Helena Schmidt

Genome-wide Analysis of Genetic Loci Associated With Alzheimer Disease

CONTEXT Genome-wide association studies (GWAS) have recently identified CLU, PICALM, and CR1 as novel genes for late-onset Alzheimer disease (AD). OBJECTIVES To identify and strengthen additional loci associated with AD and confirm these in an independent sample and to examine the contribution of recently identified genes to AD risk prediction in a 3-stage analysis of new and previously published GWAS on more than 35,000 persons (8371 AD cases). DESIGN, SETTING, AND PARTICIPANTS In stage 1, we identified strong genetic associations (P < 10(-3)) in a sample of 3006 AD cases and 14,642 controls by combining new data from the population-based Cohorts for Heart and Aging Research in Genomic Epidemiology consortium (1367 AD cases [973 incident]) with previously reported results from the Translational Genomics Research Institute and the Mayo AD GWAS. We identified 2708 single-nucleotide polymorphisms (SNPs) with P < 10(-3). In stage 2, we pooled results for these SNPs with the European AD Initiative (2032 cases and 5328 controls) to identify 38 SNPs (10 loci) with P < 10(-5). In stage 3, we combined data for these 10 loci with data from the Genetic and Environmental Risk in AD consortium (3333 cases and 6995 controls) to identify 4 SNPs with P < 1.7x10(-8). These 4 SNPs were replicated in an independent Spanish sample (1140 AD cases and 1209 controls). Genome-wide association analyses were completed in 2007-2008 and the meta-analyses and replication in 2009. MAIN OUTCOME MEASURE Presence of Alzheimer disease. RESULTS Two loci were identified to have genome-wide significance for the first time: rs744373 near BIN1 (odds ratio [OR],1.13; 95% confidence interval [CI],1.06-1.21 per copy of the minor allele; P = 1.59x10(-11)) and rs597668 near EXOC3L2/BLOC1S3/MARK4 (OR, 1.18; 95% CI, 1.07-1.29; P = 6.45x10(-9)). Associations of these 2 loci plus the previously identified loci CLU and PICALM with AD were confirmed in the Spanish sample (P < .05). However, although CLU and PICALM were confirmed to be associated with AD in this independent sample, they did not improve the ability of a model that included age, sex, and APOE to predict incident AD (improvement in area under the receiver operating characteristic curve from 0.847 to 0.849 in the Rotterdam Study and 0.702 to 0.705 in the Cardiovascular Health Study). CONCLUSIONS Two genetic loci for AD were found for the first time to reach genome-wide statistical significance. These findings were replicated in an independent population. Two recently reported associations were also confirmed. These loci did not improve AD risk prediction. While not clinically useful, they may implicate biological pathways useful for future research.

Genetic risk factors for ischaemic stroke and its subtypes (the METASTROKE collaboration): a meta-analysis of genome-wide association studies.

Summary Background Various genome-wide association studies (GWAS) have been done in ischaemic stroke, identifying a few loci associated with the disease, but sample sizes have been 3500 cases or less. We established the METASTROKE collaboration with the aim of validating associations from previous GWAS and identifying novel genetic associations through meta-analysis of GWAS datasets for ischaemic stroke and its subtypes. Methods We meta-analysed data from 15 ischaemic stroke cohorts with a total of 12 389 individuals with ischaemic stroke and 62 004 controls, all of European ancestry. For the associations reaching genome-wide significance in METASTROKE, we did a further analysis, conditioning on the lead single nucleotide polymorphism in every associated region. Replication of novel suggestive signals was done in 13 347 cases and 29 083 controls. Findings We verified previous associations for cardioembolic stroke near PITX2 (p=2·8×10−16) and ZFHX3 (p=2·28×10−8), and for large-vessel stroke at a 9p21 locus (p=3·32×10−5) and HDAC9 (p=2·03×10−12). Additionally, we verified that all associations were subtype specific. Conditional analysis in the three regions for which the associations reached genome-wide significance (PITX2, ZFHX3, and HDAC9) indicated that all the signal in each region could be attributed to one risk haplotype. We also identified 12 potentially novel loci at p<5×10−6. However, we were unable to replicate any of these novel associations in the replication cohort. Interpretation Our results show that, although genetic variants can be detected in patients with ischaemic stroke when compared with controls, all associations we were able to confirm are specific to a stroke subtype. This finding has two implications. First, to maximise success of genetic studies in ischaemic stroke, detailed stroke subtyping is required. Second, different genetic pathophysiological mechanisms seem to be associated with different stroke subtypes. Funding Wellcome Trust, UK Medical Research Council (MRC), Australian National and Medical Health Research Council, National Institutes of Health (NIH) including National Heart, Lung and Blood Institute (NHLBI), the National Institute on Aging (NIA), the National Human Genome Research Institute (NHGRI), and the National Institute of Neurological Disorders and Stroke (NINDS).

