From GWAS to the clinic: risk factors for intracranial aneurysms

Subarachnoid hemorrhage (SAH) from a ruptured intracranial aneurysm is a devastating subset of stroke, occurring in relatively young people (mean age around 50 years) of whom around a third die within the initial weeks after the bleed. Environmental and genetic risk factors both have a role in SAH. A recent genome-wide association study of intracranial aneurysms in Finnish, Dutch and Japanese cohorts totaling 5,891 cases and 14,181 controls identified three new loci strongly associated with intracranial aneurysms on chromosomes 18q11.2 and 10q24.32, and replicated two previously found loci on chromosomes 8q11.23-q12.1 and 9p21.3. However, these five intracranial aneurysm risk loci identified so far explain only up to 5% of the familial risk of intracranial aneurysms, which makes genetic risk prediction tests currently unfeasible for intracranial aneurysms. New approaches, including identification of causal variants, rare variants and copy number variants, such as insertions and deletions, may improve genetic risk prediction for SAH and intracranial aneurysms. This may lead to diagnostic tools for identifying individuals at increased risk for aneurysm formation and rupture of aneurysms. In this way, genetic diagnostic tools will identify the people who will benefit most from screening by imaging studies for aneurysms and those who are most likely to benefit from preventive treatment of incidentally discovered aneurysms

Shared Genetic Risk Factors of Intracranial, Abdominal, and Thoracic Aneurysms

Background Intracranial aneurysms (IAs), abdominal aortic aneurysms (AAAs), and thoracic aortic aneurysms (TAAs) all have a familial predisposition. Given that aneurysm types are known to co‐occur, we hypothesized that there may be shared genetic risk factors for IAs, AAAs, and TAAs. Methods and Results We performed a mega‐analysis of 1000 Genomes Project‐imputed genome‐wide association study (GWAS) data of 4 previously published aneurysm cohorts: 2 IA cohorts (in total 1516 cases, 4305 controls), 1 AAA cohort (818 cases, 3004 controls), and 1 TAA cohort (760 cases, 2212 controls), and observed associations of 4 known IA, AAA, and/or TAA risk loci (9p21, 18q11, 15q21, and 2q33) with consistent effect directions in all 4 cohorts. We calculated polygenic scores based on IA‐, AAA‐, and TAA‐associated SNPs and tested these scores for association to case‐control status in the other aneurysm cohorts; this revealed no shared polygenic effects. Similarly, linkage disequilibrium–score regression analyses did not show significant correlations between any pair of aneurysm subtypes. Last, we evaluated the evidence for 14 previously published aneurysm risk single‐nucleotide polymorphisms through collaboration in extended aneurysm cohorts, with a total of 6548 cases and 16 843 controls (IA) and 4391 cases and 37 904 controls (AAA), and found nominally significant associations for IA risk locus 18q11 near RBBP8 to AAA (odds ratio [OR]=1.11; P=4.1×10−5) and for TAA risk locus 15q21 near FBN1 to AAA (OR=1.07; P=1.1×10−3). Conclusions Although there was no evidence for polygenic overlap between IAs, AAAs, and TAAs, we found nominally significant effects of two established risk loci for IAs and TAAs in AAAs. These two loci will require further replicatio

Complement C5 contributes to brain injury after subarachnoid hemorrhage

Previous studies showed that complement activation is associated with poor functional outcome after aneurysmal subarachnoid hemorrhage (SAH). We investigated whether complement activation is underlying brain injury after aneurysmal SAH (n = 7) and if it is an appropriate treatment target. We investigated complement expression in brain tissue of aneurysmal SAH patients (n = 930) and studied the role of common genetic variants in C3 and C5 genes in outcome. We analyzed plasma levels (n = 229) to identify the functionality of a single nucleotide polymorphism (SNP) associated with outcome. The time course of C5a levels was measured in plasma (n = 31) and CSF (n = 10). In an SAH mouse model, we studied the extent of microglia activation and cell death in wild-type mice, mice lacking the C5a receptor, and in mice treated with C5-specific antibodies (n = 15 per group). Brain sections from aneurysmal SAH patients showed increased presence of complement components C1q and C3/C3b/iC3B compared to controls. The complement component 5 (C5) SNP correlated with C5a plasma levels and poor disease outcome. Serial measurements in CSF revealed that C5a was > 1400-fold increased 1 day after aneurysmal SAH and then gradually decreased. C5a in plasma was 2-fold increased at days 3–10 after aneurysmal SAH. In the SAH mouse model, we observed a ≈ 40% reduction in both microglia activation and cell death in mice lacking the C5a receptor, and in mice treated with C5-specific antibodies. These data show that C5 contributes to brain injury after experimental SAH, and support further study of C5-specific antibodies as novel treatment option to reduce brain injury and improve prognosis after aneurysmal SAH

