Ricardo Segurado

Combined Analysis from Eleven Linkage Studies of Bipolar Disorder Provides Strong Evidence of Susceptibility Loci on Chromosomes 6q and 8q

Several independent studies and meta-analyses aimed at identifying genomic regions linked to bipolar disorder (BP) have failed to find clear and consistent evidence of linkage regions. Our hypothesis is that combining the original genotype data provides benefits of increased power and control over sources of heterogeneity that outweigh the difficulty and potential pitfalls of the implementation. We conducted a combined analysis using the original genotype data from 11 BP genomewide linkage scans comprising 5,179 individuals from 1,067 families. Heterogeneity among studies was minimized in our analyses by using uniform methods of analysis and a common, standardized marker map and was assessed using novel methods developed for meta-analysis of genome scans. To date, this collaboration is the largest and most comprehensive analysis of linkage samples involving a psychiatric disorder. We demonstrate that combining original genome-scan data is a powerful approach for the elucidation of linkage regions underlying complex disease. Our results establish genomewide significant linkage to BP on chromosomes 6q and 8q, which provides solid information to guide future gene-finding efforts that rely on fine-mapping and association approaches.

Genome Scan Meta-Analysis of Schizophrenia and Bipolar Disorder, Part I: Methods and Power Analysis

This is the first of three articles on a meta-analysis of genome scans of schizophrenia (SCZ) and bipolar disorder (BPD) that uses the rank-based genome scan meta-analysis (GSMA) method. Here we used simulation to determine the power of GSMA to detect linkage and to identify thresholds of significance. We simulated replicates resembling the SCZ data set (20 scans; 1,208 pedigrees) and two BPD data sets using very narrow (9 scans; 347 pedigrees) and narrow (14 scans; 512 pedigrees) diagnoses. Samples were approximated by sets of affected sibling pairs with incomplete parental data. Genotypes were simulated and nonparametric linkage (NPL) scores computed for 20 180-cM chromosomes, each containing six 30-cM bins, with three markers/bin (or two, for some scans). Genomes contained 0, 1, 5, or 10 linked loci, and we assumed relative risk to siblings (lambda(sibs)) values of 1.15, 1.2, 1.3, or 1.4. For each replicate, bins were ranked within-study by maximum NPL scores, and the ranks were averaged (R(avg)) across scans. Analyses were repeated with weighted ranks ((sqrt)N[genotyped cases] for each scan). Two P values were determined for each R(avg): P(AvgRnk) (the pointwise probability) and P(ord) (the probability, given the bins place in the order of average ranks). GSMA detected linkage with power comparable to or greater than the underlying NPL scores. Weighting for sample size increased power. When no genomewide significant P values were observed, the presence of linkage could be inferred from the number of bins with nominally significant P(AvgRnk), P(ord), or (most powerfully) both. The results suggest that GSMA can detect linkage across multiple genome scans.

The SNP ratio test

UNLABELLED We present a tool that assesses the enrichment of significant associations from genome-wide association studies (GWAS) in a pathway context. The SNP ratio test (SRT) compares the proportion of significant to all SNPs within genes that are part of a pathway and computes an empirical P-value based on comparisons to ratios in datasets where the assignment of case/control status has been randomized. We applied the SRT to a Parkinsons disease GWAS dataset, using the KEGG database, revealing significance for Parkinsons disease and related pathways. AVAILABILITY https://sourceforge.net/projects/snpratiotest/

The Wellcome trust UK-Irish bipolar affective disorder sibling-pair genome screen: first stage report.

We have completed the first stage of a two-stage genome wide screen designed to identify chromosomal regions that may harbour susceptibility genes for bipolar affective disorder. The first stage screening sample included 509 subjects from 151 nuclear families recruited within the United Kingdom and Republic of Ireland. This sample contained 154 narrowly defined affected sibling pairs (DSM-IV BPI) and 258 broadly defined affected sibling pairs (DSM-IV BPI, SABP, BPII, BPNOS or MDD(R)), approximately two thirds of all families contained at least one other additional typed individual. All individuals were genotyped using 398 highly polymorphic microsatellite markers from Applied Biosystemss Linkage Mapping Set Version 2. The average inter-marker distance was 9.6 cM and the mean heterozygosity was 0.78. Analysis of these data using non-parametric linkage methods (MAPMAKER/SIBS) found no evidence for loci of major effect and no regions reached genome-wide significance for either suggestive or significant linkage. We identified 19 points across the genome where the MLS exceeded a value set for follow up in our second stage screen (MLS≥0.74 (equivalent to a nominal pointwise significance of 5%) under the narrowest diagnostic model). These points were on chromosomes 2, 3, 4, 6, 7, 9, 10, 12, 17, 18 & X. Some of these points overlapped with previous linkage reports both within bipolar affective disorder and other psychiatric illnesses. Under the narrowest diagnostic model, the single most significant multipoint linkage was on chromosome 18 at marker D18S452 (MLS=1.54). Overall the highest MLS was 1.70 on chromosome 2 at marker D2S125, under the broadest diagnostic model.

