (Δß) of ±0.5, and CpG-island probes as cut-offs, 199 DMGs (195 hyper-methylated, 4 hypo-methylated) were identified for VSCC. Most of the hyper-methylated genes were found to be involved in transcription regulator activity, indicating that disruption of this process plays a vital role in VSCC development. The majority of VSCCs harbored amplifications of chromosomes 3, 8, and 9. We identified a set of DMGs in this exploratory, hypothesis-generating study, which we hope will facilitate epigenetic profiling of VSCCs. Prognostic relevance of these DMGs deserves further exploration in larger cohorts of VSCC and its precursor lesions.

The role of epigenetic modifications in neurodevelopmental disorders: A systematic review.

Epigenetic processes have been suggested as key mechanisms in the etiology of neurodevelopmental disorders. This systematic review summarizes the current evidence for an association between epigenetics and Autism Spectrum Disorder (ASD) and Attention/Deficit-Hyperactivity Disorder (ADHD). Six databases were searched until the 24th of October 2017. Of the 2169 retrieved articles, 29 met our inclusion criteria. While generally associations between epigenetics and neurodevelopmental disorders were reported, only a few findings were consistent across independent analyses. Differential epigenetic markers were repeatedly identified in OR2L13, C11orf21/TSPAN32, PRRT1 and H3K27 for autism, and in VIPR2 for ADHD. Overall, evidence of an association between epigenetic modifications and ASD or ADHD should be considered preliminary and based on studies suffering from numerous caveats. We highlight the need for carefully designed investigations and for greater homogeneity and provide specific recommendations for future research. Despite the current limited understanding, the suggestive findings and rapid advances in the field hold the promise of a forthcoming elucidation of the role of epigenetic modifications in neurodevelopmental disorders.

Genome-wide methylation analysis identifies novel CpG loci for perimembranous ventricular septal defects in human.

AIM: Congenital heart diseases are the most common birth defects worldwide and leading cause of infant mortality. The perimembranous ventricular septal defect is most prevalent. Epigenetics may provide an underlying mechanism of the gene-environment interactions involved. MATERIALS & METHODS: We examined epigenome-wide DNA methylation using the Illumina HumanMethylation450 BeadChip in 84 case children and 196 control children. RESULTS: We identified differential methylation of a CpG locus (cg17001566) within the PRDM16 gene after Bonferroni correction (p = 9.17 × 10-8). This was validated by bisulfite pyrosequencing. PRDM16 functions as a repressor of TGF-ß signaling controlling tissue morphogenesis crucial during cardiogenesis. At 15% false-discovery rate, we identified seven additional CpG loci. CONCLUSION: These findings provide novel insights in the pathogenesis of perimembranous ventricular septal defect, which is of interest for future prediction and prevention.

Disease variants alter transcription factor levels and methylation of their binding sites.

Most disease-associated genetic variants are noncoding, making it challenging to design experiments to understand their functional consequences. Identification of expression quantitative trait loci (eQTLs) has been a powerful approach to infer the downstream effects of disease-associated variants, but most of these variants remain unexplained. The analysis of DNA methylation, a key component of the epigenome, offers highly complementary data on the regulatory potential of genomic regions. Here we show that disease-associated variants have widespread effects on DNA methylation in trans that likely reflect differential occupancy of trans binding sites by cis-regulated transcription factors. Using multiple omics data sets from 3,841 Dutch individuals, we identified 1,907 established trait-associated SNPs that affect the methylation levels of 10,141 different CpG sites in trans (false discovery rate (FDR) < 0.05). These included SNPs that affect both the expression of a nearby transcription factor (such as NFKB1, CTCF and NKX2-3) and methylation of its respective binding site across the genome. Trans methylation QTLs effectively expose the downstream effects of disease-associated variants.

The transcriptional landscape of age in human peripheral blood.

Disease incidences increase with age, but the molecular characteristics of ageing that lead to increased disease susceptibility remain inadequately understood. Here we perform a whole-blood gene expression meta-analysis in 14,983 individuals of European ancestry (including replication) and identify 1,497 genes that are differentially expressed with chronological age. The age-associated genes do not harbor more age-associated CpG-methylation sites than other genes, but are instead enriched for the presence of potentially functional CpG-methylation sites in enhancer and insulator regions that associate with both chronological age and gene expression levels. We further used the gene expression profiles to calculate the 'transcriptomic age' of an individual, and show that differences between transcriptomic age and chronological age are associated with biological features linked to ageing, such as blood pressure, cholesterol levels, fasting glucose, and body mass index. The transcriptomic prediction model adds biological relevance and complements existing epigenetic prediction models, and can be used by others to calculate transcriptomic age in external cohorts.