Distribution of Cardioembolic Stroke:A Cohort Study

Background: A cardiac origin in ischemic stroke is more frequent than previously assumed, but it is not clear which patients benefit from cardiac work-up if obvious cardiac pathology is absent. We hypothesized that thromboembolic stroke with a cardiac source occurs more frequently in the posterior circulation compared with thromboembolic stroke of another etiology. Methods: We performed a multicenter observational study in 3,311 consecutive patients with ischemic stroke who were enrolled in an ongoing prospective stroke registry of 8 University hospitals between September 2009 and November 2014 in The Netherlands. In this initiative, the so-called Parelsnoer Institute-Cerebrovascular Accident Study Group, clinical data, imaging, and biomaterials of patients with stroke are prospectively and uniformly collected. We compared the proportions of posterior stroke location in patients with a cardiac stroke source with those with another stroke etiology and calculated risk ratios (RR) with corresponding 95% CI with Poisson regression analyses. To assess which patient or disease characteristics were most strongly associated with a cardiac etiology in patients with ischemic stroke, we performed a stepwise backward regression analysis. Results: For the primary aim, 1,428 patients were eligible for analyses. The proportion of patients with a posterior stroke location among patients with a cardiac origin of their stroke (28%) did not differ statistically significant to those with another origin (25%), age and sex adjusted RR 1.16; 95% CI 0.96-1.41. For the secondary aim, 1,955 patients were eligible for analyses. No recent history of smoking, no hyperlipidemia, coronary artery disease, a higher age, and a higher National Institutes of Health Stroke Scale (NIHSS) score were associated with a cardiac etiology of ischemic stroke. Conclusions: We could not confirm our hypothesis that thromboembolic stroke localized in the posterior circulation is associated with a cardioembolic source of ischemic stroke, and therefore posterior stroke localization on itself does not necessitate additional cardiac examination. The lack of determinants of atherosclerosis, for example, no recent history of smoking and no hyperlipidemia, coronary artery disease, a higher age, and a higher NIHSS score are stronger risk factors for a cardiac source of ischemic stroke

Exome-chip association analysis of intracranial aneurysms

Objective: To investigate to what extent low-frequency genetic variants (with minor allele frequencies <5%) affect the risk of intracranial aneurysms (IAs). Methods: One thousand fifty-six patients with IA and 2,097 population-based controls from the Netherlands were genotyped with the Illumina HumanExome BeadChip. After quality control (QC) of samples and single nucleotide variants (SNVs), we conducted a single variant analysis using the Fisher exact test. We also performed the variable threshold (VT) test and the sequence kernel association test (SKAT) at different minor allele count (MAC) thresholds of >5 and >0 to test the hypothesis that multiple variants within the same gene are associated with IA risk. Significant results were tested in a replication cohort of 425 patients with IA and 311 controls, and results of the 2 cohorts were combined in a meta-analysis. Results: After QC, 995 patients with IA and 2,080 controls remained for further analysis. The single variant analysis comprising 46,534 SNVs did not identify significant loci at the genome-wide level. The gene-based tests showed a statistically significant association for fibulin 2 (FBLN2) (best p = 1 × 10-6 for the VT test, MAC >5). Associations were not statistically significant in the independent but smaller replication cohort (p > 0.57) but became slightly stronger in a meta-analysis of the 2 cohorts (best p = 4.8 × 10-7 for the SKAT, MAC ≥1). Conclusion: Gene-based tests indicated an association for FBLN2, a gene encoding an extracellular matrix protein implicated in vascular wall remodeling, but independent validation in larger cohorts is warranted. We did not identify any significant associations for single low-frequency genetic variants

Difference in Rupture Risk Between Familial and Sporadic Intracranial Aneurysms An Individual Patient Data Meta-analysis

