Sequential Sentinel SNP Regional Association Plots (SSS‐RAP): An Approach for Testing Independence of SNP Association Signals Using Meta‐Analysis Data

Genome‐Wide Association Studies (GWAS) frequently incorporate meta‐analysis within their framework. However, conditional analysis of individual‐level data, which is an established approach for fine mapping of causal sites, is often precluded where only group‐level summary data are available for analysis. Here, we present a numerical and graphical approach, “sequential sentinel SNP regional association plot” (SSS‐RAP), which estimates regression coefficients (beta) with their standard errors using the meta‐analysis summary results directly. Under an additive model, typical for genes with small effect, the effect for a sentinel SNP can be transformed to the predicted effect for a possibly dependent SNP through a 2×2 2‐SNP haplotypes table. The approach assumes Hardy–Weinberg equilibrium for test SNPs. SSS‐RAP is available as a Web‐tool ( http://apps.biocompute.org.uk/sssrap/sssrap.cgi ). To develop and illustrate SSS‐RAP we analyzed lipid and ECG traits data from the British Women's Heart and Health Study (BWHHS), evaluated a meta‐analysis for ECG trait and presented several simulations. We compared results with existing approaches such as model selection methods and conditional analysis. Generally findings were consistent. SSS‐RAP represents a tool for testing independence of SNP association signals using meta‐analysis data, and is also a convenient approach based on biological principles for fine mapping in group level summary data.     

Novel polymorphisms in the promoter region of the neurotrophin‐3 gene and their associations with schizophrenia

Based on the neurodevelopmental hypothesis in the etiology of schizophrenia, neurotrophic factors may be involved in the pathogenesis of the illness. We searched for polymorphisms in the promoter region of the neurotrophin‐3 (NTF3) gene by using denaturing high performance liquid chromatography (DHPLC). Six single nucleotide polymorphisms (SNPs) were found. When these polymorphisms were examined for association with schizophrenia, a weakly significant difference was observed in the genotype distribution of the G/? 3004/A polymorphism between 184 schizophrenics and 185 controls (P < 0.05), although no statistically significant association was detected in a family‐based sample of 50 trios (schizophrenics and their parents). With respect to the other polymorphisms, there was no significant association with schizophrenia. The G/? 3004/A polymorphism was in linkage disequilibrium with the CA repeat polymorphism in the first intron of the NTF3 gene. When haplotype‐based analysis was performed, an increased frequency of the haplotype containing the G(? 3004) and the “A3” ([CA]₂₃) alleles was observed for the schizophrenics compared to controls. Our results suggest that the G(? 3004)‐A3 haplotype has a modest effect of giving susceptibility to schizophrenia. © 2002 Wiley‐Liss, Inc.     

Association between ADAM33 polymorphisms and adult asthma in the Japanese population

Background ADAM33, a member of the ADAM (a disintegrin and metalloprotease) family, is a putative asthma susceptibility gene recently identified by positional cloning. It is important to know whether the association exists in ethnically diverse populations. Objective To assess whether genetic functional variants of ADAM33 relate to the susceptibility or some phenotypes in adult patients with bronchial asthma in a Japanese population. Methods We searched for single nucleotide polymorphisms (SNPs) in ADAM33 by PCR‐directed sequencing and identified 48 SNPs. Fourteen SNPs were selected with regard to the LD pattern, and genotyped by Taq‐Man and PCR–RFLP methods. We conducted an association study of ADAM33 with 504 adult asthmatic patients and 651 controls, and haplotype analyses of related variants were performed. Results Significant associations with asthma were found for the SNPs T1 (Met764Thr), T2 (Pro774Ser), S2 and V?3 (with the lowest P‐value for T1, P=0.0015; OR 0.63). We analysed the haplotype using these four polymorphisms, and found a positive association with haplotype CCTG (P=0.0024). Conclusion Our results replicate associations reported recently in other ethnic populations, and suggest that the ADAM33 gene is involved in the development of asthma through genetic polymorphisms.     

