Protocols The Dutch String-of-Pearls Stroke Study: Protocol of a large prospective multicenter genetic cohort study Paul J. Nederkoorn1, Ewoud J. van Dijk2, Peter J. Koudstaal3, Gert-Jan Luijckx4, Robert J. van Oostenbrugge5, Marieke C. Visser6, Marieke J. H. Wermer7, Ynte M. Ruigrok8, Ale Algra9,10, and L. Jaap Kappelle8* on behalf of the Dutch String-of-Pearls Stroke Study-Group† Correspondence: L. Jaap Kappelle*, Department of Neurology, University Medical Center Utrecht, Heidelberglaan 100, Utrecht 3584 GR, The Netherlands. E-mail: [email protected] 1 Department of Neurology, Academic Medical Center, Amsterdam, The Netherlands 2 Department of Neurology, Radboud University Medical Center, Nijmegen, The Netherlands 3 Department of Neurology, Erasmus Medical Center, Rotterdam, The Netherlands 4 Department of Neurology, University Medical Center, Groningen, The Netherlands 5 Department of Neurology, Maastricht University Medical Center, Amsterdam, The Netherlands 6 Department of Neurology, VU Medical Center, Amsterdam, The Netherlands 7 Department of Neurology, Leiden University Medical Center, Utrecht, The Netherlands 8 Department of Neurology, University Medical Center Utrecht, Utrecht, The Netherlands 9 Julius Center for Clinical Research and Patient Care, University Medical Center Utrecht, Utrecht, The Netherlands 10 Department of Clinical Epidemiology, Leiden University Medical Center, Leiden, The Netherlands Received: 26 February 2014; Accepted: 30 June 2014
Background In the last couple of years, genome-wide association studies have largely altered the scope in genetic research in diseases in which both environmental and genetic risk factors contribute to the disease. To date, the genetic risk loci identified in stroke have lagged behind those in other complex diseases, possibly because of the heterogeneity of stroke phenotypes. Sufficiently large cohorts with well-defined and detailed phenotyping of stroke patients are needed to identify additional genetic risk loci. Design The String-of-Pearls Institute is a unique partnership between all eight University Medical Centers in the Netherlands. It was established in 2007 by the Netherlands Federation of University Medical Centers, and it conducts a large prospective cohort study in which comprehensive clinical data, detailed phenotyping of stroke, imaging data, and biomaterials are collected in a large cohort of stroke patients. Aims The study aims (1) to collect a sufficiently large prospective cohort of stroke patients, with well-defined phenotypes; (2) to collect blood samples and DNA in a standardized infrastructure, allowing for storing and analyzing the samples in a uniform way; (3) to investigate associations between genetic risk loci and stroke; (4) to create possibilities to perform epidemiological studies in a well-defined hospital-based cohort of stroke patients; and (5) to allow for pooling data with other large ongoing genetic stroke studies. Key words: biobank, epidemiology, genetics, stroke
†
The Dutch String-of-Pearls Stroke Study Group consists of the following researchers: from University Medical Center Utrecht, Merel Luitse (PhD student), Berber Zweedijk (research nurse), L. Jaap Kappelle (PI, neurologist), Gabriel J. Rinkel (PI, neurologist), Ynte M. Ruigrok (neurologist), Ale Algra (clinical epidemiologist); from Academic Medical Center Amsterdam, Paul J. Nederkoorn (local PI, neurologist), Irem Baharoglu (PhD student), Mineke Ek-Post (research nurse); from University Medical Center Groningen: Gert-Jan Luijckx (local PI, neurologist), Gert Messchendorp and Friedus van de Minne (research nurses); from Maastricht University Medical Center, Robert J. Oostenbrugge (local PI, neurologist), Caroline M. J. Loos (PhD student), and Annemieke Lindl (research nurse); from Erasmus Medical Center Rotterdam, Peter J. Koudstaal (local PI, neurologist), Susanne Fonville (PhD student), and Maaike Scheele (physician assistant); from Leiden University Medical Center, Marieke J. H. Wermer (local PI, neurologist), Arend M. Hamming (PhD student), and Kirsten Janssen (research coordinator); and from Radboud University Medical Center Nijmegen, Ewoud J. van Dijk (local PI, neurologist), Joyce Wilbers (PhD student). Conflict of interest: None declared. Funding: The work described in this study was carried out in the context of the Parelsnoer Institute (PSI). PSI is part of and funded by the Dutch Federation of University Medical Centers and has received initial funding from the Dutch Government (from 2007–2011). DOI: 10.1111/ijs.12359
