Alzheimer’s & Dementia 10 (2014) 609-618 Two rare AKAP9 variants are associated with Alzheimer’s disease in African Americans Mark W. Loguea,b,1, Matthew Schua,1, Badri N. Vardarajana, John Farrella, David A. Bennettc, Joseph D. Buxbaumd, Goldie S. Byrde, Nilufer Ertekin-Tanerf, Denis Evansg, Tatiana Foroudh, Alison Goatei, Neill R. Graff-Radfordf, M. Ilyas Kambohj, Walter A. Kukullk, Jennifer J. Manlyl, Alzheimer Disease Genetics Consortium2, Jonathan L. Hainesm, Richard Mayeuxl, Margaret A. Pericak-Vancen, Gerard D. Schellenbergo, Kathryn L. Lunettab, Clinton T. Baldwina, M. Daniele Fallinp, Lindsay A. Farrera,b,q,r,s,* aDepartment of Medicine (Biomedical Genetics), Boston University School of Medicine, Boston, MA, USA bDepartment of Biostatistics, Boston University School of Public Health, Boston, MA, USA cRush Alzheimer’s Disease Center, Rush University Medical Center, Chicago, IL, USA dDepartments of Neuroscience and Genetics & Genomic Sciences, Mount Sinai School of Medicine, New York, NY, USA eDepartment of Biology, North Carolina A & T State University, Greensboro, NC, USA fDepartments of Neuroscience and Neurology, Mayo Clinic, Jacksonville, FL, USA gRush Institute for Healthy Aging, Department of Internal Medicine, Rush University Medical Center, Chicago, IL, USA hDepartment of Medical and Molecular Genetics, Indiana University, Indianapolis, IN, USA iDepartment of Psychiatry and Hope Center Program on Protein Aggregation and Neurodegeneration, Washington University School of Medicine, St. Louis, MI, USA jDepartment of Human Genetics and Alzheimer’s Disease Research Center, University of Pittsburgh, Pittsburgh, PA, USA kNational Alzheimer’s Coordinating Center and Department of Epidemiology, University of Washington, Seattle, WA, USA lDepartment of Neurology and the Taub Institute, Columbia University, New York, NY, USA mDepartment of Epidemiology and Biostatistics, Case Western Reserve University, Cleveland, OH, USA nThe John P. Hussman Institute for Human Genomics, University of Miami, Miami, FL, USA oDepartment of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA pDepartment of Epidemiology, Johns Hopkins University School of Public Health, Baltimore, MD, USA qDepartment of Neurology, Boston University School of Medicine, Boston, MA, USA rDepartment of Ophthalmology, Boston University School of Medicine, Boston, MA, USA sDepartment of Epidemiology, Boston University School of Public Health, Boston, MA, USA Abstract Background: Less is known about the genetic basis of Alzheimer’s disease (AD) in African Amer- icans (AAs) than in non-Hispanic whites. Methods: Whole exome sequencing (WES) was performed on seven AA AD cases. Disease associ- ation with potentially AD-related variants from WES was assessed in an AA discovery cohort of 422 cases and 394 controls. Replication was sought in an AA sample of 1037 cases and 1869 controls from the Alzheimer Disease Genetics Consortium (ADGC). Results: Forty-four single nucleotide polymorphisms (SNPs) from WES passed filtering criteria and were successfully genotyped. Nominally significant (P , .05) association to AD was observed with two African-descent specific AKAP9 SNPs in tight linkage disequilibrium: rs144662445 (P 5 .014) and rs149979685 (P 5.037). These associations were replicated in the ADGC sample (rs144662445: P 5 .0022, odds ratio [OR] 5 2.75; rs149979685: P 5 .0022, OR 5 3.61). 1These two authors contributed equally to this work. *Corresponding author. 2See Acknowledgments for additional ADGC members who contributed to E-mail address: [email protected] this study. 1552-5260/$ - see front matter Ó 2014 The Alzheimer’s Association. All rights reserved. http://dx.doi.org/10.1016/j.jalz.2014.06.010 610 M.W. Logue et al. / Alzheimer’s& Dementia 10 (2014) 609-618 Conclusions: Because AKAP9 was not previously linked to AD risk, this study indicates a potential new disease mechanism. Ó 2014 The Alzheimer’s Association. All rights reserved. Keywords: Whole-exome sequencing; Late-onset Alzheimer’s disease; Rare variant; Genetic association; African American; AKAP9 1. Background design to detect highly penetrant disease variants entails sequencing individuals from families with Late onset Alzheimer’s disease (AD) is the most common multiple affected members who are more likely than form of dementia in the elderly. It has a substantial genetic sporadic cases to possess such variants. Potentially component, with heritability as high as 75% [1]. The apoli- disease-causing variants identified in familial cases can poprotein E (APOE) gene ε4 variant has been shown to be a then be genotyped in other family members or in a larger major component of this risk [2]; among non-Hispanics sample of unrelated individuals to evaluate association whites (NHW), ε4/ε4 homozygotes are estimated to have with disease. This study design was recently used in up to 15-fold increased odds of AD compared with those Caucasian samples to identify rare variants in the phos- with the most common ε3/ε3 genotype [3]. Genome-wide pholipase D3 (PLD3)gene[18]. Resequencing PLD3 association studies (GWAS) including very