应用连锁不平衡图谱的关联研究分析方法

目的提出一种对高通量单核苷酸多态位点(single nucleotide polymorphism,SNP)关联研究的数据分析方法。方法在160名上海地区中国人中进行754个SNP的基因型检测,分别构建病例组和对照组的连锁不平衡(linkage disequilibrium,LD)图谱,通过比较两组间染色体区域LD图谱随物理距离的变化趋势寻找与疾病相关的位点,并与传统LD分析以及SNP单点、单倍型分析进行比较。结果LD图谱的分析能判断出两组间LD存在差异的染色体区域,并且该区域SNP等位基因频率和单倍型频率在两组间分布存在统计学差异或差异趋势。结论可应用该方法对高通量SNP的关联研究进行数据分析。     

Probability that a Two‐Stage Genome‐Wide Association Study will Detect a Disease‐Associated SNP and Implications for Multistage Designs

Large two‐stage genome‐wide association studies (GWASs) have been shown to reduce required genotyping with little loss of power, compared to a one‐stage design, provided a substantial fraction of cases and controls, π_sample , is included in stage 1. However, a number of recent GWASs have used π_sample < 0.2 . Moreover, standard power calculations are not applicable because SNPs are selected in stage 1 by ranking their p‐values, rather than comparing each SNP's statistic to a fixed critical value. We define the detection probability (DP) of a two‐stage design as the probability that a given disease‐associated SNP will have a p‐value among the lowest ranks of p‐values at stage 1, and, among those SNPs selected at stage 1, at stage 2. For 8000 cases and 8000 controls available for study and for odds ratios per allele in the range 1.1‐1.3, we show that DP is substantially reduced for designs with π_sample≤ 0.25 , and that DP cannot be appreciably increased by analyzing the stage 1 and stage 2 data jointly. These results suggest that multistage designs with small first stages (e.g. π_sample≤ 0.25 ) should be avoided, and that additional genotyping in earlier studies with small first stages will yield previously unselected disease‐associated SNPs.     

单核苷酸多态性与连锁不平衡研究进展

单核苷酸多态性(single nucleotide polymorphism,SNP)是人类基因组中最广泛的多态性现象,也是造成个体差异的最主要的遗传原因,发现和研究SNP的工作在目前人类基因组研究中倍受关注。连锁不平衡是不同遗传标记问存在着的非随机组合现象,SNP作为极具优势的遗传标记为深入研究连锁不平衡、以及利用连锁不平衡进行群体遗传学的参数估计、基因精细定位、关联分析等提供了良好的先决条件。最近,在SNP研究及连锁不平衡的度量和连锁不平衡性质的研究方面取得的一系列进展为遗传学在将来发展奠定了基础。     

基于标签单核苷酸多态性单倍型和单倍域的构建及其在关联研究中的应用

人类基因组中单倍型(haplotype)和单倍域(haplotype block)的结构提供了人类进化的宝贵信息,并成为发现人类复杂疾病易感基因的有效策略。一个单倍域可分割成多个具有有限单倍型多样性的离散的区域,代表每个区域结构特征的少量标签单核苷酸多态性(tag single nucleotide polymorphism,tSNP)可使绝大部分单倍型相互区分开来。因此,标签SNP在单倍型和单倍域的构建和关联研究中具有重要地位。构建单倍型和单倍域的方法分为两类,分别是基于大家系中基因分型数据和基于统计学的算法。通过系统回顾几种单倍型和单倍域的构建方法,了解它们在不同的疾病模型或根据不同的分割标准,进行关联研究的检验效能,客观评价每种方法的优、缺点、应用前景及其在关联研究中的应用。随着国际人类基因组单倍型图的完成和单倍型构建统计学运算规则的完善,融合数学、物理学、计算机科学等学科的单倍型构建方法将对人类遗传学、复杂疾病易感基因的定位和克隆鉴定等生命科学的相关领域产生深远的影响。     

Linkage Disequilibrium and Haplotype Architecture for two ABC Transporter Genes (ABCC1 and ABCG2) in Chinese Population: Implications for Pharmacogenomic Association Studies

Information about linkage disequilibrium (LD) patterns and haplotype structures for candidate genes is instructive for the design and analysis of genetic association studies for complex diseases and drug response. ABCC1 and ABCG2 are genes coding for two multidrug resistance (MDR) associated transporters; they are also related to some pathophysiological traits. To pinpoint the LD profiles of these MDR genes in Chinese, we systemically screened 27 unrelated individuals for single nucleotide polymorphisms (SNPs) in the coding and regulatory regions of these genes, and thereby characterized their haplotype structures. Despite marked variations in haplotype diversity, LD pattern and intragenic recombination intensity between the two genes, both loci could be partitioned into several LD blocks, in which a modest number of haplotypes accounted for a high fraction of the sampled chromosomes. We concluded that each locus has its own genomic LD profile, but that they still share a common segmental LD architecture with low haplotype diversity. Our data will benefit genetic association studies of complex traits and drug response possibly related to these genes.     

