NCRAD study

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Poster Presentation’s P1

Conclusions: These results provide valuable clues to understand the role of Blmh in the pathological process of AD . P1-278

RESULTS FROM DENSE SNP GENOTYPING OF THE 9Q22-Q31 CRI IN THE NIMH AND NCRAD AD COHORTS

Rodney Perry, Howard Wiener, Rodney Go, University of Alabama at Birmingham, Birmingham, Alabama, United States. Background: Linkage scans of Alzheimer’s disease (AD) families have identified the 9q22 region as a candidate region of interest (CRI). Second to APOE, the 9q22 signal was the most suggestive from the linkage scan of the NIMH Alzheimer’s Disease Genetic Initiative (ADGI). Additional microsatellite markers, narrowed the 1 LOD region to 11.5 cm (91.5-103 Mb). Four candidate genes located between 2-6 Mb proximal to this peak have been found significantly associated with AD. Methods: To determine more precisely the location of these signals, dense SNP genotyping of the 9q21-9q31 region, covering a total of w18 Mb which includes this region and the proximal 6 Mb was performed. Haplotag SNPs and unblocked SNPs were chosen from HapMap. SNPs in reported CNVs from this region, and SNPs covering four genes CRI, CLU, PICALM, and TOMM40 were also genotyped, resulting in a total of w 5800 SNPs. Genotyping used the Illumina iSelect platform. Results: Dense SNP genotyping was performed in the NIMH and NCRAD cohorts. Using Family Based Association Testing, besides confirming the APOE/TOMM40 and NTRK2 associations, the strongest and most consistent signals for additive and dominant models were at 86.6 Mb, 91.3 Mb, 93.2 Mb, 96.25 Mb, 97.8 Mb, 100.3 Mb, and 102.5 Mb. Conclusions: HIATL1, HABP4, GABBR2, and PRG-3, are genes in these key regions known to be expressed in brain and involved in synaptic transmission and neuroplasticity. In depth analyses including possible CNVs, and results from deep sequencing of these genes and others to identify causal variants will be presented. P1-279

REST REGULATES DYRK1A TRANSCRIPTION IN A NEGATIVE FEEDBACK LOOP

Xiulian Sun, Shandong University, Jinan, China. Background: Dual-specificity tyrosine-(Y)-phosphorylation regulated kinase 1A (DYRK1A) has been shown to be involved in learning and memory impairments in Alzheimer’s disease (AD) and Down Syndrome (DS). As a homolog of Drosophila mnb (minibrain) gene, DYRK1A also plays important roles in neurodevelopment; however, the function and regulatory mechanism of DYRK1A in neurodevelopment remain elusive. RE1-silencing transcription factor (REST) plays vital roles in neuronal differentiation. Methods: Gene regulation. Post-translational modifications. Results: Here, we found that REST can activate DYRK1A transcription via a neuron-restrictive silencer element (NRSE) at bp -833 to -815 of human DYRK1A promoter. The coordinated expression of DYRK1A and REST in mouse brain further supports the cross interaction of DYRK1A and REST during neurodevelopment. Moreover, we showed that DYRK1A dosage imbalance reduced REST protein stability and transcriptional activity through facilitating ubiquitination and subsequent degradation of REST protein. Conclusions: Therefore, the regulation of DYRK1A by REST in a negative feedback loop suggests that DYRK1A and REST are closely related in neurodevelopment. P1-280

ASSESS THE IMPORTANCE OF THE OBJECTIVE MEMORY DEFICIT IN THE ASSOCIATION OF APOE4 WITH MILD COGNITIVE IMPAIRMENT AND ITS RELATION WITH A FAST-PROGRESSION TO ALZHEIMER’S DISEASE

