TagSNP approach for HLA risk allele genotyping of Saudi celiac disease patients: effectiveness and pitfalls

Background: Celiac disease (CD) is a genetically complex autoimmune disease which is triggered by dietary gluten. Human leukocyte antigen (HLA) class II genes are known to act as high-risk markers for CD, where >95% of CD patients carry (HLA), DQ2 and/or DQ8 alleles. Therefore, the present study was conducted to investigate the distribution of HLA haplotypes among Saudi CD patients and healthy controls by using the tag single nucleotide polymorphisms (SNP).Methods: HLA-tag SNPs showing strong linkage value (r²>0.99) were used to predict the HLA DQ2 and DQ8 genotypes in 101 Saudi CD patients and in 103 healthy controls by using real-time polymerase chain reaction technique. Genotype calls were further validated by Sanger sequencing method.Results: A total of 63.7% of CD cases and of 60.2% of controls were predicted to carry HLA-DQ2 and DQ8 heterodimers, either in the homozygous or heterozygous states. The prevalence of DQ8 in our CD patients was predicted to be higher than the patients from other ethnic populations (35.6%). More than 32% of the CD patients were found to be non-carriers of HLA risk haplotypes as predicted by the tag SNPs.Conclusion: The present study highlights that the Caucasian specific HLA-tag SNPs would be of limited value to accurately predict CD specific HLA haplotypes in Saudi population, when compared with the Caucasian groups. Prediction of risk haplotypes by tag SNPs in ethnic groups is a good alternate approach as long as the tag SNPs were identified from the local population genetic variant databases.     

Genome-wide pathway analysis of genome-wide association studies on systemic lupus erythematosus and rheumatoid arthritis

The aim of this study was to explore candidate single nucleotide polymorphisms (SNPs) and candidate mechanisms of systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA). Two SLE genome-wide association studies (GWASs) datasets were included in this study. Meta-analysis was conducted using 737,984 SNPs in 1,527 SLE cases and 3,421 controls of European ancestry, and 4,429 SNPs that met a threshold of p?<?0.01 in a Korean RA GWAS dataset was used. ICSNPathway (identify candidate causal SNPs and pathways) analysis was applied to the meta-analysis results of the SLE GWAS datasets, and a RA GWAS dataset. The most significant result of SLE GWAS meta-analysis concerned rs2051549 in the human leukocyte antigen (HLA) region (p?=?3.36E?22). In the non-HLA region, meta-analysis identified 6 SNPs associated with SLE with genome-wide significance (STAT4, TNPO3, BLK, FAM167A, and IRF5). ICSNPathway identified five candidate causal SNPs and 13 candidate causal pathways. This pathway-based analysis provides three hypotheses of the biological mechanism involved. First, rs8084 and rs7192?→?HLA-DRA?→?bystander B cell activation. Second, rs1800629?→?TNF?→?cytokine network. Third, rs1150752 and rs185819?→?TNXB?→?collagen metabolic process. ICSNPathway analysis identified three candidate causal non-HLA SNPs and four candidate causal pathways involving the PADI4, MTR, PADI2, and TPH2 genes of RA. We identified five candidate SNPs and thirteen pathways, involving bystander B cell activation, cytokine network, and collagen metabolic processing, which may contribute to SLE susceptibility, and we revealed candidate causal non-HLA SNPs, genes, and pathways of RA.     

Effective detection of human leukocyte antigen risk alleles in celiac disease using tag single nucleotide polymorphisms

Background

The HLA genes, located in the MHC region on chromosome 6p21.3, play an important role in many autoimmune disorders, such as celiac disease (CD), type 1 diabetes (T1D), rheumatoid arthritis, multiple sclerosis, psoriasis and others. Known HLA variants that confer risk to CD, for example, include DQA1*05/DQB1*02 (DQ2.5) and DQA1*03/DQB1*0302 (DQ8). To diagnose the majority of CD patients and to study disease susceptibility and progression, typing these strongly associated HLA risk factors is of utmost importance. However, current genotyping methods for HLA risk factors involve many reactions, and are complicated and expensive. We sought a simple experimental approach using tagging SNPs that predict the CD-associated HLA risk factors.