© 2014 World Stroke Organization
Background The String-of-Pearls Institute The String-of-Pearls Institute is a unique partnership between all eight University Medical Centers (UMCs) in the Netherlands. It was established in 2007 by the Netherlands Federation of University Medical Centers, and it conducts a large prospective cohort study in which comprehensive clinical data, imaging data, and biomaterials are collected from patients from all participating institutions (1). The general aims are to build a strong collaborative infrastructure, to allow all participants to prospectively collect their data, and to store biomaterials in a uniform and standardized format. Initially, the project focused on nine groups of medical conditions, its so-called pearls: stroke, diabetes mellitus, hereditary colorectal cancer, inflammatory bowel diseases, leukemia, neurodegenerative diseases such as Alzheimer’s disease, renal failure, rheumatoid arthritis and arthrosis, and congenital heart diseases. Currently, activities have been expanded and include other conditions such as malignancies. Genetics in stroke Previously, a genetic basis of a disease could only be proven in rare familial conditions and often consisted of single nucleotide type Vol ••, •• 2014, ••–••
1
Protocols polymorphisms (SNPs) in a specific gene. However, the majority of cases of ischemic stroke are multifactorial in etiology (2). In the last couple of years, genome-wide association studies (GWASs) have largely altered the scope in genetic research of so-called common complex diseases, in which both environmental and genetic risk factors contribute to the disease. In GWASss hundreds of thousands of variants (SNPs) across the genome are genotyped in thousands of patients and controls and analyzed for association with the risk of disease. Thereby, genetic risk loci contributing to risk of common diseases can now be unraveled (3). Ischemic stroke has several different etiologies, such as carotid stenosis, atrial fibrillation, or small-vessel disease in case of lacunar infarcts, which makes it a heterogeneous entity. Intracranial hemorrhage (ICH) can also have different causes. In about 15% of patients, a macrovascular cause is found, such as an arteriovenous malformation or fistula. In approximately 85%, ICH is supposed to be caused by small-vessel disease, classically associated with hypertension in patients with ICH in a deep location of the brain (thalamus, basal ganglia), or by cerebral amyloid angiopathy in elderly patients with a lobar ICH. Current knowledge suggests that the different stroke subtypes, ischemic and hemorrhagic, have different genetic architecture. This heterogeneity of the stroke phenotype hampers the success of GWASs in stroke. Large cohorts consisting of a specific stroke subtype are needed to achieve sufficient power to detect additional genetic risk loci. Aims The general aims of the Dutch String-of-Pearls Stroke Study are (1) to collect a sufficiently large prospective cohort of stroke patients, with well-defined phenotypes; (2) to collect blood samples and DNA in a standardized infrastructure, allowing for storing and analyzing the samples in a uniform way; (3) to investigate associations between genetic risk loci and stroke subtypes; (4) to create possibilities to perform epidemiological studies in a well-defined hospital-based cohort of stroke patients; and (5) to allow for pooling data with other large ongoing genetic stroke studies.