Prenatal parental tobacco smoking, gene specific DNA methylation, and newborns size: the Generation R study.

BACKGROUND: Deleterious effects of prenatal tobacco smoking on fetal growth and newborn weight are well-established. One of the proposed mechanisms underlying this relationship is alterations in epigenetic programming. We selected 506 newborns from a population-based prospective birth cohort in the Netherlands. Prenatal parental tobacco smoking was assessed using self-reporting questionnaires. Information on birth outcomes was obtained from medical records. The deoxyribonucleic acid (DNA) methylation of the growth genes IGF2DMR and H19 was measured in newborn umbilical cord white blood cells. Associations were assessed between parental tobacco smoking and DNA methylation using linear mixed models and adjusted for potential confounders. RESULTS: The DNA methylation levels of IGF2DMR and H19 in the non-smoking group were median (90 % range), 54.0 % (44.6-62.0), and 30.0 % (25.5-34.0), in the first trimester only smoking group 52.2 % (44.5-61.1) and 30.8 % (27.1-34.1), and in the continued smoking group 51.6 % (43.9-61.3) and 30.2 % (23.7-34.8), respectively. Continued prenatal maternal smoking was inversely associated with IGF2DMR methylation (ß = -1.03, 95 % CI -1.76; -0.30) in a dose-dependent manner (P-trend = 0.030). This association seemed to be slightly more profound among newborn girls (ß = -1.38, 95 % CI -2.63; -0.14) than boys (ß = -0.72, 95 % CI -1.68; 0.24). H19 methylation was also inversely associated continued smoking <5 cigarettes/day (ß = -0.96, 95 % CI -1.78; -0.14). Moreover, the association between maternal smoking and newborns small for gestational age seems to be partially explained by IGF2DMR methylation (ß = -0.095, 95 % CI -0.249; -0.018). Among non-smoking mothers, paternal tobacco smoking was not associated with IGF2DMR or H19 methylation. CONCLUSIONS: Maternal smoking is inversely associated with IGF2DMR methylation in newborns, which can be one of the underlying mechanisms through which smoking affects fetal growth.

Adverse life events and allele-specific methylation of the serotonin transporter gene (SLC6A4) in adolescents: the TRAILS study.

OBJECTIVES: Adverse life events increase vulnerability to affective disorders later in life, possibly mediated by methylation of the serotonin transporter gene (SLC6A4). We investigated the relationship of SLC6A4 methylation with various types of adversity (perinatal adversity, traumatic youth experiences and stressful life events [SLEs]), as well as with the timing of SLEs (during childhood [0-11 years] or during adolescence [12-15 years]). In addition, we investigated whether different serotonin-transporter-linked polymorphic region genotypes were equally sensitive to SLE-related methylation. METHODS: In a population sample of 939 adolescents (mean age = 16.2 years), we assessed SLC6A4 methylation, SLC6A4 functionality (serotonin-transporter-linked polymorphic region "long" and "short" alleles, and rs25531), and adverse life events. RESULTS: Only a higher number of SLEs was positively associated with higher SLC6A4 methylation (B = 0.11, p = .011). Adolescent SLEs were associated with higher SLC6A4 methylation (B = 0.13, p = .004) independently of childhood SLEs (B = 0.02, p = .57). L-allele homozygotes showed a greater impact of SLEs on methylation (B = 0.37, p < .001) than did s-allele carriers (B = 0.04, p = .66), resulting in higher levels of SLC6A4 methylation for l-allele homozygotes among those experiencing high levels of SLEs. CONCLUSIONS: Our findings demonstrate a higher level of SLC6A4 methylation after SLEs in adolescents, with a more pronounced association for SLEs during adolescence than during childhood. Considering the allele-specific sensitivity of SLC6A4 methylation to SLEs, this study may help clarify the role of SLC6A4 in the development of affective disorders.