Methodology

Our tagging approach exploits linkage disequilibrium between single nucleotide polymorphism (SNPs) and the CD-associated HLA risk factors DQ2.5 and DQ8 that indicate direct risk, and DQA1*0201/DQB1*0202 (DQ2.2) and DQA1*0505/DQB1*0301 (DQ7) that attribute to the risk of DQ2.5 to CD. To evaluate the predictive power of this approach, we performed an empirical comparison of the predicted DQ types, based on these six tag SNPs, with those executed with current validated laboratory typing methods of the HLA-DQA1 and -DQB1 genes in three large cohorts. The results were validated in three European celiac populations.

Conclusion

Using this method, only six SNPs were needed to predict the risk types carried by >95% of CD patients. We determined that for this tagging approach the sensitivity was >0.991, specificity >0.996 and the predictive value >0.948. Our results show that this tag SNP method is very accurate and provides an excellent basis for population screening for CD. This method is broadly applicable in European populations.     

Cost-effective HLA typing with tagging SNPs predicts celiac disease risk haplotypes in the Finnish, Hungarian, and Italian populations

Human leukocyte antigen (HLA) genes, located on chromosome 6p21.3, have a crucial role in susceptibility to various autoimmune and inflammatory diseases, such as celiac disease and type 1 diabetes. Certain HLA heterodimers, namely DQ2 (encoded by the DQA1*05 and DQB1*02 alleles) and DQ8 (DQA1*03 and DQB1*0302), are necessary for the development of celiac disease. Traditional genotyping of HLA genes is laborious, time-consuming, and expensive. A novel HLA-genotyping method, using six HLA-tagging single-nucleotide polymorphisms (SNPs) and suitable for high-throughput approaches, was described recently. Our aim was to validate this method in the Finnish, Hungarian, and Italian populations. The six previously reported HLA-tagging SNPs were genotyped in patients with celiac disease and in healthy individuals from Finland, Hungary, and two distinct regions of Italy. The potential of this method was evaluated in analyzing how well the tag SNP results correlate with the HLA genotypes previously determined using traditional HLA-typing methods. Using the tagging SNP method, it is possible to determine the celiac disease risk haplotypes accurately in Finnish, Hungarian, and Italian populations, with specificity and sensitivity ranging from 95% to 100%. In addition, it predicts homozygosity and heterozygosity for a risk haplotype, allowing studies on genotypic risk effects. The method is transferable between populations and therefore suited for large-scale research studies and screening of celiac disease among high-risk individuals or at the population level. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Lotta Koskinen and Jihane Romanos are authors with equal contribution.     

Improving the estimation of celiac disease sibling risk by non-HLA genes

Celiac Disease (CD) is a polygenic trait, and HLA genes explain less than half of the genetic variation. Through large GWAs more than 40 associated non-HLA genes were identified, but they give a small contribution to the heritability of the disease. The aim of this study is to improve the estimate of the CD risk in siblings, by adding to HLA a small set of non-HLA genes. One-hundred fifty-seven Italian families with a confirmed CD case and at least one other sib and both parents were recruited. Among 249 sibs, 29 developed CD in a 6 year follow-up period. All individuals were typed for HLA and 10 SNPs in non-HLA genes: CCR1/CCR3 (rs6441961), IL12A/SCHIP1 and IL12A (rs17810546 and rs9811792), TAGAP (rs1738074), RGS1 (rs2816316), LPP (rs1464510), OLIG3 (rs2327832), REL (rs842647), IL2/IL21 (rs6822844), SH2B3 (rs3184504). Three associated SNPs (in LPP, REL, and RGS1 genes) were identified through the Transmission Disequilibrium Test and a Bayesian approach was used to assign a score (BS) to each detected HLA+SNPs genotype combination. We then classified CD sibs as at low or at high risk if their BS was respectively < or ≥ median BS value within each HLA risk group. A larger number (72%) of CD sibs showed a BS ≥ the median value and had a more than two fold higher OR than CD sibs with a BS value < the median (O.R = 2.53, p = 0.047). Our HLA+SNPs genotype classification, showed both a higher predictive negative value (95% vs 91%) and diagnostic sensitivity (79% vs 45%) than the HLA only. In conclusion, the estimate of the CD risk by HLA+SNPs approach, even if not applicable to prevention, could be a precious tool to improve the prediction of the disease in a cohort of first degree relatives, particularly in the low HLA risk groups.     

Genome-wide association study reveals the genetic architecture of flowering time in rapeseed (Brassica napus L.)