Genetic variants associated with cardiac structure and function: A meta-analysis and replication of genome-wide association data

CONTEXT Echocardiographic measures of left ventricular (LV) structure and function are heritable phenotypes of cardiovascular disease. OBJECTIVE To identify common genetic variants associated with cardiac structure and function by conducting a meta-analysis of genome-wide association data in 5 population-based cohort studies (stage 1) with replication (stage 2) in 2 other community-based samples. DESIGN, SETTING, AND PARTICIPANTS Within each of 5 community-based cohorts comprising the EchoGen consortium (stage 1; n = 12 612 individuals of European ancestry; 55% women, aged 26-95 years; examinations between 1978-2008), we estimated the association between approximately 2.5 million single-nucleotide polymorphisms (SNPs; imputed to the HapMap CEU panel) and echocardiographic traits. In stage 2, SNPs significantly associated with traits in stage 1 were tested for association in 2 other cohorts (n = 4094 people of European ancestry). Using a prespecified P value threshold of 5 x 10(-7) to indicate genome-wide significance, we performed an inverse variance-weighted fixed-effects meta-analysis of genome-wide association data from each cohort. MAIN OUTCOME MEASURES Echocardiographic traits: LV mass, internal dimensions, wall thickness, systolic dysfunction, aortic root, and left atrial size. RESULTS In stage 1, 16 genetic loci were associated with 5 echocardiographic traits: 1 each with LV internal dimensions and systolic dysfunction, 3 each with LV mass and wall thickness, and 8 with aortic root size. In stage 2, 5 loci replicated (6q22 locus associated with LV diastolic dimensions, explaining <1% of trait variance; 5q23, 12p12, 12q14, and 17p13 associated with aortic root size, explaining 1%-3% of trait variance). CONCLUSIONS We identified 5 genetic loci harboring common variants that were associated with variation in LV diastolic dimensions and aortic root size, but such findings explained a very small proportion of variance. Further studies are required to replicate these findings, identify the causal variants at or near these loci, characterize their functional significance, and determine whether they are related to overt cardiovascular disease.

Apolipoprotein E ε4 is associated with rapid progression of multiple sclerosis

Objective: The apolipoprotein E (APOE) polymorphism is known to impact on various neurologic disorders and has differential effects on the immune system and on CNS repair. Previous findings concerning a possible modulation of the clinical course of MS have been inconsistent, however. Methods: In a cross-sectional study, the authors investigated 374 patients with clinically definite MS and a disease duration of at least 3 years and related their clinical and demographic findings to the allelic polymorphism of the APOE gene. The genotype distribution of patients with MS was compared with a cohort of 389 asymptomatic, randomly selected elderly volunteers. Results: The authors found no significant differences in the distribution of genotypes between patients with MS and controls. However, patients with MS with the ε4 allele (n = 85) had a significantly higher progression index of disability (0.46 ± 0.4 versus 0.33 ± 0.26; p < 0.004) and a worse ranked MS severity score (5.1 ± 1.9 versus 5.7 ± 1.7; p = 0.05) than their non-ε4 counterparts, despite significantly more frequent long-term immunotherapy in ε4 carriers (74% versus 58%; p < 0.007). The annual relapse rate in ε4 carriers (0.87 ± 0.56) was significantly higher than in patients with MS without an ε4 allele (0.71 ± 0.47; p = 0.03). Conclusions: These results suggest no effect of the APOE genotype on susceptibility to MS, but indicate an association of the APOE ε4 allele with a more severe course of the disease.