large NHW sam- exons in a larger number of cases and controls identified ples have identified genes harboring additional modest- additional AD-associated PLD3 variants including effect AD risk susceptibility loci (odds ratios between 1.1 Ala442Ala, which was subsequently shown to be nomi- and 1.3) [4–8]. nally significantly associated (P 5 .03) with AD in a Interestingly, even though the proportion of risk due to modest AA sample (130 cases, 172 controls). In this genetic influences is similar in African-Americans (AA) study, we performed whole-exome sequencing (WES) and NHWs [9], there is evidence that the genetic architecture of seven familial AA late-onset AD cases and subse- of the disease differs between these two populations. For quently tested association of AD risk with 44 rare vari- APOE ε example, the effect of 4 on AD risk is considerably ants selected on the basis of frequency, sequencing weaker in AAs than NHWs [3,10,11], although this quality, and relationship to previously implicated AD difference may be accounted for in part by the age- ε genes in 422 AA cases and 394 cognitively normal AA dependent effect of 4 [11]. Several genes associated with controls. We identified two African-descent specific ge- AD and AD-related hippocampal atrophy measured by mag- netic variants in the A kinase anchor protein 9 netic resonance imaging in NHWs also show evidence of as- (AKAP9) gene which are significantly enriched in AA sociation in AAs, however, most SNPs associated with AD cases versus controls, and subsequently replicated NHWs have not been replicated in AAs [12–14]. While it this association in a large independent AA cohort. The may be that the AA studies are underpowered compared studydesignissummarizedinFigure 1. with studies with NHWs, it also may be due to differences in the linkage disequilibrium (LD) structure between AAs and NHWs or population-specific causal variants underlying 2. Methods the associations. Interestingly, two GWAS replicated the as- 2.1. Study samples sociation with ABCA7 in AAs [12,14] and the risk associated with ABCA7 (odds ratio [OR] 5 1.79) approaches that of The discovery sample included AA participants of the APOE ε4 in AAs (OR 5 2.31) [14]. Nevertheless, a large Multi-Institutional Research on Alzheimer’s Genetic Epide- proportion of heritability in AAs remains unexplained. miology (MIRAGE) Study and the Genetic and Environ- Much of the heritability may be ascribed to uncommon mental Risk Factors for Alzheimer’s Disease among (frequency ,5%) or rare (frequency ,1%) variants which African Americans Study (GenerAAtions). Ascertainment would be undetectable in GWAS except in very large sam- methods, inclusion criteria, and genome-wide genotyping ples [15]. Causal high-penetrant rare variants in APP, in the MIRAGE and GenerAAtions cohorts are described PSEN1, and PSEN2 have been identified primarily in the in detail elsewhere [9,12]. The MIRAGE cohort consists of early onset familial form of the disease [16]. However, to nuclear families most of which are discordant sib pairs. date no causal variants in these genes have been reported There are also several affected sib pairs and larger in AAs. sibships. Seven MIRAGE-Study AD cases with multiple Next-generation sequencing methodologies have the affected family members, and thus more likely to be en- potential to identify rare or private risk variants many riched with rare AD-related variants, were selected for of which would be expected to exert moderate to high ef- WES. Descriptive statistics for these subjects are summa- fects on disease risk compared with most common rized in Supplementary Table 1. Four of these subjects disease-associated variants [15,17]. One efficient study were affected sib-pairs. The genotypes of single nucleotide M.W. Logue et al. / Alzheimer’s& Dementia 10 (2014) 609-618 611 Fig. 1. The overall study design including a whole-exome sequencing (WES) of seven African-American (AA) late-onset AD cases, followed by variant prioritization and filtering, genotyping in a larger AA discovery sample, and replication in a large AA Alzheimer’s Disease Genetics Consortium (ADGC) cohort. polymorphisms (SNPs) identified as potentially AD related 2.2. Sequencing and genotyping in WES were genotyped in 218 cases (including the seven sequenced subjects) and 237 controls from the MIRAGE A detailed description of WES, SNP genotyping cohort and 223 cases and 190 controls from the GenerAA- methods and quality control, and Sanger-sequencing used tions cohort. Analyses were performed on a discovery cohort to confirm genotypes of the most significant SNPs from made up of the genotyped subjects excluding the seven the analysis of the discovery data is provided in sequenced AD cases and subjects less than 60 years at the Supplementary Methods. time of assessment. The discovery cohort includes 199 cases and 209 controls from the MIRAGE cohort and 223 cases 2.3. Variant filtering and 185 controls from the GenerAAtions cohort. Character- istics of the discovery cohort are summarized in Variants identified by WES and passing quality control Supplementary Table 1.