Alberto Marcos, Hospital Clınico San Carlos, Madrid, Spain. Background: Although the association of Mild Cognitive Impairment (MCI) with the presence of the APOE 4 allele has been described, its usefulness as a marker of evolution is controversial. Our goal was to determine whether in patients with MCI, divided according to their objective memory loss, the presence of the 4 allele could be a marker of rapid progression to Alz-

heimer’s disease (AD). Methods: A prospective, longitudinal study was carried out in Clinico Hospital, Madrid, Spain from May 2008 to December 2009. We studied 92 patients diagnosed of MCI (according to Petersen criteria). 53 of them fulfilled criteria of objective memory loss according to the California Verbal Learning Test (CVLT). Patients were evaluated periodically to complete a period of 20 months or until conversion to AD (NINCS-ADRDA criteria). We also included a control group of 40 healthy subjects matched for age and sex. The analysis of APOE genotypes by PCR and subsequent restriction enzyme digestion was performed. Results: In the 92 MCI patients, 37 (43.5%) were carriers of allele 4 vs. 5 (14.7%) control subjects (OR 4.5 95% 1.6 to 12.7). When we analyzed 53 patients with objective memory loss, 28 (57.1%) were carriers of APOE4 compared with 4 controls (15.4%) (OR 7.3 95% CI 2.2 to 24.5). 55 of these patients were followed during 20 months. 60% of all carriers of APOE4 and 58.8% of carriers of APOE4 with memory loss, objectified by the CVLT, progressed to AD. Conclusions: The presence of the APOE 4 allele is more related to the risk of MCI with objective memory loss than the patient’s progression to AD. P1-281

MULTILOCUS GENOTYPE PATTERN AT PICALM, CLU AND APOE GENES IS ASSOCIATED WITH EPISODIC MEMORY IN THE NIA-LOAD/NCRAD STUDY

David A. Bennett1, Thomas Bird2, Bradley Boeve3, Ramon Diaz-Arrastia4, M. Farlow5, Tatiana Foroud5, Alison Goate6, Neill Graff-Radford3, Robert Sweet7, Robert Wilson1, Richard Mayeux8, 1Rush University Medical Center, Chicago, Illinois, United States; 2University of Washington, Seattle, Washington, United States; 3Mayo Clinic, Jacksonville, Florida, United States; 4University of Texas Southwestern Medical Center, Dallas, Texas, United States; 5Indiana University School of Medicine, Indianapolis, Indiana, United States; 6Washington University School of Medicine, St. Louis, Missouri, United States; 7University of Pittsburgh, Pittsburgh, Pennsylvania, United States; 8Columbia University, New York City, New York, United States. Background: Genomewide association studies (GWAS) have identified new susceptibility genes for Late Onset Alzheimer’s Disease (AD). Additional efforts also confirmed endophenotypes as powerful disease’s surrogates that overcome phenotypic heterogeneity. However, GWAS also revealed very modest effect sizes, reinforcing the multi factorial nature of AD, suggesting that single SNP approaches might be underpowered. We combined several genetic variants at different AD susceptibility genes into multi-locus genotype patterns (MLGP) and evaluated their association with memory performance. Methods: We used 2091 AD cases and 2200 non demented individuals of European ancestry from the National Institute on Aging-Late Onset Alzheimer’s Disease Family Study (NIA-LOAD)that were genotyped using Illumina 610quad platform. All participants completed a battery of cognitive tests and a composite measure of episodic memory was used as cognitive endophenotype. We tabulated multi-locus genotype patterns using genotypes at CLU, PICALM and APOE genes. Using Generalized Estimating Equations (to adjust for family structure), we modeled episodic memory as the dependent endophenotype, MLGPas a predictor variable and sex, age and education as covariates. Analysis was carried in all subjects and in unaffected subjects only. Results: Two of the CLU and PICALM patterns significantly influenced episodic memory performance. The genotypic pattern TT-GG is a nominally significant genetic risk factor of episodic performance in all subjects (ß¼-0.24, SE ¼ 0.12, p ¼ 0.039) and after excluding AD cases (ß¼-0.32, SE ¼ 0.14, p ¼ 0.021). After adding APOE genotype, the effect of the genotype pattern TT-GG-e4 increased (ß¼-0.47, SE ¼ 0.19, p ¼ 0.016). The genotype pattern CC-AG is a nominally significant predictor of a better episodic performance in all subjects (ß¼ 0.15, SE ¼ 0.06, p ¼ 0.020) and showed a trend to significance in unaffected subjects (ß ¼0.15, SE ¼ 0.08, p ¼ 0.057). After adding APOE, the genotype pattern was no longer significant associated with episodic performance (ß¼-0.09, 0.10, p ¼ 0.354 in all subjects; ß¼-0.15, SE ¼ 0.14, p ¼ 0.305 after excluding AD cases). However, when we considered e3e3 homozygotes, the genotype pattern CC-AG-e3e3 is significant associated with episodic memory performance in all subjects (ß¼0.27, SE ¼ 0.10, p ¼ 0.005) and unaffected subjects (ß¼0.26, SE ¼