Protocol Study design The Dutch String-of-Pearls Stroke Study is a prospective, multicenter cohort study in which detailed clinical data and blood samples for genetic analyses are collected. The String-of-Pearls Institute has provided a regulatory framework in which ethical and legal rules and guidelines have been described (1). Patients Patients with recent transient ischemic attack (TIA) or stroke presenting in one of the eight Dutch UMCs within one-month after starting of the symptoms are eligible for this study. Patients with different cerebrovascular diseases, including TIA, ischemic stroke, ICH, subarachnoid hemorrhage, and cerebral venous sinus thrombosis, can be enrolled. All patients should give written informed consent. Clinical data Patient data and biomaterials (blood for genetic analyses as well as plasma and serum) are collected and stored according to uniform
2
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P. J. Nederkoorn et al. standard operating procedures (SOPs), and in accordance with a process information model (PIM). The use of a PIM guarantees harmonized data collection across UMCs. From each patient, detailed data are collected on history, family history, risk factors, and medication use. Precise time of onset of symptoms is recorded. Deficits at neurological examination at baseline are recorded according to the National Institutes of Health Stroke Scale. Both hemorrhagic and ischemic stroke patients are included. The subtype of ischemic stroke is defined according to both the TOAST classification and the ASCO system (4,5). Furthermore, detailed information is collected on treatment, such as intravenous thrombolysis; start of oral anticoagulation treatment in case of atrial fibrillation; and start of secondary prophylactic treatment. Data are also collected on laboratory investigations and imaging tests, that is, CT or MRI of the brain, CT or MR angiography, and duplex ultrasound investigations. Blood samples Samples of venous blood (10 cc clotted blood for serum, 8 cc EDTA blood for plasma, and 6 cc EDTA blood for DNA isolation) are collected and stored in the String-of-Pearls biobank. Samples are processed and stored according to the SOP for biomaterials. Serum and plasma samples are divided into 0·5 cc samples and stored at −80°C, and DNA is directly isolated and stored at 4°C in each local biobank. Functional outcome After three-months a structured interview by telephone is performed in order to assess functional outcome, expressed as a modified Rankin Scale score. Ethical considerations The local medical ethical committees have approved the study. The research is performed according to the principles of the Declaration of Helsinki and in accordance with the Medical Research Involving Human Subjects Act and codes on ‘good use’ of clinical data and biological samples as developed by the Dutch Federation of Medical Scientific Societies (6). Genetic analyses Currently up to 5000 stroke patients have been included in our study, with approximately 80% of them having had ischemic stroke. Inclusion is still ongoing. The current sample size does not allow for discovery of novel genetic risk loci with a GWAS approach because of limited power. Therefore, for GWAS analyses, we will aim to pool our data with other genetic stroke cohorts. Our current sample size, however, does allow replication studies of previous candidate genetic risk loci and putative newly identified genetic risk loci, especially for the ischemic stroke subtype. For comparison, publicly available previously genotyped controls will be used. Associations between SNPs or haplotypes and case/ control status will be investigated with an additive model in binary logistic regression, with adjustment for possible confounders such as age, sex, hypertension, diabetes mellitus, and smoking. We will assess statistical significance using a Bonferroni correction for the total number of SNPs or haplotypes investigated. In addition to the studies on the genetic causes of stroke, we will perform outcome studies by relating both genetic information © 2014 World Stroke Organization
Protocols
P. J. Nederkoorn et al. and clinical information (as defined by baseline characteristics), including treatment characteristics (i.e., response to intravenous thrombolysis), to outcome at three-months. To assess the relation of these genetic and clinical characteristics with outcome, Poisson regression will be used, with adjustment for possible confounders. Types of publications We anticipate two different types of publications in the String-ofPearls Cerebrovascular Diseases studies: (1) epidemiological studies on the clinical data only, without genetic analyses involved (because in this cohort, all stroke patient data will be collected in a complete and uniform way among all eight UMCs, which will form a unique prospective ‘stroke database’ allowing for detailed epidemiology association studies), and (2) genetic studies/ GWASs. In addition, other blood sample results (not genetic) can be included in the analyses.
Discussion The genetic architecture of cerebrovascular disease is still largely undiscovered. In a recent review, Rost stated that ‘apart from a few single-gene mutations associated with cerebral ischemia or intracerebral hemorrhage, stroke is a complex genetic phenotype that requires careful ascertainment and robust association testing for discovery and validation analyses’ (7). Reasons for failure to achieve more robust results in genetic stroke studies are the still too-small sample sizes of individual studies and failure to phenotype stroke subtypes in detail. An additional problem with singlecandidate gene studies is that associations can only be identified in already known ‘suspected’ genes, and completely novel genes cannot be identified by investigating these separately. Several large cohort studies are currently collecting patients for genetic studies, such as the Stroke Genetics Network (SiGN) study (8) and the Welcome Trust Genome-Wide Association Study of Ischaemic Stroke (9). The investigators of the SiGN study, which is based on meta-analyses of existing genomewide data, outlined the need to focus on precise phenotyping and on subtypes of ischemic stroke for future gene pattern discovery with genome-wide screening. Secondly, they mentioned that a sufficiently large and prospectively collected cohort is mandatory. And finally, the SIGN investigators emphasized the need to pool data and perform meta-analyses to increase power for the detection of genetic patterns related to the different subtypes of stroke.