Determinants of maternal pregnancy one-carbon metabolism and newborn human DNA methylation profiles.

Maternal one-carbon (1-C) metabolism provides methylgroups for fetal development and programing by DNA methylation as one of the underlying epigenetic mechanisms. We aimed to investigate maternal 1-C biomarkers, folic acid supplement use, and MTHFR C677T genotype as determinants of 1-C metabolism in early pregnancy in association with newborn DNA methylation levels of fetal growth and neurodevelopment candidate genes. The participants were 463 mother-child pairs of Dutch national origin from a large population-based birth cohort in Rotterdam, The Netherlands. In early pregnancy (median 13.0 weeks, 90% range 10.4-17.1), we assessed the maternal folate and homocysteine blood concentrations, folic acid supplement use, and the MTHFR C677T genotype in mothers and newborns. In newborns, DNA methylation was measured in umbilical cord blood white blood cells at 11 regions of the seven genes: NR3C1, DRD4, 5-HTT, IGF2DMR, H19, KCNQ1OT1, and MTHFR. The associations between the 1-C determinants and DNA methylation were examined using linear mixed models. An association was observed between maternal folate deficiency and lower newborn DNA methylation, which attenuated after adjustment for potential confounders. The maternal MTHFR TT genotype was significantly associated with lower DNA methylation. However, maternal homocysteine and folate concentrations, folic acid supplement use, and the MTHFR genotype in the newborn were not associated with newborn DNA methylation. The maternal MTHFR C677T genotype, as a determinant of folate status and 1-C metabolism, is associated with variations in the epigenome of a selection of genes in newborns. Research on the implications of these variations in methylation on gene expression and health is recommended.

Inhibin alpha-subunit (INHA) expression in adrenocortical cancer is linked to genetic and epigenetic INHA promoter variation.

Adrenocortical carcinoma (ACC) is a rare, but highly malignant tumor of unknown origin. Inhibin α-subunit (Inha) knockout mice develop ACCs following gonadectomy. In man, INHA expression varies widely within ACC tissues and its circulating peptide inhibin pro-αC has been described as a novel tumor marker for ACC. We investigated whether genetic and epigenetic changes of the INHA gene in human ACC cause loss or variation of INHA expression. To this end, analyses of INHA sequence, promoter methylation and mRNA expression were performed in human adrenocortical tissues. Serum inhibin pro-αC levels were also measured in ACC patients. INHA genetic analysis in 37 unique ACCs revealed 10 novel, heterozygous rare variants. Of the 3 coding bases affected, one variant was synonymous and two were missense variants: S72F and S184F. The minor allele of rs11893842 at -124 bp was observed at a low frequency (24%) in ACC samples and was associated with decreased INHA mRNA levels: 4.7±1.9 arbitrary units for AA, compared to 26±11 for AG/GG genotypes (P = 0.034). The methylation of four proximal INHA promoter CpGs was aberrantly increased in five ACCs (47.7±3.9%), compared to normal adrenals (18.4±0.6%, P = 0.0052), whereas the other 14 ACCs studied showed diminished promoter methylation (9.8±1.1%, P = 0.020). CpG methylation was inversely correlated to INHA mRNA levels in ACCs (r = -0.701, p = 0.0036), but not associated with serum inhibin pro-αC levels. In conclusion, aberrant methylation and common genetic variation in the INHA promoter occur in human ACCs and are associated with decreased INHA expression.

Methylation of migraine-related genes in different tissues of the rat.

17ß-Estradiol, an epigenetic modulator, is involved in the increased prevalence of migraine in women. Together with the prophylactic efficacy of valproate, which influences DNA methylation and histone modification, this points to the involvement of epigenetic mechanisms. Epigenetic studies are often performed on leukocytes, but it is unclear to what extent methylation is similar in other tissues. Therefore, we investigated methylation of migraine-related genes that might be epigenetically regulated (CGRP-ergic pathway, estrogen receptors, endothelial NOS, as well as MTHFR) in different migraine-related tissues and compared this to methylation in rat as well as human leukocytes. Further, we studied whether 17ß-estradiol has a prominent role in methylation of these genes. Female rats (n = 35) were ovariectomized or sham-operated and treated with 17ß-estradiol or placebo. DNA was isolated and methylation was assessed through bisulphite treatment and mass spectrometry. Human methylation data were obtained using the Illumina 450k genome-wide methylation array in 395 female subjects from a population-based cohort study. We showed that methylation of the Crcp, Calcrl, Esr1 and Nos3 genes is tissue-specific and that methylation in leukocytes was not correlated to that in other tissues. Interestingly, the interindividual variation in methylation differed considerably between genes and tissues. Furthermore we showed that methylation in human leukocytes was similar to that in rat leukocytes in our genes of interest, suggesting that rat may be a good model to study human DNA methylation in tissues that are difficult to obtain. In none of the genes a significant effect of estradiol treatment was observed.