OBJECTIVE: We combined individual patient data (IPD) from prospective cohorts of patients with unruptured intracranial aneurysms (UIA) to assess to what extent patients with familial UIA have a higher rupture risk than those with sporadic UIA. METHODS: For this IPD meta-analysis we performed an Embase and Pubmed search for studies published up to December 1, 2020. We included studies that 1) had a prospective study design; 2) included 50 or more patients with UIA; 3) studied the natural course of UIA and risk factors for aneurysm rupture including family history for aneurysmal subarachnoid haemorrhage and UIA; and 4) had aneurysm rupture as an outcome. Cohorts with available IPD were included. All studies included patients with newly diagnosed UIA visiting one of the study centers. The primary outcome was aneurysmal rupture. Patients with polycystic kidney disease and moyamoya disease were excluded. We compared rupture rates of familial versus sporadic UIA using a Cox proportional hazard regression model adjusted for the PHASES score and smoking. We performed two analyses: 1. only studies defining first-degree relatives as parents, children, and siblings and 2. all studies, including those in which first-degree relatives are defined as only parents and children, but not siblings. RESULTS: We pooled IPD from eight cohorts with a low and moderate risk of bias. First-degree relatives were defined as parents, siblings and children in six cohorts (29% Dutch, 55% Finnish, 15% Japanese), totalling 2,297 patients (17% familial, 399 patients) with 3,089 UIA and 7,301 person-years follow-up. Rupture occurred in 10 familial patients (rupture rate: 0·89%/person-year; 95% CI:0·45-1·59) and 41 sporadic patients (0·66%/person-year; 95% CI:0·48-0·89); adjusted HR for familial patients 2·56 (95% CI: 1·18-5·56). After adding also the two cohorts excluding siblings as first-degree relatives resulting in 9,511 patients the adjusted HR was 1·44 (95% CI: 0·86-2·40). CONCLUSION: The risk of rupture of UIA is two and a half times higher, with a range from a 1.2 to 5 times higher risk, in familial than in sporadic UIA. When assessing the risk of rupture in UIA, family history should be taken into account

Distribution of Cardioembolic Stroke: A Cohort Study

Background: A cardiac origin in ischemic stroke is more frequent than previously assumed, but it is not clear which patients benefit from cardiac work-up if obvious cardiac pathology is absent. We hypothesized that thromboembolic stroke with a cardiac source occurs more frequently in the posterior circulation compared with thromboembolic stroke of another etiology. Methods: We performed a multicenter observational study in 3,311 consecutive patients with ischemic stroke who were enrolled in an ongoing prospective stroke registry of 8 University hospitals between September 2009 and November 2014 in The Netherlands. In thi

Sex Difference and Rupture Rate of Intracranial Aneurysms : An Individual Patient Data Meta-Analysis

Background and Purpose: In previous studies, women had a higher risk of rupture of intracranial aneurysms than men, but female sex was not an independent risk factor. This may be explained by a higher prevalence of patient- or aneurysm-related risk factors for rupture in women than in men or by insufficient power of previous studies. We assessed sex differences in rupture rate taking into account other patient- and aneurysm-related risk factors for aneurysmal rupture. Methods: We searched Embase and Pubmed for articles published until December 1, 2020. Cohorts with available individual patient data were included in our meta-analysis. We compared rupture rates of women versus men using a Cox proportional hazard regression model adjusted for the PHASES score (Population, Hypertension, Age, Size of Aneurysm, Earlier Subarachnoid Hemorrhage From Another Aneurysm, Site of Aneurysm), smoking, and a positive family history of aneurysmal subarachnoid hemorrhage. Results: We pooled individual patient data from 9 cohorts totaling 9940 patients (6555 women, 66%) with 12 193 unruptured intracranial aneurysms, and 24 357 person-years follow-up. Rupture occurred in 163 women (rupture rate 1.04%/person-years [95% CI, 0.89-1.21]) and 63 men (rupture rate 0.74%/person-years [95% CI, 0.58-0.94]). Women were older (61.9 versus 59.5 years), were less often smokers (20% versus 44%), more often had internal carotid artery aneurysms (24% versus 17%), and larger sized aneurysms (>= 7 mm, 24% versus 23%) than men. The unadjusted women-to-men hazard ratio was 1.43 (95% CI, 1.07-1.93) and the adjusted women/men ratio was 1.39 (95% CI, 1.02-1.90). Conclusions: Women have a higher risk of aneurysmal rupture than men and this sex difference is not explained by differences in patient- and aneurysm-related risk factors for aneurysmal rupture. Future studies should focus on the factors explaining the higher risk of aneurysmal rupture in women.Peer reviewe