© 2014 World Stroke Organization
A limitation of our study is selection bias, because it is carried out in University centers only. As such, we may include relatively many patients with more complicated strokes. To estimate magnitude of this possible bias we will compare our baseline characteristics and case-mix with nationwide numbers and other (genetic) stroke cohorts. Identifying novel genetic factors in stroke subtypes may contribute to treating disease by identifying new pathways involved in disease pathogenesis. In addition, genetic predisposition of individual patients may be related to stroke subtype and severity at onset and may predict their response to several interventions, such as medical treatment with antiplatelet agents or statins in ischemic stroke and functional outcome in general. Results of large prospective multicenter genetic cohort studies, as presented in this paper, may lead to individual genetic risk profiles and eventually even to individualized optimal medical treatment. With the Dutch String-of-Pearls Stroke Study, we specifically aim for precise phenotyping of subtypes of stroke for future gene pattern discovery. We also aim for pooling with other large genetic stroke cohorts. In addition, the current study, with prospective comprehensive clinical and imaging data collection, well-defined stroke phenotypes, and protocolled follow-up, will give us a unique possibility to perform high-quality epidemiological studies on a pooled cohort of stroke patients of all eight Dutch University Medical centers.
References 1 Parelsnoer Institute. PSI: Parelsnoer Institute. Available at http:// www.string-of-pearls.org/ (accessed 4 August 2014). 2 Hassan A, Markus HS. Genetics and ischaemic stroke. Brain 2000; 123:1784–812. 3 Dichgans M. Genetics of ischaemic stroke. Lancet Neurol 2007; 6:149– 61. 4 Adams HP Jr, Bendixen BH, Kapelle LJ et al. Classification of subtype of acute ischemic stroke: definitions for use in a multicenter clinical trial. Stroke 1993; 24:35–41. 5 Amarenco P, Bogousslavsky J, Caplan LR, Donnan GA, Wolf ME, Hennerici MG. The ASCOD phenotyping of ischemic stroke (updated ASCO phenotyping). Cerebrovasc Dis 2013; 36:1–5. 6 World Medical Association. WMA Declaration of Helsinki – ethical principles for medical research involving human subjects. 2013. Available at http://www.wma.net/en/10home/index.html (accessed 4 August 2014). 7 Rost NS. Clinical neurogenetics: stroke. Neurol Clin 2013; 31:915–28. 8 Meschia JF, Arnett DK, Ay H et al. Stroke Genetics Network (SiGN) Study: design and rationale for a genome-wide association study of ischaemic stroke subtypes. Stroke 2013; 44:2694–702. 9 Markus HS. Wellcome Trust genome-wide association study of ischemic stroke. Stroke 2013; 44:S20–2.
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Background In the last couple of years, genome-wide association studies have largely altered the scope in genetic research in diseases in which both environmental and genetic risk factors contribute to the disease. To date, the genetic risk loci identified in stroke have lagged behind those in other complex diseases, possibly because of the heterogeneity of stroke phenotypes. Sufficiently large cohorts with well-defined and detailed phenotyping of stroke patients are needed to identify additional genetic risk loci. Design The String-of-Pearls Institute is a unique partnership between all eight University Medical Centers in the Netherlands. It was established in 2007 by the Netherlands Federation of University Medical Centers, and it conducts a large prospective cohort study in which comprehensive clinical data, detailed phenotyping of stroke, imaging data, and biomaterials are collected in a large cohort of stroke patients. Aims The study aims (1) to collect a sufficiently large prospective cohort of stroke patients, with well-defined phenotypes; (2) to collect blood samples and DNA in a standardized infrastructure, allowing for storing and analyzing the samples in a uniform way; (3) to investigate associations between genetic risk loci and stroke; (4) to create possibilities to perform epidemiological studies in a well-defined hospital-based cohort of stroke patients; and (5) to allow for pooling data with other large ongoing genetic stroke studies. Key words: biobank, epidemiology, genetics, stroke
†