Poster Presentation’s P1 0.10, p ¼ 0.010). Conclusions: MLGP combining the effects of different associated SNPs might be a powerful analytical approach to predict individual’s genetic risk associated with memory performance. P1-282

ANNAMARIA CONFALONI PHD

Paola Piscopo1, Giuseppina Talarico2, Daniela Galimberti3, Saviana Barbati1, Marina Gasparini2, Chiara Belli1, Angelo Toto1, Anna Poleggi1, Alessio Crestini1, Elio Scarpini3, Giuseppe Bruno2, Annamaria Confaloni1, 1Istituto Superiore di Sanita, Rome, Italy; 2 University of Rome “Sapienza”, Rome, Italy; 3University of Milan, Ospedale Policlinico, Milan, Italy. Background: Sortilin-related receptor 1 (SORL1) plays a key role in the recycling of the amyloid precursor protein (APP) from the cell surface via the endocytic pathways. In fact, the APP holoprotein is synthesized in the endoplasmic reticulum and Golgi; then, it is processed by alpha- or beta- secretase and, successively, by the gamma-secretase complex generating A-beta fragments. Alternatively, SORL1 binds APP holoprotein acting as a sorting receptor. Absence of SORL1 switches APP away from the retromer recycling pathway directing it into the beta-secretase pathway and so increasing A-beta peptide production. Recently, Rogaeva described an association between some SORL1 genetic variants and late-onset Alzheimer’s disease (AD). Methods: After obtaining informed consent, blood samples were taken from patients and control subjects. Genomic DNAwas extracted from peripheral blood leukocytes. Genetic analysis was performed using Direct sequencing by DNA sequencer Beckman CEQ 8000 and HRM analysis by RotorGene 6000 real-time analyzer. Results: The aim of this study was to study allelic variants of SORL1 to verify if they could be genetics markers of AD and/or predictors of MCI to AD conversion. We carried out a genetic analysis on SORL1 gene using tagging SNPs representative of three different LD blocks: rs641120, rs1010159 and rs641120. We found that rs641120 and rs1010159 frequencies of SORL1 were increased in the cases compared to controls. Moreover, rs641120 SNP seems to be more associated to the MCI subjects converted in AD respect to no-converted MCI group. Conclusions: Our results support the evidence that genetic variants of SORL1 affect the susceptibility to develop AD; moreover, this gene could be considered a genetic determinant to predict the progression of clinical phenotype from MCI to Alzheimer’s disease. P1-283

MITOCHONDRIAL DNA HAPLOGROUP, AD INCIDENCE, AND COGNITIVE FUNCTION: THE CACHE COUNTY STUDY ON MEMORY, HEALTH, AND AGING

John Kauwe1, Richard Cawthon2, Richard Kerber3, JoAnn Tschanz4, David Ward5, Maria Norton4, Peter Zandi6, Kathleen Welsh-Bohmer7, John Breitner8, Ronald Munger4, 1Brigham Young University, Provo, Utah, United States; 2University of Utah, Salt Lake City, Utah, United States; 3 University of Louisville, Louisville, Kentucky, United States; 4Utah State University, Logan, Utah, United States; 5University of Hawaii, Honolulu, Hawaii, United States; 6Johns Hopkins University, Baltimore, Maryland, United States; 7Duke University, Durham, North Carolina, United States; 8 McGill University, Montreal, Quebec, Canada. Background: Previous findings from case-control studies of Alzheimer’s disease (AD) and mitochondrial DNA (mtDNA) haplogroup have identified some significant associations, including higher AD risk among some populations for individuals in haplogroup H. Further examination of AD incidence in tandem with quantitative cognitive phenotypes leading to AD onset may help to refine our understanding of the relationship between haplogroup and AD risk. Methods: mtDNA haplogroups were determined for 1009 persons without dementia at baseline from the Cache County Memory Study cohort, comprised almost entirely by individuals of European ancestry. For 284 of these subjects, mtDNA sequences were typed from stored blood samples. Haplogroups for the remaining 768 individuals were imputed based on matrilineages determined from the Utah Population Database (UPDB). Five major mtDNA haplogroups were identified, including H, UK, WIX, U, and JT. Cognition was assessed with the Modified Mini-Mental State Exam (3MS). Linear mixed models examined the effects of haplogroup on average 3MS and rate of 3MS decline, controlling also for APOE genotype, age, gender, and education. Risk for AD was assessed