Common variants at 12q14 and 12q24 are associated with hippocampal volume

Aging is associated with reductions in hippocampal volume that are accelerated by Alzheimers disease and vascular risk factors. Our genome-wide association study (GWAS) of dementia-free persons (n = 9,232) identified 46 SNPs at four loci with P values of <4.0 × 10−7. In two additional samples (n = 2,318), associations were replicated at 12q14 within MSRB3-WIF1 (discovery and replication; rs17178006; P = 5.3 × 10−11) and at 12q24 near HRK-FBXW8 (rs7294919; P = 2.9 × 10−11). Remaining associations included one SNP at 2q24 within DPP4 (rs6741949; P = 2.9 × 10−7) and nine SNPs at 9p33 within ASTN2 (rs7852872; P = 1.0 × 10−7); along with the chromosome 12 associations, these loci were also associated with hippocampal volume (P < 0.05) in a third younger, more heterogeneous sample (n = 7,794). The SNP in ASTN2 also showed suggestive association with decline in cognition in a largely independent sample (n = 1,563). These associations implicate genes related to apoptosis (HRK), development (WIF1), oxidative stress (MSR3B), ubiquitination (FBXW8) and neuronal migration (ASTN2), as well as enzymes targeted by new diabetes medications (DPP4), indicating new genetic influences on hippocampal size and possibly the risk of cognitive decline and dementia.

Heterogeneity in age-related white matter changes

White matter changes occur endemically in routine magnetic resonance imaging (MRI) scans of elderly persons. MRI appearance and histopathological correlates of white matter changes are heterogeneous. Smooth periventricular hyperintensities, including caps around the ventricular horns, periventricular lining and halos are likely to be of non-vascular origin. They relate to a disruption of the ependymal lining with subependymal widening of the extracellular space and have to be differentiated from subcortical and deep white matter abnormalities. For the latter a distinction needs to be made between punctate, early confluent and confluent types. Although punctate white matter lesions often represent widened perivascular spaces without substantial ischemic tissue damage, early confluent and confluent lesions correspond to incomplete ischemic destruction. Punctate abnormalities on MRI show a low tendency for progression, while early confluent and confluent changes progress rapidly. The causative and modifying pathways involved in the occurrence of sporadic age-related white matter changes are still incompletely understood, but recent microarray and genome-wide association approaches increased the notion of pathways that might be considered as targets for therapeutic intervention. The majority of differentially regulated transcripts in white matter lesions encode genes associated with immune function, cell cycle, proteolysis, and ion transport. Genome-wide association studies identified six SNPs mapping to a locus on chromosome 17q25 to be related to white matter lesion load in the general population. We also report first and preliminary data that demonstrate apolipoprotein E (ApoE) immunoreactivity in white matter lesions and support epidemiological findings indicating that ApoE is another factor possibly related to white matter lesion occurrence. Further insights come from modern MRI techniques, such as diffusion tensor and magnetization transfer imaging, as they provide tools for the characterization of normal-appearing brain tissue beyond what can be expected from standard MRI scans. There is a need for additional pre- and postmortem studies in humans, including these new imaging techniques.

Genome-wide association studies of cerebral white matter lesion burden

White matter hyperintensities (WMHs) detectable by magnetic resonance imaging are part of the spectrum of vascular injury associated with aging of the brain and are thought to reflect ischemic damage to the small deep cerebral vessels. WMHs are associated with an increased risk of cognitive and motor dysfunction, dementia, depression, and stroke. Despite a significant heritability, few genetic loci influencing WMH burden have been identified.

Markers of Endothelial and Hemostatic Activation and Progression of Cerebral White Matter Hyperintensities. Longitudinal Results of the Austrian Stroke Prevention Study