The Dutch String-of-Pearls Stroke Study Group consists of the following researchers: from University Medical Center Utrecht, Merel Luitse (PhD student), Berber Zweedijk (research nurse), L. Jaap Kappelle (PI, neurologist), Gabriel J. Rinkel (PI, neurologist), Ynte M. Ruigrok (neurologist), Ale Algra (clinical epidemiologist); from Academic Medical Center Amsterdam, Paul J. Nederkoorn (local PI, neurologist), Irem Baharoglu (PhD student), Mineke Ek-Post (research nurse); from University Medical Center Groningen: Gert-Jan Luijckx (local PI, neurologist), Gert Messchendorp and Friedus van de Minne (research nurses); from Maastricht University Medical Center, Robert J. Oostenbrugge (local PI, neurologist), Caroline M. J. Loos (PhD student), and Annemieke Lindl (research nurse); from Erasmus Medical Center Rotterdam, Peter J. Koudstaal (local PI, neurologist), Susanne Fonville (PhD student), and Maaike Scheele (physician assistant); from Leiden University Medical Center, Marieke J. H. Wermer (local PI, neurologist), Arend M. Hamming (PhD student), and Kirsten Janssen (research coordinator); and from Radboud University Medical Center Nijmegen, Ewoud J. van Dijk (local PI, neurologist), Joyce Wilbers (PhD student). Conflict of interest: None declared. Funding: The work described in this study was carried out in the context of the Parelsnoer Institute (PSI). PSI is part of and funded by the Dutch Federation of University Medical Centers and has received initial funding from the Dutch Government (from 2007–2011). DOI: 10.1111/ijs.12359
© 2014 World Stroke Organization
Background The String-of-Pearls Institute The String-of-Pearls Institute is a unique partnership between all eight University Medical Centers (UMCs) in the Netherlands. It was established in 2007 by the Netherlands Federation of University Medical Centers, and it conducts a large prospective cohort study in which comprehensive clinical data, imaging data, and biomaterials are collected from patients from all participating institutions (1). The general aims are to build a strong collaborative infrastructure, to allow all participants to prospectively collect their data, and to store biomaterials in a uniform and standardized format. Initially, the project focused on nine groups of medical conditions, its so-called pearls: stroke, diabetes mellitus, hereditary colorectal cancer, inflammatory bowel diseases, leukemia, neurodegenerative diseases such as Alzheimer’s disease, renal failure, rheumatoid arthritis and arthrosis, and congenital heart diseases. Currently, activities have been expanded and include other conditions such as malignancies. Genetics in stroke Previously, a genetic basis of a disease could only be proven in rare familial conditions and often consisted of single nucleotide type Vol ••, •• 2014, ••–••
1
Protocols polymorphisms (SNPs) in a specific gene. However, the majority of cases of ischemic stroke are multifactorial in etiology (2). In the last couple of years, genome-wide association studies (GWASs) have largely altered the scope in genetic research of so-called common complex diseases, in which both environmental and genetic risk factors contribute to the disease. In GWASss hundreds of thousands of variants (SNPs) across the genome are genotyped in thousands of patients and controls and analyzed for association with the risk of disease. Thereby, genetic risk loci contributing to risk of common diseases can now be unraveled (3). Ischemic stroke has several different etiologies, such as carotid stenosis, atrial fibrillation, or small-vessel disease in case of lacunar infarcts, which makes it a heterogeneous entity. Intracranial hemorrhage (ICH) can also have different causes. In about 15% of patients, a macrovascular cause is found, such as an arteriovenous malformation or fistula. In approximately 85%, ICH is supposed to be caused by small-vessel disease, classically associated with hypertension in patients with ICH in a deep location of the brain (thalamus, basal ganglia), or by cerebral amyloid angiopathy in elderly patients with a lobar ICH. Current knowledge suggests that the different stroke subtypes, ischemic and hemorrhagic, have different genetic architecture. This heterogeneity of the stroke phenotype hampers the success of GWASs in stroke. Large cohorts consisting of a specific stroke subtype are needed to achieve sufficient power to detect additional genetic risk loci. Aims The general aims of the Dutch String-of-Pearls Stroke Study are (1) to collect a sufficiently large prospective cohort of stroke patients, with well-defined phenotypes; (2) to collect blood samples and DNA in a standardized infrastructure, allowing for storing and analyzing the samples in a uniform way; (3) to investigate associations between genetic risk loci and stroke subtypes; (4) to create possibilities to perform epidemiological studies in a well-defined hospital-based cohort of stroke patients; and (5) to allow for pooling data with other large ongoing genetic stroke studies.