Flowering time adaptation is a major breeding goal in the allopolyploid species Brassica napus. To investigate the genetic architecture of flowering time, a genome-wide association study (GWAS) of flowering time was conducted with a diversity panel comprising 523 B. napus cultivars and inbred lines grown in eight different environments. Genotyping was performed with a Brassica 60K Illumina Infinium SNP array. A total of 41 single-nucleotide polymorphisms (SNPs) distributed on 14 chromosomes were found to be associated with flowering time, and 12 SNPs located in the confidence intervals of quantitative trait loci (QTL) identified in previous researches based on linkage analyses. Twenty-five candidate genes were orthologous to Arabidopsis thaliana flowering genes. To further our understanding of the genetic factors influencing flowering time in different environments, GWAS was performed on two derived traits, environment sensitivity and temperature sensitivity. The most significant SNPs were found near Bn-scaff_16362_1-p380982, just 13 kb away from BnaC09g41990D, which is orthologous to A. thaliana CONSTANS (CO), an important gene in the photoperiod flowering pathway. These results provide new insights into the genetic control of flowering time in B. napus and indicate that GWAS is an effective method by which to reveal natural variations of complex traits in B. napus.     

Identification of Non-HLA Genes Associated with Celiac Disease and Country-Specific Differences in a Large,International Pediatric Cohort

Objectives

There are significant geographical differences in the prevalence and incidence of celiac disease that cannot be explained by HLA alone. More than 40 loci outside of the HLA region have been associated with celiac disease. We investigated the roles of these non-HLA genes in the development of tissue transglutaminase autoantibodies (tTGA) and celiac disease in a large international prospective cohort study.

Methods

A total of 424,788 newborns from the US and European general populations and first-degree relatives with type 1 diabetes were screened for specific HLA genotypes. Of these, 21,589 carried 1 of the 9 HLA genotypes associated with increased risk for type 1 diabetes and celiac disease; we followed 8676 of the children in a 15 y prospective follow-up study. Genotype analyses were performed on 6010 children using the Illumina ImmunoChip. Levels of tTGA were measured in serum samples using radio-ligand binding assays; diagnoses of celiac disease were made based on persistent detection of tTGA and biopsy analysis. Data were analyzed using Cox proportional hazards analyses.

Results

We found 54 single-nucleotide polymorphisms (SNPs) in 5 genes associated with celiac disease (TAGAP, IL18R1, RGS21, PLEK, and CCR9) in time to celiac disease analyses (10⁻⁴>P>5.8x10⁻⁶). The hazard ratios (HR) for the SNPs with the smallest P values in each region were 1.59, 1.45, 2.23, 2.64, and 1.40, respectively. Outside of regions previously associated with celiac disease, we identified 10 SNPs in 8 regions that could also be associated with the disease (P<10⁻⁴). A SNP near PKIA (rs117128341, P = 6.5x10⁻⁸, HR = 2.8) and a SNP near PFKFB3 (rs117139146, P<2.8x10⁻⁷, HR = 4.9) reached the genome-wide association threshold in subjects from Sweden. Analyses of time to detection of tTGA identified 29 SNPs in 2 regions previously associated with celiac disease (CTLA4, P = 1.3x10⁻⁶, HR = 0.76 and LPP, P = 2.8x10⁻⁵, HR = .80) and 6 SNPs in 5 regions not previously associated with celiac disease (P<10⁻⁴); non-HLA genes are therefore involved in development of tTGA.

Conclusions

In conclusion, using a genetic analysis of a large international cohort of children, we associated celiac disease development with 5 non-HLA regions previously associated with the disease and 8 regions not previously associated with celiac disease. We identified 5 regions associated with development of tTGA. Two loci associated with celiac disease progression reached a genome-wide association threshold in subjects from Sweden.     

No allelic variation in genes with high gliadin homology in patients with celiac disease and type 1 diabetes

Celiac disease (CD) is a complex inflammatory disorder of the small intestine, induced by dietary gluten in genetically susceptible individuals. CD is strongly associated with HLA-DQ2 and it has recently been established that gut-derived DQ2-restricted T cells from patients with CD predominantly recognize gluten-derived peptides in which specific glutamine residues are deamidated to glutamic acid by tissue transglutaminase. Recently, intestinally expressed human genes with high homology to DQ2-gliadin celiac T-cell epitopes have been identified. Single or double point mutations which would increase the celiac T-cell epitope homology, and mutation in these genes, leading to the expression of glutamic acid at particular positions, could hypothetically be involved in the initiation of CD in HLA-DQ2-positive children. Six gene regions with high celiac T-cell epitope homology were investigated for single-nucleotide polymorphisms using direct sequencing of DNA from 20 CD patients, 27 type 1 diabetes mellitus (T1DM) patients with associated CD, 24 patients with T1DM without CD and 110 healthy controls, all of Caucasian origin. No variants in any of these genes in any of the investigated groups were found. We conclude that gut-expressed human celiac epitope homologous peptides are unlikely to represent non-HLA risk factors in the development of celiac disease in Caucasians.     