Background and Purpose— The pathogenesis of cerebral small vessel disease (SVD) is poorly understood, but endothelial activation and dysfunction may play a causal role. Cross-sectional studies have found increased circulating markers of endothelial activation, but this study design cannot exclude causality from secondary elevations. Confluent white matter hyperintensities (WMHs) on magnetic resonance imaging (MRI) appear to represent asymptomatic cerebral SVD. In a prospective study, we determined whether circulating markers of endothelial activation predicted progression of WMH. Methods— In the community-based Austrian Stroke Prevention Study, MRI was performed at baseline in 296 subjects and repeated at 3 and 6 years. The following were measured on baseline plasma samples: intercellular adhesion molecule (ICAM), thrombomodulin, tissue factor plasma inhibitor, prothrombin fragments 1 and 2, and D-dimers. Results— ICAM was associated with age- and gender-adjusted WMH lesion progression at both 3 and 6 years, respectively; (odds ratio [OR], 1.007; 95% confidence interval [CI], 1.002 to 1.012; P=0.004; and OR, 1.004; 95% CI, 1.000 to 1.009 per ng/mL; P=0.057). After multivariate analysis controlling for other cardiovascular risk factors and C-reactive protein, 3-year OR was 1.010 (95% CI, 1.004 to 1.017; P=0.001) and 6-year OR was 1.008 (1.002 to 1.014 per ng/mL; P=0.006). Baseline log lesion volume was a strong independent predictor of progression but associations remained after controlling for this (3-year OR, 1.011; 95% CI, 1.002 to 1.020; P=0.013; and 6-year OR, 1.009; 95% CI, 1.000 to 1.017; P=0.039 per ng/mL). There was no association between WMH progression and other markers. Conclusions— ICAM levels are related to progression of WMH on MRI. The prospective study design increases the likelihood that this association is causal and supports a role of endothelial cell activation in disease pathogenesis. In contrast, we found no evidence for coagulation activation being important.

APOE epsilon variation in multiple sclerosis susceptibility and disease severity: some answers.

Background: Previous studies have examined the role of APOE variation in multiple sclerosis (MS), but have lacked the statistical power to detect modest genetic influences on risk and disease severity. The meta- and pooled analyses presented here utilize the largest collection, to date, of MS cases, controls, and families genotyped for the APOE epsilon polymorphism. Methods: Studies of MS and APOE were identified by searches of PubMed, Biosis, Web of Science, Cochrane Review, and Embase. When possible, authors were contacted for individual genotype data. Meta-analyses of MS case-control data and family-based analyses were performed to assess the association of APOE epsilon genotype with disease risk. Pooled analyses of MS cases were also performed to assess the influence of APOE epsilon genotype on disease severity. Results: A total of 22 studies (3,299 MS cases and 2,532 controls) were available for meta-analysis. No effect of ε2 or ε4 status on MS risk was observed (summary OR 1.14, 95% CI 0.96–1.34 and OR 0.89, 95% CI 0.78–1.01). Results obtained from analyses of APOE genotypes in 1,279 MS families were also negative (p = 0.61). Finally, results from pooled analyses of 4,048 MS cases also argue strongly that APOE epsilon status does not distinguish a relapsing-remitting from primary progressive disease course, or influence disease severity, as measured by the Expanded Disability Status Scale and disease duration. Conclusion: Overall, these findings do not support a role for APOE in multiple sclerosis, and underscore the importance of using large sample sizes to detect modest genetic effects, particularly in studies of genotype-phenotype relationships.

C-Reactive Protein, Carotid Atherosclerosis, and Cerebral Small-Vessel Disease Results of the Austrian Stroke Prevention Study

Background and Purpose— C-reactive protein (CRP) is an inflammatory marker known to be a risk factor for stroke. We examined the associations between CRP, carotid atherosclerosis, white matter lesions, and lacunes as manifestations of cerebral large- and small-vessel disease. Methods— In the community-based Austrian Stroke Prevention Study, CRP concentrations were measured by a highly sensitive assay in 700 participants at baseline. All underwent carotid duplex scanning, and a subset of 505 subjects underwent brain magnetic resonance imaging. Imaging was repeated after 3 and 6 years. We graded carotid atherosclerosis in both common and internal carotid arteries on a 5-point scale and calculated the sum of scores as an index of the severity of carotid atherosclerosis. The volume of white matter lesions and the number of lacunes were considered small vessel disease–related brain abnormalities. Results— After adjustment for vascular risk factors, the severity and progression of extracranial carotid atherosclerosis increased with increasing quintiles of CRP. Only study participants in the fourth and fifth quintile (>2.50 mg/L) had significantly more baseline atherosclerosis and greater progression when we used the first quintile (<0.80 mg/L) as a reference. No interactions were seen between CRP quintiles and vascular risk factors for carotid atherosclerosis. The associations between severity and progression of small vessel disease–related brain abnormalities and CRP were nonsignificant. Conclusions— We found evidence for differential effects of CRP in different beds of the arterial brain supply. CRP was a marker for active carotid atherosclerosis but not for small vessel disease–related brain lesions.