Protocol Study design The Dutch String-of-Pearls Stroke Study is a prospective, multicenter cohort study in which detailed clinical data and blood samples for genetic analyses are collected. The String-of-Pearls Institute has provided a regulatory framework in which ethical and legal rules and guidelines have been described (1). Patients Patients with recent transient ischemic attack (TIA) or stroke presenting in one of the eight Dutch UMCs within one-month after starting of the symptoms are eligible for this study. Patients with different cerebrovascular diseases, including TIA, ischemic stroke, ICH, subarachnoid hemorrhage, and cerebral venous sinus thrombosis, can be enrolled. All patients should give written informed consent. Clinical data Patient data and biomaterials (blood for genetic analyses as well as plasma and serum) are collected and stored according to uniform
2
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P. J. Nederkoorn et al. standard operating procedures (SOPs), and in accordance with a process information model (PIM). The use of a PIM guarantees harmonized data collection across UMCs. From each patient, detailed data are collected on history, family history, risk factors, and medication use. Precise time of onset of symptoms is recorded. Deficits at neurological examination at baseline are recorded according to the National Institutes of Health Stroke Scale. Both hemorrhagic and ischemic stroke patients are included. The subtype of ischemic stroke is defined according to both the TOAST classification and the ASCO system (4,5). Furthermore, detailed information is collected on treatment, such as intravenous thrombolysis; start of oral anticoagulation treatment in case of atrial fibrillation; and start of secondary prophylactic treatment. Data are also collected on laboratory investigations and imaging tests, that is, CT or MRI of the brain, CT or MR angiography, and duplex ultrasound investigations. Blood samples Samples of venous blood (10 cc clotted blood for serum, 8 cc EDTA blood for plasma, and 6 cc EDTA blood for DNA isolation) are collected and stored in the String-of-Pearls biobank. Samples are processed and stored according to the SOP for biomaterials. Serum and plasma samples are divided into 0·5 cc samples and stored at −80°C, and DNA is directly isolated and stored at 4°C in each local biobank. Functional outcome After three-months a structured interview by telephone is performed in order to assess functional outcome, expressed as a modified Rankin Scale score. Ethical considerations The local medical ethical committees have approved the study. The research is performed according to the principles of the Declaration of Helsinki and in accordance with the Medical Research Involving Human Subjects Act and codes on ‘good use’ of clinical data and biological samples as developed by the Dutch Federation of Medical Scientific Societies (6). Genetic analyses Currently up to 5000 stroke patients have been included in our study, with approximately 80% of them having had ischemic stroke. Inclusion is still ongoing. The current sample size does not allow for discovery of novel genetic risk loci with a GWAS approach because of limited power. Therefore, for GWAS analyses, we will aim to pool our data with other genetic stroke cohorts. Our current sample size, however, does allow replication studies of previous candidate genetic risk loci and putative newly identified genetic risk loci, especially for the ischemic stroke subtype. For comparison, publicly available previously genotyped controls will be used. Associations between SNPs or haplotypes and case/ control status will be investigated with an additive model in binary logistic regression, with adjustment for possible confounders such as age, sex, hypertension, diabetes mellitus, and smoking. We will assess statistical significance using a Bonferroni correction for the total number of SNPs or haplotypes investigated. In addition to the studies on the genetic causes of stroke, we will perform outcome studies by relating both genetic information © 2014 World Stroke Organization
Protocols
P. J. Nederkoorn et al. and clinical information (as defined by baseline characteristics), including treatment characteristics (i.e., response to intravenous thrombolysis), to outcome at three-months. To assess the relation of these genetic and clinical characteristics with outcome, Poisson regression will be used, with adjustment for possible confounders. Types of publications We anticipate two different types of publications in the String-ofPearls Cerebrovascular Diseases studies: (1) epidemiological studies on the clinical data only, without genetic analyses involved (because in this cohort, all stroke patient data will be collected in a complete and uniform way among all eight UMCs, which will form a unique prospective ‘stroke database’ allowing for detailed epidemiology association studies), and (2) genetic studies/ GWASs. In addition, other blood sample results (not genetic) can be included in the analyses.