Preferential Transmission of Paternal Alleles at Risk Genes in Attention-Deficit/Hyperactivity Disorder

Family, twin, and adoption studies have demonstrated a significant genetic contribution to the etiology of attention-deficit/hyperactivity disorder (ADHD). Pharmacological, neuroimaging, and animal-model findings suggest imbalances in monoaminergic (dopaminergic, serotonergic, and noradrenergic) neurotransmission in ADHD. We have examined monoaminergic candidate genes for possible genetic association with ADHD in the Irish population, focusing particularly on genes of the dopaminergic and serotonergic systems. We have observed that several of these genes are associated with ADHD, including DAT1, DBH, DRD4, DRD5, and 5HT1B. Here, we present what appears to be a systematic overtransmission of paternal alleles at candidate genes associated with ADHD. For the nine genes included in the analysis, the overall odds ratio for paternal transmission was 2, compared with 1.3 for maternal transmission (paternal vs. maternal chi 2=9.6; P=.0019). Transmission to females, from either parent, was significantly stronger than to males. Possible reasons for this preferential transmission include imprinting and ascertainment bias, although results of further analyses show that the latter is unlikely.

Stage 2 of the Wellcome Trust UK-Irish bipolar affective disorder sibling-pair genome screen: evidence for linkage on chromosomes 6q16-q21, 4q12-q21, 9p21, 10p14-p12 and 18q22

Bipolar affective disorder (BPAD) is a common psychiatric disorder with complex genetic aetiology. We have undertaken a genome-wide scan in one of the largest samples of bipolar affected sibling pairs (ASPs) using a two-stage approach combining sample splitting and marker grid tightening. In this second stage analysis, we have examined 17 regions that achieved a nominally significant maximum likelihood LOD score (MLS) threshold of 0.74 (or 1.18 for the X-chromosome) in stage one. The second stage has added 135 ASP families to bring the total stage 2 sample to 395 ASPs. In total, 494 microsatellite markers have been used to screen the human genome at a density of 10 cM in the first stage sample (260 ASPs) and 5 cM in the second stage. Under the broad diagnostic model, two markers gave LOD scores exceeding 3 with two-point analysis: D4S392 (LOD=3.30) and D10S197 (LOD=3.18). Multipoint analysis demonstrated suggestive evidence of linkage between BPAD and chromosomal regions 6q16–q21 (MLS=2.61) and 4q12–q21 (MLS=2.38). 6q16–q21 is of particular interest because our data, together with those from two recent genome scans, make this the best supported linkage region in BPAD. Further, our data show evidence of a gender effect at this locus with increased sharing predominantly within the male–male pairs. Our scan also provides support for linkage (MLS≥1.5) at several other regions that have been implicated in meta-analyses of bipolar disorder and/or schizophrenia including 9p21, 10p14–p12 and 18q22.

Association analysis of AKT1 and schizophrenia in a UK case control sample.

AKT1 (V-akt murine thyoma viral oncogene homolog 1) is involved in intracellular signalling pathways postulated as of aetiological importance in schizophrenia. Markers in the AKT1 gene have also recently been associated with schizophrenia in two samples of European origin and in Japanese and Iranian samples. Aiming to replicate these findings, we examined ten SNPs spanning AKT1 in a UK case-control sample (schizophrenia cases n=673, controls n=716). These included all SNPs previously reported to be associated in European, Japanese and Iranian samples, alone or in haplotypes, as well as additional markers defined by the Haploview Tagger program (pair-wise tagging, minimum r(2)=0.8, minor allele frequency=0.02). We found no association with single markers (min p=0.17). We found weak evidence for association (p=0.04) with a four marker haplotype reported as significant in the original positive European sample of Emamian et al. [Emamian, E.S., Hall, D., Birnbaum, M.J., Karayiorgou, M., Gogos, J.A., 2004. Convergent evidence for impaired AKT1-GSK3beta signaling in schizophrenia. Nat. Genet. 36, 131-137] and also an overlapping three marker haplotype (p=0.016) that had previously been reported as significant in a Japanese sample. Nominal p-values for these haplotypes did not survive correction for multiple testing. Our study provides at best weak support for the hypothesis that AKT1 is a susceptibility gene for schizophrenia. Examination of our own data and those of other groups leads us to conclude that overall, the evidence for association of AKT1 as a susceptibility gene for schizophrenia is weakly positive, but not yet convincing.