Pooled sample-based GWAS: a cost-effective alternative for identifying colorectal and prostate cancer risk variants in the Polish population

Background

Prostate cancer (PCa) and colorectal cancer (CRC) are the most commonly diagnosed cancers and cancer-related causes of death in Poland. To date, numerous single nucleotide polymorphisms (SNPs) associated with susceptibility to both cancer types have been identified, but their effect on disease risk may differ among populations.

Methods

To identify new SNPs associated with PCa and CRC in the Polish population, a genome-wide association study (GWAS) was performed using DNA sample pools on Affymetrix Genome-Wide Human SNP 6.0 arrays. A total of 135 PCa patients and 270 healthy men (PCa sub-study) and 525 patients with adenoma (AD), 630 patients with CRC and 690 controls (AD/CRC sub-study) were included in the analysis. Allele frequency distributions were compared with t-tests and χ²-tests. Only those significantly associated SNPs with a proxy SNP (p<0.001; distance of 100 kb; r²>0.7) were selected. GWAS marker selection was conducted using PLINK. The study was replicated using extended cohorts of patients and controls. The association with previously reported PCa and CRC susceptibility variants was also examined. Individual patients were genotyped using TaqMan SNP Genotyping Assays.

Results

The GWAS selected six and 24 new candidate SNPs associated with PCa and CRC susceptibility, respectively. In the replication study, 17 of these associations were confirmed as significant in additive model of inheritance. Seven of them remained significant after correction for multiple hypothesis testing. Additionally, 17 previously reported risk variants have been identified, five of which remained significant after correction.

Conclusion

Pooled-DNA GWAS enabled the identification of new susceptibility loci for CRC in the Polish population. Previously reported CRC and PCa predisposition variants were also identified, validating the global nature of their associations. Further independent replication studies are required to confirm significance of the newly uncovered candidate susceptibility loci.     

A whole genome analyses of genetic variants in two Kelantan Malay individuals

BackgroundThe sequencing of two members of the Royal Kelantan Malay family genomes will provide insights on the Kelantan Malay whole genome sequences. The two Kelantan Malay genomes were analyzed for the SNP markers associated with thalassemia and Helicobacter pylori infection. Helicobacter pylori infection was reported to be low prevalence in the north-east as compared to the west coast of the Peninsular Malaysia and beta-thalassemia was known to be one of the most common inherited and genetic disorder in Malaysia.ResultBy combining SNP information from literatures, GWAS study and NCBI ClinVar, 18 unique SNPs were selected for further analysis. From these 18 SNPs, 10 SNPs came from previous study of Helicobacter pylori infection among Malay patients, 6 SNPs were from NCBI ClinVar and 2 SNPs from GWAS studies. The analysis reveals that both Royal Kelantan Malay genomes shared all the 10 SNPs identified by Maran (Single Nucleotide Polymorphims (SNPs) genotypic profiling of Malay patients with and without Helicobacter pylori infection in Kelantan, 2011) and one SNP from GWAS study. In addition, the analysis also reveals that both Royal Kelantan Malay genomes shared 3 SNP markers; HBG1 (rs1061234), HBB (rs1609812) and BCL11A (rs766432) where all three markers were associated with beta-thalassemia.ConclusionsOur findings suggest that the Royal Kelantan Malays carry the SNPs which are associated with protection to Helicobacter pylori infection. In addition they also carry SNPs which are associated with beta-thalassemia. These findings are in line with the findings by other researchers who conducted studies on thalassemia and Helicobacter pylori infection in the non-royal Malay population.     

Association of Genetic Markers with CSF Oligoclonal Bands in Multiple Sclerosis Patients

Objective

to explore the association between genetic markers and Oligoclonal Bands (OCB) in the Cerebro Spinal Fluid (CSF) of Italian Multiple Sclerosis patients.