Identification of Association of Common AGGF1 Variants with Susceptibility for Klippel-Trenaunay Syndrome Using the Structure Association Program

Klippel-Trenaunay syndrome (KTS) is a severe congenital disorder characterized by capillary malformations, venous malformations or varicose veins, and hypertrophy of the affected tissues. The angiogenic factor gene AGGF1 was previously identified as a candidate susceptibility gene for KTS, but further genetic studies are needed to firmly establish the genetic relationship between AGGF1 and KTS. We analyzed HapMap data and identified two tagSNPs, rs13155212 and rs7704267 that capture information for all common variants in AGGF1 . The two SNPs were genotyped in 173 Caucasian KTS patients and 477 Caucasian non-KTS controls, and both significantly associated with susceptibility for KTS ( P = 0.004 and 0.013, respectively). Permutation testing also showed a significant empirical P value for the association (empirical P = 0.006 and 0.015, respectively). To control for potential confounding due to population stratification, the population structure for both cases and controls was characterized by genotyping of 38 ancestry-informative markers (AIMs) and the STRUCTURE program. The association between the AGGF1 SNPs and KTS remained significant after multivariate analysis by incorporating the inferred cluster scores as a covariate or after removal of outlier individuals identified by STRUCTURE. These results suggest that common AGGF1 variants confer risk of KTS.     

Genome‐Wide Association Study of Pre‐Eclampsia Detects Novel Maternal Single Nucleotide Polymorphisms and Copy‐Number Variants in Subsets of the Hyperglycemia and Adverse Pregnancy Outcome (HAPO) Study Cohort

A genome‐wide association study was undertaken to identify maternal single nucleotide polymorphisms (SNPs) and copy‐number variants (CNVs) associated with pre‐eclampsia. Case‐control analysis was performed on 1070 Afro‐Caribbean (n = 21 cases and 1049 controls) and 723 Hispanic (n = 62 cases and 661 controls) mothers and 1257 mothers of European ancestry (n = 50 cases and 1207 controls) from the Hyperglycemia and Adverse Pregnancy Outcome (HAPO) study. European ancestry subjects were genotyped on Illumina Human610‐Quad and Afro‐Caribbean and Hispanic subjects were genotyped on Illumina Human1M‐Duo BeadChip microarrays. Genome‐wide SNP data were analyzed using PLINK. CNVs were called using three detection algorithms (GNOSIS, PennCNV, and QuantiSNP), merged using CNVision, and then screened using stringent criteria. SNP and CNV findings were compared to those of the Study of Pregnancy Hypertension in Iowa (SOPHIA), an independent pre‐eclampsia case‐control dataset of Caucasian mothers (n = 177 cases and 116 controls). A list of top SNPs were identified for each of the HAPO ethnic groups, but none reached Bonferroni‐corrected significance. Novel candidate CNVs showing enrichment among pre‐eclampsia cases were also identified in each of the three ethnic groups. Several variants were suggestively replicated in SOPHIA. The discovered SNPs and copy‐number variable regions present interesting candidate genetic variants for pre‐eclampsia that warrant further replication and investigation.     