Genes of the Interleukin-18 Pathway Are Associated With Susceptibility to Barrett's Esophagus and Esophageal Adenocarcinoma

OBJECTIVES:To investigate the association of genetic polymorphisms of the interleukin-18 (IL-18) pathway to Barretts esophagus (BE) and esophageal adenocarcinoma (EAC). Most cases of EAC arise in a background of reflux-induced BE. Genetic influences in this pathway are poorly understood. IL-18 is a multifunctional cytokine implicated in anti-tumor immunity. A number of polymorphisms of the IL-18 and IL-18 receptor-accessory protein (IL-18RAP) genes have been reported to alter gene expression and have recently been linked to inflammatory processes and various tumors, but have not heretofore been studied in BE and EAC.METHODS:Two IL-18 promoter polymorphisms −137 G/C and −607 C/A, (rs187238 and rs1946518) and one IL-18RAP polymorphism (rs917997, C/T) were analyzed. Each single-nucleotide polymorphism (SNP) was genotyped in the following groups: EAC, BE, reflux esophagitis (RE), and controls and analyzed for association with disease status.RESULTS:The IL-18RAP rs917997C allele is strongly associated with a protective effect in BE (P=0.0002) and EAC (P=6 × 10−7), which approaches genome-wide levels of significance for allele association without incurring significant multiple testing. The CC genotype at IL-18RAP locus rs917997 was associated with a protective effect against esophageal disease (P=6 × 10−4, odds ratio (OR)=0.59, and 95% confidence interval (CI) 0.43–0.80 for BE; and P=2 × 10−6, OR=0.46, and 95% CI 0.34–0.64 for EAC). The genotype frequencies of IL-18−607 C/A were weakly associated with BE (P=0.02), and this trend was also seen between controls and EAC (P=0.07). The CC genotype was associated with an increased risk of BE (OR=1.45, 95% CI 1.07–1.98) and approached significance for EAC (OR=1.34, 95% CI 0.98–1.82). Allele and genotype frequencies at these loci were not significantly different between the RE group and controls. Although no significant association was observed between the disease groups at the –137 G/C locus, the −137G/−607C haplotype was associated with increased risk of BE (P=0.006) with haplotype frequencies of 55% in controls and 65% in BE.CONCLUSIONS:These data show a strong association of the IL-18RAP SNP rs917997 locus with BE and EAC and suggestive association of the Barretts population with the IL-18−607 C/A promoter polymorphism. As both of these SNPs have been demonstrated as expression quantitative trait loci affecting expression of the respective genes, this strongly implicates IL-18 signaling in susceptibility to BE and EAC.

The Association of Vitamin D Status with Acute Respiratory Morbidity in Preterm Infants

OBJECTIVE To assess the association between serum 25-hydroxyvitamin D (25OHD) levels and outcomes in preterm infants (<32 weeks gestation). STUDY DESIGN Serum 25OHD was measured in mothers and their infants within 24 hours of birth, before the start of enteral vitamin D supplementation, and at discharge from the neonatal intensive care unit. We evaluated the associations between vitamin D status and various early preterm outcomes. RESULTS Ninety-four preterm infants and their mothers were included; 92% of the infants had a 25OHD level≤50 nmol/L (20 ng/mL), and 64% had a 25OHD level<30 nmol/L (12 ng/mL). A low 25OHD level (<30 nmol/L) in preterm infants at birth was associated with increased oxygen requirement (P=.008), increased duration of intermittent positive-pressure ventilation during resuscitation at delivery (P=.032), and greater need for assisted ventilation (P=.013). CONCLUSION We observed a high prevalence of low 25OHD (<30 nmol/L), and found an association between vitamin D status and acute respiratory morbidity in preterm infants after birth.

Genome Wide Linkage Analysis of 972 Bipolar Pedigrees Using Single Nucleotide Polymorphisms

Because of the high costs associated with ascertainment of families, most linkage studies of Bipolar I disorder (BPI) have used relatively small samples. Moreover, the genetic information content reported in most studies has been less than 0.6. Although microsatellite markers spaced every 10 cM typically extract most of the genetic information content for larger multiplex families, they can be less informative for smaller pedigrees especially for affected sib pair kindreds. For these reasons we collaborated to pool family resources and carried out higher density genotyping. Approximately 1100 pedigrees of European ancestry were initially selected for study and were genotyped by the Center for Inherited Disease Research using the Illumina Linkage Panel 12 set of 6090 single-nucleotide polymorphisms. Of the ∼1100 families, 972 were informative for further analyses, and mean information content was 0.86 after pruning for linkage disequilibrium. The 972 kindreds include 2284 cases of BPI disorder, 498 individuals with bipolar II disorder (BPII) and 702 subjects with recurrent major depression. Three affection status models (ASMs) were considered: ASM1 (BPI and schizoaffective disorder, BP cases (SABP) only), ASM2 (ASM1 cases plus BPII) and ASM3 (ASM2 cases plus recurrent major depression). Both parametric and non-parametric linkage methods were carried out. The strongest findings occurred at 6q21 (non-parametric pairs LOD 3.4 for rs1046943 at 119 cM) and 9q21 (non-parametric pairs logarithm of odds (LOD) 3.4 for rs722642 at 78 cM) using only BPI and schizoaffective (SA), BP cases. Both results met genome-wide significant criteria, although neither was significant after correction for multiple analyses. We also inspected parametric scores for the larger multiplex families to identify possible rare susceptibility loci. In this analysis, we observed 59 parametric LODs of 2 or greater, many of which are likely to be close to maximum possible scores. Although some linkage findings may be false positives, the results could help prioritize the search for rare variants using whole exome or genome sequencing.