Methods

We genotyped 1115 Italian patients for HLA-DRB1*15 and HLA-A*02. In a subset of 925 patients we tested association with 52 non-HLA SNPs associated with MS susceptibility and we calculated a weighted Genetic Risk Score. Finally, we performed a Genome Wide Association Study (GWAS) with OCB status on a subset of 562 patients. The best associated SNPs of the Italian GWAS were replicated in silico in Scandinavian and Belgian populations, and meta-analyzed.

Results

HLA-DRB1*15 is associated with OCB+: p = 0.03, Odds Ratio (OR) = 1.6, 95% Confidence Limits (CL) = 1.1–2.4. None of the 52 non-HLA MS susceptibility loci was associated with OCB, except one SNP (rs2546890) near IL12B gene (OR: 1.45; 1.09–1.92). The weighted Genetic Risk Score mean was significantly (p = 0.0008) higher in OCB+ (7.668) than in OCB− (7.412) patients. After meta-analysis on the three datasets (Italian, Scandinavian and Belgian) for the best associated signals resulted from the Italian GWAS, the strongest signal was a SNP (rs9320598) on chromosome 6q (p = 9.4×10⁻⁷) outside the HLA region (65 Mb).

Discussion

genetic factors predispose to the development of OCB.     

HLA and non-HLA genes in Beh?et’s disease: a multicentric study in the Spanish population

Introduction

According to genome wide association (GWA) studies as well as candidate gene approaches, Behçet’s disease (BD) is associated with human leukocyte antigen (HLA)-A and HLA-B gene regions. The HLA-B51 has been consistently associated with the disease, but the role of other HLA class I molecules remains controversial. Recently, variants in non-HLA genes have also been associated with BD. The aims of this study were to further investigate the influence of the HLA region in BD and to explore the relationship with non-HLA genes recently described to be associated in other populations.

Methods

This study included 304 BD patients and 313 ethnically matched controls. HLA-A and HLA-B low resolution typing was carried out by PCR-SSOP Luminex. Eleven tag single nucleotide polymorphisms (SNPs) located outside of the HLA-region, previously described associated with the disease in GWA studies and having a minor allele frequency in Caucasians greater than 0.15 were genotyped using TaqMan assays. Phenotypic and genotypic frequencies were estimated by direct counting and distributions were compared using the χ² test.

Results

In addition to HLA-B*51, HLA-B*57 was found as a risk factor in BD, whereas, B*35 was found to be protective. Other HLA-A and B specificities were suggestive of association with the disease as risk (A*02 and A*24) or protective factors (A*03 and B*58). Regarding the non-HLA genes, the three SNPs located in IL23R and one of the SNPs in IL10 were found to be significantly associated with susceptibility to BD in our population.

Conclusion

Different HLA specificities are associated with Behçet’s disease in addition to B*51. Other non-HLA genes, such as IL23R and IL-10, play a role in the susceptibility to the disease.     

Investigation of a genome wide association signal for obesity: synthetic association and haplotype analyses at the melanocortin 4 receptor gene locus

Background

Independent genome-wide association studies (GWAS) showed an obesogenic effect of two single nucleotide polymorphisms (SNP; rs12970134 and rs17782313) more than 150 kb downstream of the melanocortin 4 receptor gene (MC4R). It is unclear if the SNPs directly influence MC4R function or expression, or if the SNPs are on a haplotype that predisposes to obesity or includes functionally relevant genetic variation (synthetic association). As both exist, functionally relevant mutations and polymorphisms in the MC4R coding region and a robust association downstream of the gene, MC4R is an ideal model to explore synthetic association.

Methodology/Principal Findings

We analyzed a genomic region (364.9 kb) encompassing the MC4R in GWAS data of 424 obesity trios (extremely obese child/adolescent and both parents). SNP rs12970134 showed the lowest p-value (p = 0.004; relative risk for the obesity effect allele: 1.37); conditional analyses on this SNP revealed that 7 of 78 analyzed SNPs provided independent signals (p≤0.05). These 8 SNPs were used to derive two-marker haplotypes. The three best (according to p-value) haplotype combinations were chosen for confirmation in 363 independent obesity trios. The confirmed obesity effect haplotype includes SNPs 3′ and 5′ of the MC4R. Including MC4R coding variants in a joint model had almost no impact on the effect size estimators expected under synthetic association.