DNA methylation profiles at birth and child ADHD symptoms.

Attention deficit/hyperactivity disorder (ADHD) is a common and highly heritable psychiatric disorder. In addition, early life environmental factors contribute to the occurrence of ADHD. Recently, DNA methylation has emerged as a mechanism potentially mediating genetic and environmental effects. Here, we investigated whether newborn DNA methylation patterns of selected candidate genes involved in psychiatric disorders or fetal growth are associated with ADHD symptoms in childhood. Participants were 426 children from a large population based cohort of Dutch national origin. Behavioral data were obtained at age 6 years with the Child Behavior Checklist. For the current study, 11 regions at 7 different genes were selected. DNA methylation levels of cord blood DNA were measured for the 11 regions combined and for each region separately. We examined the association between DNA methylation levels at different regions and ADHD symptoms with linear mixed models. DNA methylation levels were negatively associated with ADHD symptom score in the overall analysis of all 11 regions. This association was largely explained by associations of DRD4 and 5-HTT regions. Other candidate genes showed no association between DNA methylation levels and ADHD symptom score. Associations between DNA methylation levels and ADHD symptom score were attenuated by co-occurring Oppositional defiant disorder and total symptoms. Lower DNA methylation levels of the 7 genes assessed at birth, were associated with more ADHD symptoms of the child at 6 years of age. Further studies are needed to confirm our results and to investigate the possible underlying mechanism.

DNA methylation of IGF2DMR and H19 is associated with fetal and infant growth: the generation R study.

Changes in epigenetic programming of embryonic growth genes during pregnancy seem to affect fetal growth. Therefore, in a population-based prospective birth cohort in the Netherlands, we examined associations between fetal and infant growth and DNA methylation of IGF2DMR, H19 and MTHFR. For this study, we selected 69 case children born small-for-gestational age (SGA, birth weight <-2SDS) and 471 control children. Fetal growth was assessed with serial ultrasound measurements. Information on birth outcomes was retrieved from medical records. Infant weight was assessed at three and six months. Methylation was assessed in DNA extracted from umbilical cord white blood cells. Analyses were performed using linear mixed models with DNA methylation as dependent variable. The DNA methylation levels of IGF2DMR and H19 in the control group were, median (90% range), 53.6% (44.5-61.6) and 30.0% (25.6-34.2) and in the SGA group 52.0% (43.9-60.9) and 30.5% (23.9-32.9), respectively. The MTHFR region was found to be hypomethylated with limited variability in the control and SGA group, 2.5% (1.4-4.0) and 2.4% (1.5-3.8), respectively. SGA was associated with lower IGF2DMR DNA methylation (ß = -1.07, 95% CI -1.93; -0.21, P-value = 0.015), but not with H19 methylation. A weight gain in the first three months after birth was associated with lower IGF2DMR DNA methylation (ß = -0.53, 95% CI -0.91; -0.16, P-value = 0.005). Genetic variants in the IGF2/H19 locus were associated with IGF2DMR DNA methylation (P-value<0.05), but not with H19 methylation. Furthermore, our results suggest a possibility of mediation of DNA methylation in the association between the genetic variants and SGA. To conclude, IGF2DMR and H19 DNA methylation is associated with fetal and infant growth.

Epigenetic profiles in children with a neural tube defect; a case-control study in two populations.