Association of SNP rs17465637 on chromosome 1q41 and rs599839 on 1p13.3 with myocardial infarction in an American caucasian population

Recent genome-wide single nucleotide polymorphism (SNP) association studies (GWAS) have identified a number of SNPs that were significantly associated with coronary artery disease and myocardial infarction (MI). However, many independent replication studies in other populations are needed to unequivocally confirm the GWAS association. To assess GWAS association, we have established a case-control cohort consisting of 1231 well-characterised MI patients and 560 controls without detectable coronary stenosis, all selected from the Cleveland Genebank population. The Genebank cohort has sufficient power to detect the association between MI and four GWAS SNPs, including rs17465637 within the MIA3 gene, rs2943634 (intergenic), rs6922269 in MTHFD1L, and rs599839 near SORT1. SNPs were genotyped by TaqMan assays and follow-up multivariate logistic regression analysis with incorporation of significant covariates showed significant association with MI for MIA3 SNP rs17465637 (P-adj= 0.0034) and SORT1 SNP rs599839 (P-adj= 0.009). The minor allele G of rs599839 was also associated with a decreased LDL-C level of 5-9 mg/dL per allele, but not with HDL-C or triglyceride levels. No association for MI or lipid levels was found for SNPs rs2943634 and rs6922269 (P-adj > 0.05). Our results establish two SNPs, rs17465637 in MIA3 and rs599839 near SORT1 as significant risk factors for MI in the American Genebank Caucasian population.     

OX40基因rs2298212位点与汉族人群冠状动脉粥样硬化疾病的关联研究

目的 探讨OX40基因(TNFRSF4)rs2298212G/A位点与山东汉族人群冠状动脉粥样硬化疾病的相关性.方法 在山东大学齐鲁医院心内科和健康体检中心分别收集到冠状动脉粥样硬化疾病患者536例和正常对照544名,采用聚合酶链反应-限制性片段长度多态性方法 对OX40基因rs2298212G/A多态性位点进行基因分型,并对数据进行统计分析.结果 基因型与等位基因频率分布在病例组与对照组之间差异均无统计学意义(P＞0.05).在回归校正了年龄、性别、体重指数、收缩压、舒张压、血糖、总胆固醇及甘油三酯等因素的影响后,基因型频率分布差异仍无统计学意义(P＞0.05).在对冠状动脉受累支数进行的分层分析发现,受累1支与受累3支之间,基因型与等位基因频率分布差异均有统计学意义(P＜0.05).结论 OX40基因rs2298212G/A多态位点同山东汉族人群冠状动脉粥样硬化疾病之间无关联性存在,但该位点可能与冠状动脉粥样硬化的严重程度相关.     

Association and Expression Study of <Emphasis Type="Italic">PRKCH</Emphasis> Gene in a French Caucasian Population with Rheumatoid Arthritis

We study the association between three protein kinase C, eta gene polymorphisms (+8134C/T, rs912620, rs959728), and susceptibility to rheumatoid arthritis. One hundred French Caucasian rheumatoid arthritis trio families were genotyped. Relative quantification of protein kinase C, eta mRNA expression was performed from whole blood in 24 unrelated rheumatoid arthritis patients and in 16 healthy controls. Our results showed no significant association or linkage between the protein kinase C, eta polymorphisms, and rheumatoid arthritis. The protein kinase C, eta mRNA was expressed at lower level in rheumatoid arthritis unrelated patients than in healthy controls. This study shows that protein kinase C, eta gene is not a Rheumatoid Arthritis major susceptibility genetic factor in the French Caucasian population. Furthermore, the lower expression of this gene in rheumatoid arthritis patients comparing to healthy controls suggests that protein kinase C, eta could be associated with the patho-physiologic mechanism of rheumatoid arthritis.     

Linkage and Association Study of Late‐Onset Alzheimer Disease Families Linked to 9p21.3

A chromosomal locus for late‐onset Alzheimer disease (LOAD) has previously been mapped to 9p21.3. The most significant results were reported in a sample of autopsy‐confirmed families. Linkage to this locus has been independently confirmed in AD families from a consanguineous Israeli‐Arab community. In the present study we analyzed an expanded clinical sample of 674 late‐onset AD families, independently ascertained by three different consortia. Sample subsets were stratified by site and autopsy‐confirmation. Linkage analysis of a dense array of SNPs across the chromosomal locus revealed the most significant results in the 166 autopsy‐confirmed families of the NIMH sample. Peak HLOD scores of 4.95 at D9S741 and 2.81 at the nearby SNP rs2772677 were obtained in a dominant model. The linked region included the cyclin‐dependent kinase inhibitor 2A gene (CDKN2A), which has been suggested as an AD candidate gene. By re‐sequencing all exons in the vicinity of CDKN2A in 48 AD cases, we identified and genotyped four novel SNPs, including a non‐synonymous, a synonymous, and two variations located in untranslated RNA sequences. Family‐based allelic and genotypic association analysis yielded significant results in CDKN2A (rs11515: PDT p = 0.003, genotype‐PDT p = 0.014). We conclude that CDKN2A is a promising new candidate gene potentially contributing to AD susceptibility on chromosome 9p.     