Conclusions/Significance

A haplotype reaching from a region 5′ of the MC4R to a region at least 150 kb from the 3′ end of the gene showed a stronger association to obesity than single SNPs. Synthetic association analyses revealed that MC4R coding variants had almost no impact on the association signal. Carriers of the haplotype should be enriched for relevant mutations outside the MC4R coding region and could thus be used for re-sequencing approaches. Our data also underscore the problems underlying the identification of relevant mutations depicted by GWAS derived SNPs.     

Identification of novel genetic markers associated with clinical phenotypes of systemic sclerosis through a genome-wide association strategy

The aim of this study was to determine, through a genome-wide association study (GWAS), the genetic components contributing to different clinical sub-phenotypes of systemic sclerosis (SSc). We considered limited (lcSSc) and diffuse (dcSSc) cutaneous involvement, and the relationships with presence of the SSc-specific auto-antibodies, anti-centromere (ACA), and anti-topoisomerase I (ATA). Four GWAS cohorts, comprising 2,296 SSc patients and 5,171 healthy controls, were meta-analyzed looking for associations in the selected subgroups. Eighteen polymorphisms were further tested in nine independent cohorts comprising an additional 3,175 SSc patients and 4,971 controls. Conditional analysis for associated SNPs in the HLA region was performed to explore their independent association in antibody subgroups. Overall analysis showed that non-HLA polymorphism rs11642873 in IRF8 gene to be associated at GWAS level with lcSSc (P = 2.32×10⁻¹², OR = 0.75). Also, rs12540874 in GRB10 gene (P = 1.27 × 10⁻⁶, OR = 1.15) and rs11047102 in SOX5 gene (P = 1.39×10⁻⁷, OR = 1.36) showed a suggestive association with lcSSc and ACA subgroups respectively. In the HLA region, we observed highly associated allelic combinations in the HLA-DQB1 locus with ACA (P = 1.79×10⁻⁶¹, OR = 2.48), in the HLA-DPA1/B1 loci with ATA (P = 4.57×10⁻⁷⁶, OR = 8.84), and in NOTCH4 with ACA P = 8.84×10⁻²¹, OR = 0.55) and ATA (P = 1.14×10⁻⁸, OR = 0.54). We have identified three new non-HLA genes (IRF8, GRB10, and SOX5) associated with SSc clinical and auto-antibody subgroups. Within the HLA region, HLA-DQB1, HLA-DPA1/B1, and NOTCH4 associations with SSc are likely confined to specific auto-antibodies. These data emphasize the differential genetic components of subphenotypes of SSc.     

High Frequency of Haplotype HLA-DQ7 in Celiac Disease Patients from South Italy: Retrospective Evaluation of 5,535 Subjects at Risk of Celiac Disease

Background

Celiac disease (CD) has a strong genetic component mainly due to HLA DQ2/DQ8 encoding genes. However, a minority of CD patients are DQ2/DQ8-negative. To address this issue, we retrospectively characterized HLA haplotypes in 5,535 subjects at risk of CD (either relatives of CD patients or subjects with CD-like symptoms) referred to our center during a 10-year period.

Methods

We identified loci DQA1/DQB1/DRB1 by sequence-specific oligonucleotide-PCR and sequence-specific primer-PCR; anti-transglutaminase IgA/IgG and anti-endomysium IgA by ELISA and indirect immunofluorescence, respectively.

Results

We diagnosed CD in 666/5,535 individuals, 4.2% of whom were DQ2/DQ8-negative. Interestingly, DQ7 was one of the most abundant haplotypes in all CD patients and significantly more frequent in DQ2/DQ8-negative (38%) than in DQ2/DQ8-positive CD patients (24%) (p<0.05).

Conclusion

Our data lend support to the concept that DQ7 represents an additive or independent CD risk haplotype with respect to DQ2/DQ8 haplotypes but this finding should be verified in other large CD populations.     