Folate deficiency is implicated in the causation of neural tube defects (NTDs). The preventive effect of periconceptional folic acid supplement use is partially explained by the treatment of a deranged folate-dependent one carbon metabolism, which provides methyl groups for DNA-methylation as an epigenetic mechanism. Here, we hypothesize that variations in DNA-methylation of genes implicated in the development of NTDs and embryonic growth are part of the underlying mechanism. In 48 children with a neural tube defect and 62 controls from a Dutch case-control study and 34 children with a neural tube defect and 78 controls from a Texan case-control study, we measured the DNA-methylation levels of imprinted candidate genes (IGF2-DMR, H19, KCNQ1OT1) and non-imprinted genes (the LEKR/CCNL gene region associated with birth weight, and MTHFR and VANGL1 associated with NTD). We used the MassARRAY EpiTYPER assay from Sequenom for the assessment of DNA-methylation. Linear mixed model analysis was used to estimate associations between DNA-methylation levels of the genes and a neural tube defect. In the Dutch study group, but not in the Texan study group we found a significant association between the risk of having an NTD and DNA methylation levels of MTHFR (absolute decrease in methylation of -0.33% in cases, P-value = 0.001), and LEKR/CCNL (absolute increase in methylation: 1.36% in cases, P-value = 0.048), and a borderline significant association for VANGL (absolute increase in methylation: 0.17% in cases, P-value = 0.063). Only the association between MTHFR and NTD-risk remained significant after multiple testing correction. The associations in the Dutch study were not replicated in the Texan study. We conclude that the associations between NTDs and the methylation of the MTHFR gene, and maybe VANGL and LEKKR/CNNL, are in line with previous studies showing polymorphisms in the same genes in association with NTDs and embryonic development, respectively.

Tissue-specific DNA methylation profiles in newborns.

BACKGROUND: Epidemiological studies demonstrate that foetal growth restriction and low birth weight affect long-term health. Derangements in tissue-specific epigenetic programming of foetal and placental tissues are a suggested underlying mechanism of which DNA methylation is best understood. DNA methylation has been mostly investigated in DNA from white blood cells. To improve baseline understanding of tissue-specific DNA methylation, we examined variation in DNA methylation profiles of the imprinted foetal growth genes IGF2 and H19 in three different tissues from the same newborn obtained at the same time. FINDINGS: We obtained DNA from umbilical cord blood mononuclear cells (MNC; CD34+ and CD34-, n = 6), foetal side of the placenta (n = 5) and umbilical cord Wharton jelly (n = 5). DNA methylation of the IGF2 differentially methylated region (DMR) and H19 DMR was measured using quantitative mass spectrometry. Analysis of variance testing showed no statistical difference between total mean methylation of CD34+ and CD34- MNC. Further comparisons were made with the pooled total MNC fraction. Mean IGF2 DMR methylation of Wharton jelly was 1.3 times higher (P = 0.001) than mean methylation of the pooled MNC. Placental mean methylation was 0.8 times lower (P <0.001) and Wharton jelly 0.9 times lower (P <0.001) than the pooled MNC of H19 DMR. CONCLUSION: The total MNC fraction is a rather homogeneous cell population for methylation studies of imprinted genes in umbilical cord blood white blood cells, but may not always reflect the methylation levels of IGF2 and H19 in other organs.

A large-scale association analysis of 68 thyroid hormone pathway genes with serum TSH and FT4 levels.

OBJECTIVE: Minor variation in serum thyroid hormone (TH) levels can have important effects on various clinical endpoints. Although 45-65% of the inter-individual variation in serum TH levels is due to genetic factors, the causative genes are not well established. We therefore studied the effects of genetic variation in 68 TH pathway genes on serum TSH and free thyroxine (FT(4)) levels. DESIGN AND METHODS: Sixty-eight genes (1512 polymorphisms) were studied in relation to serum TSH and FT(4) levels in 1121 Caucasian subjects. Promising hits (P<0.01) were studied in three independent Caucasian populations (2656 subjects) for confirmation. A meta-analysis of all four studies was performed. RESULTS: For TSH, eight PDE8B polymorphisms (P=4×10(-17)) remained significant in the meta-analysis. For FT(4), two DIO1 (P=8×10(-12)) and one FOXE1 (P=0.0003) polymorphisms remained significant in the meta-analysis. Suggestive associations were detected for one FOXE1 (P=0.0028) and three THRB (P=0.0045) polymorphisms with TSH, and one SLC16A10 polymorphism (P=0.0110) with FT(4), but failed to reach the significant multiple-testing corrected P value (P<0.0022 and P<0.0033 respectively). CONCLUSIONS: Using a large-scale association analysis, we replicated previously reported associations with genetic variation in PDE8B, THRB, and DIO1. We demonstrate effects of genetic variation in FOXE1 on serum FT(4) levels, and borderline significant effects on serum TSH levels. A suggestive association of genetic variation in SLC16A10 with serum FT(4) levels was found. These data provide insight into the molecular basis of inter-individual variation in TH serum levels.