Discussion The genetic architecture of cerebrovascular disease is still largely undiscovered. In a recent review, Rost stated that ‘apart from a few single-gene mutations associated with cerebral ischemia or intracerebral hemorrhage, stroke is a complex genetic phenotype that requires careful ascertainment and robust association testing for discovery and validation analyses’ (7). Reasons for failure to achieve more robust results in genetic stroke studies are the still too-small sample sizes of individual studies and failure to phenotype stroke subtypes in detail. An additional problem with singlecandidate gene studies is that associations can only be identified in already known ‘suspected’ genes, and completely novel genes cannot be identified by investigating these separately. Several large cohort studies are currently collecting patients for genetic studies, such as the Stroke Genetics Network (SiGN) study (8) and the Welcome Trust Genome-Wide Association Study of Ischaemic Stroke (9). The investigators of the SiGN study, which is based on meta-analyses of existing genomewide data, outlined the need to focus on precise phenotyping and on subtypes of ischemic stroke for future gene pattern discovery with genome-wide screening. Secondly, they mentioned that a sufficiently large and prospectively collected cohort is mandatory. And finally, the SIGN investigators emphasized the need to pool data and perform meta-analyses to increase power for the detection of genetic patterns related to the different subtypes of stroke.
© 2014 World Stroke Organization
A limitation of our study is selection bias, because it is carried out in University centers only. As such, we may include relatively many patients with more complicated strokes. To estimate magnitude of this possible bias we will compare our baseline characteristics and case-mix with nationwide numbers and other (genetic) stroke cohorts. Identifying novel genetic factors in stroke subtypes may contribute to treating disease by identifying new pathways involved in disease pathogenesis. In addition, genetic predisposition of individual patients may be related to stroke subtype and severity at onset and may predict their response to several interventions, such as medical treatment with antiplatelet agents or statins in ischemic stroke and functional outcome in general. Results of large prospective multicenter genetic cohort studies, as presented in this paper, may lead to individual genetic risk profiles and eventually even to individualized optimal medical treatment. With the Dutch String-of-Pearls Stroke Study, we specifically aim for precise phenotyping of subtypes of stroke for future gene pattern discovery. We also aim for pooling with other large genetic stroke cohorts. In addition, the current study, with prospective comprehensive clinical and imaging data collection, well-defined stroke phenotypes, and protocolled follow-up, will give us a unique possibility to perform high-quality epidemiological studies on a pooled cohort of stroke patients of all eight Dutch University Medical centers.
References 1 Parelsnoer Institute. PSI: Parelsnoer Institute. Available at http:// www.string-of-pearls.org/ (accessed 4 August 2014). 2 Hassan A, Markus HS. Genetics and ischaemic stroke. Brain 2000; 123:1784–812. 3 Dichgans M. Genetics of ischaemic stroke. Lancet Neurol 2007; 6:149– 61. 4 Adams HP Jr, Bendixen BH, Kapelle LJ et al. Classification of subtype of acute ischemic stroke: definitions for use in a multicenter clinical trial. Stroke 1993; 24:35–41. 5 Amarenco P, Bogousslavsky J, Caplan LR, Donnan GA, Wolf ME, Hennerici MG. The ASCOD phenotyping of ischemic stroke (updated ASCO phenotyping). Cerebrovasc Dis 2013; 36:1–5. 6 World Medical Association. WMA Declaration of Helsinki – ethical principles for medical research involving human subjects. 2013. Available at http://www.wma.net/en/10home/index.html (accessed 4 August 2014). 7 Rost NS. Clinical neurogenetics: stroke. Neurol Clin 2013; 31:915–28. 8 Meschia JF, Arnett DK, Ay H et al. Stroke Genetics Network (SiGN) Study: design and rationale for a genome-wide association study of ischaemic stroke subtypes. Stroke 2013; 44:2694–702. 9 Markus HS. Wellcome Trust genome-wide association study of ischemic stroke. Stroke 2013; 44:S20–2.
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