Structural and functional studies of trans-encoded HLA-DQ2.3 (DQA1*03:01/DQB1*02:01) protein molecule

MHC class II molecules are composed of one α-chain and one β-chain whose membrane distal interface forms the peptide binding groove. Most of the existing knowledge on MHC class II molecules comes from the cis-encoded variants where the α- and β-chain are encoded on the same chromosome. However, trans-encoded class II MHC molecules, where the α- and β-chain are encoded on opposite chromosomes, can also be expressed. We have studied the trans-encoded class II HLA molecule DQ2.3 (DQA1*03:01/DQB1*02:01) that has received particular attention as it may explain the increased risk of certain individuals to type 1 diabetes. We report the x-ray crystal structure of this HLA molecule complexed with a gluten epitope at 3.05 Å resolution. The gluten epitope, which is the only known HLA-DQ2.3-restricted epitope, is preferentially recognized in the context of the DQ2.3 molecule by T-cell clones of a DQ8/DQ2.5 heterozygous celiac disease patient. This preferential recognition can be explained by improved HLA binding as the epitope combines the peptide-binding motif of DQ2.5 (negative charge at P4) and DQ8 (negative charge at P1). The analysis of the structure of DQ2.3 together with all other available DQ crystal structures and sequences led us to categorize DQA1 and DQB1 genes into two groups where any α-chain and β-chain belonging to the same group are expected to form a stable heterodimer.     

Genetic variants associated with breast cancer risk for Ashkenazi Jewish women with strong family histories but no identifiable BRCA1/2 mutation

The ability to establish genetic risk models is critical for early identification and optimal treatment of breast cancer. For such a model to gain clinical utility, more variants must be identified beyond those discovered in previous genome-wide association studies (GWAS). This is especially true for women at high risk because of family history, but without BRCA1/2 mutations. This study incorporates three datasets in a GWAS analysis of women with Ashkenazi Jewish (AJ) homogeneous ancestry. Two independent discovery cohorts comprised 239 and 238 AJ women with invasive breast cancer or preinvasive ductal carcinoma in situ and strong family histories of breast cancer, but lacking the three BRCA1/2 founder mutations, along with 294 and 230 AJ controls, respectively. An independent, third cohort of 203 AJ cases with familial breast cancer history and 263 healthy controls of AJ women was used for validation. A total of 19 SNPs were identified as associated with familial breast cancer risk in AJ women. Among these SNPs, 13 were identified from a panel of 109 discovery SNPs, including an FGFR2 haplotype. In addition, six previously identified breast cancer GWAS SNPs were confirmed in this population. Seven of the 19 markers were significant in a multivariate predictive model of familial breast cancer in AJ women, three novel SNPs [rs17663555(5q13.2), rs566164(6q21), and rs11075884(16q22.2)], the FGFR2 haplotype, and three previously published SNPs [rs13387042(2q35), rs2046210(ESR1), and rs3112612(TOX3)], yielding moderate predictive power with an area under the curve (AUC) of the ROC (receiver-operator characteristic curve) of 0.74. Population-specific genetic variants in addition to variants shared with populations of European ancestry may improve breast cancer risk prediction among AJ women from high-risk families without founder BRCA1/2 mutations.     

Improvement in Prediction of Coronary Heart Disease Risk over Conventional Risk Factors Using SNPs Identified in Genome-Wide Association Studies

Objective

We examined whether a panel of SNPs, systematically selected from genome-wide association studies (GWAS), could improve risk prediction of coronary heart disease (CHD), over-and-above conventional risk factors. These SNPs have already demonstrated reproducible associations with CHD; here we examined their use in long-term risk prediction.

Study Design and Setting

SNPs identified from meta-analyses of GWAS of CHD were tested in 840 men and women aged 55–75 from the Edinburgh Artery Study, a prospective, population-based study with 15 years of follow-up. Cox proportional hazards models were used to evaluate the addition of SNPs to conventional risk factors in prediction of CHD risk. CHD was classified as myocardial infarction (MI), coronary intervention (angioplasty, or coronary artery bypass surgery), angina and/or unspecified ischaemic heart disease as a cause of death; additional analyses were limited to MI or coronary intervention. Model performance was assessed by changes in discrimination and net reclassification improvement (NRI).

Results

There were significant improvements with addition of 27 SNPs to conventional risk factors for prediction of CHD (NRI of 54%, P<0.001; C-index 0.671 to 0.740, P = 0.001), as well as MI or coronary intervention, (NRI of 44%, P<0.001; C-index 0.717 to 0.750, P = 0.256). ROC curves showed that addition of SNPs better improved discrimination when the sensitivity of conventional risk factors was low for prediction of MI or coronary intervention.

Conclusion

There was significant improvement in risk prediction of CHD over 15 years when SNPs identified from GWAS were added to conventional risk factors. This effect may be particularly useful for identifying individuals with a low prognostic index who are in fact at increased risk of disease than indicated by conventional risk factors alone.