Loci at chromosomes 13, 19 and 20 influence age at natural menopause.

We conducted a genome-wide association study for age at natural menopause in 2,979 European women and identified six SNPs in three loci associated with age at natural menopause: chromosome 19q13.4 (rs1172822; -0.4 year per T allele (39%); P = 6.3 × 10(-11)), chromosome 20p12.3 (rs236114; +0.5 year per A allele (21%); P = 9.7 × 10(-11)) and chromosome 13q34 (rs7333181; +0.5 year per A allele (12%); P = 2.5 × 10(-8)). These common genetic variants regulate timing of ovarian aging, an important risk factor for breast cancer, osteoporosis and cardiovascular disease.

Regional deletion and amplification on chromosome 6 in a uveal melanoma case without abnormalities on chromosomes 1p, 3 and 8.

Uveal melanoma (UM) is the most common primary intraocular malignancy in adults. Loss of the long arm and gain of the short arm of chromosome 6 are frequently observed chromosomal aberrations in UM, together with loss of chromosome 1p36, loss of chromosome 3 and gain of chromosome 8. This suggests the presence of one or more oncogenes on 6p and tumor suppressor genes at 6q that are involved in UM development. Both regions, however, have not been well defined yet. Furthermore in other neoplasms gain of 6p and loss of 6q are frequently occurring events. In this case report, we describe the delineation of a partial gain on chromosome 6p and a partial deletion on 6q in a UM with the objective to pinpoint smaller candidate regions on chromosome 6 involved in UM development. Conventional cytogenetics, comparative genomic hybridization (CGH) and fluorescence in-situ hybridization (FISH) were used to delineate regions of loss and gain on chromosome 6 in this UM patient. With conventional cytogenetics a deleted region was found on chromosome 6q that was further delineated to a region ranging from 6q16.1 to 6q22 using CGH and FISH. A region of gain from 6pter to 6p21.2 was also demarcated with CGH and FISH. No other deletions or amplifications on recurrently involved chromosomes were found in this patient. This study indicates the presence of one or more tumor suppressor genes on chromosomal region 6q16.1-6q22 and the presence of one or more oncogenes on chromosomal region 6pter-6p21.2, which are likely to be important in UM and other tumors.

Expression of APITD1 is not related to copy number changes of chromosomal region 1p36 or the prognosis of uveal melanoma.

PURPOSE: Concurrent loss of chromosome arm 1p, region 36, and chromosome 3 leads to decreased disease-free survival in patients with uveal melanoma. A candidate tumor-suppressor gene APITD1 is located on the critical region on chromosome arm 1p, and it was therefore hypothesized that lower expression levels of this gene could lead to decreased survival in patients, with concurrent loss of a region on chromosome arm 1p and chromosome 3. Using neuroblastoma cells, which, like uveal melanoma, originate from neural crest cells, a former study showed that APITD1 has cell growth and/or cell death properties. In this study, APITD1 expression was analyzed to determine whether it corresponds with the DNA copy number and is related to survival in uveal melanoma. METHODS: To detect whether loss in the copy number of APITD1 results in lowered mRNA expression of the gene, FISH analysis was combined with real-time PCR. In addition, the effect of APITD1 expression on survival was studied by using Kaplan-Meier survival analysis. RESULTS: Expression of APITD1 mRNA was not related to DNA copy number (P = 0.956) or chromosome 3 status (P = 0.958). Kaplan-Meier survival analysis showed very similar survival curves for tumors with high and low APITD1 expression, with a log-rank significance value of P = 0.9682. CONCLUSIONS: These results indicate that APITD1 is not the tumor suppressor gene on 1p36 responsible for the negative prognostic effect in uveal melanoma with concurrent loss of chromosome arm 1p, region 36, and chromosome 3.