The Impact of Incomplete Linkage Disequilibrium and Genetic Model Choice on the Analysis and Interpretation of Genome‐wide Association Studies

When conducting a genetic association study, it has previously been observed that a multiplicative risk model tends to fit better at a disease‐associated marker locus than at the ungenotyped causative locus. This suggests that, while overall risk decreases as linkage disequilibrium breaks down, non‐multiplicative components are more affected. This effect is investigated here, in particular the practical consequences it has on testing for trait/marker associations and the estimation of mode of inheritance and risk once an associated locus has been found. The extreme significance levels required for genome‐wide association studies define a restricted range of detectable allele frequencies and effect sizes. For such parameters there is little to be gained by using a test that models the correct mode of inheritance rather than the multiplicative; thus the Cochran‐Armitage trend test, which assumes a multiplicative model, is preferable to a more general model as it uses fewer degrees of freedom. Equally when estimating risk, it is likely that a multiplicative risk model will provide a good fit to the data, regardless of the underlying mode of inheritance at the true susceptibility locus. This may lead to problems in interpreting risk estimates.     

Association between the COMT locus and obsessive‐compulsive disorder in females but not males

A polymorphism in the coding region of catechol‐O‐methyltransferase gene (COMT) was previously reported to be associated with obsessive‐compulsive disorder (OCD), particularly in male probands. We attempted to replicate the previous finding using a family‐based genetic design in haplotype relative risk (HRR) and transmission disequilibrium (TDT) analyses. Fifty‐six OCD probands and their parents were genotyped for the COMT locus using established methods. Analysis of allele and genotype frequencies between the proband genotypes and the control (parental nontransmitted) genotypes failed to replicate the previous finding of gender divergence, gave no evidence of overall association, nor was linkage detected by TDT. However, further analysis of the COMT allele frequencies by proband gender gave evidence of a mildly significant association with the low‐activity COMT allele in female probands (P = 0.049), but not in male probands. These findings indicate that COMT may be etiologically relevant to OCD in a gender‐specific manner opposite to that shown in previous studies. © 2001 Wiley‐Liss, Inc.     

Association between interleukin-1 receptor antagonist gene and asthma-related traits in a German adult population

Background:  A recent study in German and Italian families associated variants in the interleukin-1 receptor antagonist (IL1RA) gene with asthma. The aim of the present study was to further investigate the role of single nucleotide polymorphisms (SNPs) in the IL1RA gene in the development of atopy and lifelong asthma in a population-based study.
Methods:  DNA samples from the German centres of the European Community Respiratory Health Survey were analysed for genetic variants in the IL1RA gene and the development of asthma, atopy and bronchial hyperreactivity.
Results:  Carriers of the rare G allele of SNP rs447713 had a significantly increased risk of developing asthma ( P  = 0.0013) and allergic sensitization ( P  = 0.0119). Carriers of the rare C allele of SNP rs3087271 had an increased risk of asthma ( P  = 0.0227) and high immunoglobulin E (IgE) levels ( P  = 0.0232). A haplotype built from eight SNPs in the IL1RA gene (A-C-A-G-A-C-G-A) was associated with a higher prevalence of asthma ( P  = 0.007) and high total IgE ( P  = 0.02). Bronchial hyperreactivity was positively associated with the haplotype A-C-G-G-A-C-G-C ( P  = 0.02) and negatively with the A-C-G-G-A-C-T-C ( P  = 0.03).
Conclusion:  A previously described association between IL1RA and asthma in families could be reproduced in a population-based sample. The genetic variants of IL1RA gene do not to seem to affect asthma alone, but to act as modulators of asthma-related traits as well, where different haplotypes drive the development of different phenotypes.