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using logistic regression. Results: Of the 968 persons with available mtDNA data, 45.0% were classified in the H haplogroup, 22.1% in JT, 23.9% in UK, 4.0% in V, and 5.0% in WIX. There were no significance differences between haplogroups with respect to gender, age, APOE genotype, or education level. Individuals in haplogroups H, JT, and WIX had significantly greater AD risk relative to UK and V (OR ¼ 1.97, 95% CI ¼ (1.07,3.66), p ¼ 0.04). There was additionally a significant association between mtDNA haplogroup and baseline 3MS (p ¼ 0.02). In adjusted linear mixed models, we found significant overall differences in average 3MS in that individuals classified as H, JT, and WIX had significantly lower 3MS scores over time in comparison to the UK and V haplogroups (p ¼ 0.04), although over the course of study their rates of cognitive decline were not significantly different. Conclusions: In this sample, there was evidence of an association between mtDNA haplogroup and both AD incidence and average cognitive function, although there was not a significant association with rate of cognitive change. P1-284

FUNCTIONAL ANALYSIS OF THE MAPT PROMOTER HAPLOTYPES

Tara Caffrey, Richard Wade-Martins, University of Oxford, Oxford, United Kingdom. Background: Two major haplotypes have been defined spanning the microtubule associated protein tau (MAPT) genomic locus, H1 and H2, of which H1 is associated with Parkinson’s disease and progressive supranuclear palsy. Despite consistent and strong association of the H1 haplotype with tauopathies, the mechanisms by which H1 polymorphisms confer susceptibility to neurodegeneration have not been defined. One theory to explain the association of neurodegenerative disease with MAPT H1 polymorphisms proposes that there are overall expression differences between the two haplotypes influenced by variants within the promoter region. The aim of this study is to examine haplotype-specific expression of the MAPT promoter haplotypes in cell culture models using genomic DNA reporter gene vectors. Methods: Our lab has developed an efficient viral delivery and expression system for genomic DNA loci up to 150 kb called the infectious bacterial artificial chromosome (iBAC). Delivery of whole genomic loci using this system allows for the study of complex expression patterns influenced by non-coding regulatory sequences. Using RecE/RecT homologous recombination in bacteria, we can modify large genomic DNA expression vectors with base pair accuracy. By harnessing this technology, we are able to insert reporter genes in frame with our gene of interest. Results: We have generated iBAC-MAPT H1 and H2 genomic DNA expression vectors containing the luciferase reporter gene. The luciferase reporter gene is expressed by the native MAPT promoter at the 3’ end of the MAPT coding sequence; This design permits luciferase to report expression of all tau isoforms. Luciferase is separated from the MAPT coding sequence by a viral 2A linker sequence which allows equimolar expression of the proteins while eliminating confounding effects of altered sub-cellular localization or sequestration of the reporter. Conclusions: The significant advantage of the proposed expression vector design over past MAPT promoter investigations is luciferase expression is under the control of the native MAPT regulatory sequences including 8 kb of promoter, and therefore will be expressed at physiologically relevant levels and subject to the same cis-acting regulatory sequences as the MAPT gene. This system will allow us to investigate the effect of haplotype-specific polymorphisms on the expression of the MAPT locus. P1-285

SIGMA NON-OPIOID INTRACELLULAR RECEPTOR 1 AND NEURODEGENERATION: PREVALENCE OF MUTATIONS AND THERAPEUTIC IMPLICATIONS

Carol Dobson-Stone1, Agnes Luty1, Marianne Hallupp1, Kirsten Coupland1, William Brooks1, Peter Panegyres2, Olivier Piguet1, G Anthony Broe1, Tomasz Sobow3, Cezary Zekanowski4, Glenda Halliday1, Peter Schofield1, John Kwok1, 1Neuroscience Research Australia, Sydney, Australia; 2Department of Health, Perth, Australia; 3Medical University of Lodz, Lodz, Poland; 4Polish Academy of Sciences, Warsaw, Poland. Background: Frontotemporal lobar degeneration (FTLD) is a common cause of presenile dementia and shows significant symptomatic and