Genes influencing coagulation and the risk of aneurysmal subarachnoid hemorrhage, and subsequent complications of secondary cerebral ischemia and rebleeding

We investigated whether genes influencing coagulation are associated with the occurrence of aneurysmal subarachnoid hemorrhage (SAH) and with secondary cerebral ischemia and rebleeding in patients with aneurysmal SAH. Genotyping for factor V Leiden (G1691A), prothrombin G20210A, methylenetetetrahydrofolate reductase (MTHFR) C677T, factor XIII subunit A Val34Leu, Tyr204Phe and Pro564Leu, and factor XIII subunit B His95Arg was performed in 208 patients with aneurysmal SAH and in 925 controls. Secondary cerebral ischemia occurred in 49 (24%) patients and rebleeding in 28 (14%) during their clinical course of 3 months after the aneurysmal SAH. The risk of aneurysmal SAH was assessed as odds ratio (OR) with 95% confidence interval (95% CI). The risk of secondary cerebral ischemia and rebleeding was assessed as hazard ratio (HR) with 95% CI using Cox regression. Carriers of the subunit B His95Arg factor XIII polymorphism had an increased risk of aneurysmal SAH with 23% of the patients homozygous or heterozygous for the variant allele compared to 17% of control subjects (OR 1.5, 95% CI 1.0-2.2). For the remaining genetic variants no effect on the risk of aneurysmal SAH could be demonstrated. A clear relation with the risk of secondary cerebral ischemia and of rebleeding could not be established for any of the genetic variants. We found that aneurysmal SAH patients are more often carriers of the subunit B His95Arg factor XIII polymorphism compared to controls. This suggests that carriers of the subunit B His95Arg factor XIII polymorphism have an increased risk of aneurysmal SAH. Larger studies should confirm our results. As aneurysmal SAH patients who died soon after admission could not be included in the present study, our results only apply to a population of patients who survived the initial hours after the hemorrhage. For the other studied genetic factors involved in coagulation, no association with the occurrence of aneurysmal SAH or with the occurrence of secondary cerebral ischemia or rebleeding after aneurysmal SAH could be demonstrated

Genetic Risk Score for Intracranial Aneurysms:Prediction of Subarachnoid Hemorrhage and Role in Clinical Heterogeneity

BACKGROUND: Recently, common genetic risk factors for intracranial aneurysm (IA) and aneurysmal subarachnoid hemorrhage (ASAH) were found to explain a large amount of disease heritability and therefore have potential to be used for genetic risk prediction. We constructed a genetic risk score to (1) predict ASAH incidence and IA presence (combined set of unruptured IA and ASAH) and (2) assess its association with patient characteristics. METHODS: A genetic risk score incorporating genetic association data for IA and 17 traits related to IA (so-called metaGRS) was created using 1161 IA cases and 407 392 controls from the UK Biobank population study. The metaGRS was validated in combination with risk factors blood pressure, sex, and smoking in 828 IA cases and 68 568 controls from the Nordic HUNT population study. Furthermore, we assessed association between the metaGRS and patient characteristics in a cohort of 5560 IA patients. RESULTS: Per SD increase of metaGRS, the hazard ratio for ASAH incidence was 1.34 (95% CI, 1.20-1.51) and the odds ratio for IA presence 1.09 (95% CI, 1.01-1.18). Upon including the metaGRS on top of clinical risk factors, the concordance index to predict ASAH hazard increased from 0.63 (95% CI, 0.59-0.67) to 0.65 (95% CI, 0.62-0.69), while prediction of IA presence did not improve. The metaGRS was statistically significantly associated with age at ASAH (β=-4.82×10(-3) per year [95% CI, -6.49×10(-3) to -3.14×10(-3)]; P=1.82×10(-8)), and location of IA at the internal carotid artery (odds ratio=0.92 [95% CI, 0.86-0.98]; P=0.0041). CONCLUSIONS: The metaGRS was predictive of ASAH incidence, although with limited added value over clinical risk factors. The metaGRS was not predictive of IA presence. Therefore, we do not recommend using this metaGRS in daily clinical care. Genetic risk does partly explain the clinical heterogeneity of IA warranting prioritization of clinical heterogeneity in future genetic prediction studies of IA and ASAH