ABSTRACT
The genetics of complex disease produce alterations in the molecular interactions of cellular pathways whose collective effect may become clear through the organized structure of molecular networks. To characterize molecular systems associated with late-onset Alzheimer's disease (LOAD), we constructed gene-regulatory networks in 1,647 postmortem brain tissues from LOAD patients and nondemented subjects, and we demonstrate that LOAD reconfigures specific portions of the molecular interaction structure. Through an integrative network-based approach, we rank-ordered these network structures for relevance to LOAD pathology, highlighting an immune- and microglia-specific module that is dominated by genes involved in pathogen phagocytosis, contains TYROBP as a key regulator, and is upregulated in LOAD. Mouse microglia cells overexpressing intact or truncated TYROBP revealed expression changes that significantly overlapped the human brain TYROBP network. Thus the causal network structure is a useful predictor of response to gene perturbations and presents a framework to test models of disease mechanisms underlying LOAD.
Subject(s)
Alzheimer Disease/genetics , Brain/metabolism , Gene Regulatory Networks , Adaptor Proteins, Signal Transducing/metabolism , Alzheimer Disease/metabolism , Animals , Bayes Theorem , Brain/pathology , Humans , Membrane Proteins/metabolism , Mice , Microglia/metabolismABSTRACT
BACKGROUND: Persons with mixed hyperlipidemia are at risk for atherosclerotic cardiovascular disease due to an elevated non-high-density lipoprotein (HDL) cholesterol level, which is driven by remnant cholesterol in triglyceride-rich lipoproteins. The metabolism and clearance of triglyceride-rich lipoproteins are down-regulated through apolipoprotein C3 (APOC3)-mediated inhibition of lipoprotein lipase. METHODS: We carried out a 48-week, phase 2b, double-blind, randomized, placebo-controlled trial evaluating the safety and efficacy of plozasiran, a hepatocyte-targeted APOC3 small interfering RNA, in patients with mixed hyperlipidemia (i.e., a triglyceride level of 150 to 499 mg per deciliter and either a low-density lipoprotein [LDL] cholesterol level of ≥70 mg per deciliter or a non-HDL cholesterol level of ≥100 mg per deciliter). The participants were assigned in a 3:1 ratio to receive plozasiran or placebo within each of four cohorts. In the first three cohorts, the participants received a subcutaneous injection of plozasiran (10 mg, 25 mg, or 50 mg) or placebo on day 1 and at week 12 (quarterly doses). In the fourth cohort, participants received 50 mg of plozasiran or placebo on day 1 and at week 24 (half-yearly dose). The data from the participants who received placebo were pooled. The primary end point was the percent change in fasting triglyceride level at week 24. RESULTS: A total of 353 participants underwent randomization. At week 24, significant reductions in the fasting triglyceride level were observed with plozasiran, with differences, as compared with placebo, in the least-squares mean percent change from baseline of -49.8 percentage points (95% confidence interval [CI], -59.0 to -40.6) with the 10-mg-quarterly dose, -56.0 percentage points (95% CI, -65.1 to -46.8) with the 25-mg-quarterly dose, -62.4 percentage points (95% CI, -71.5 to -53.2) with the 50-mg-quarterly dose, and -44.2 percentage points (95% CI, -53.4 to -35.0) with the 50-mg-half-yearly dose (P<0.001 for all comparisons). Worsening glycemic control was observed in 10% of the participants receiving placebo, 12% of those receiving the 10-mg-quarterly dose, 7% of those receiving the 25-mg-quarterly dose, 20% of those receiving the 50-mg-quarterly dose, and 21% of those receiving the 50-mg-half-yearly dose. CONCLUSIONS: In this randomized, controlled trial involving participants with mixed hyperlipidemia, plozasiran, as compared with placebo, significantly reduced triglyceride levels at 24 weeks. A clinical outcomes trial is warranted. (Funded by Arrowhead Pharmaceuticals; MUIR ClinicalTrials.gov number NCT04998201.).
ABSTRACT
Using expression profiles from postmortem prefrontal cortex samples of 624 dementia patients and non-demented controls, we investigated global disruptions in the co-regulation of genes in two neurodegenerative diseases, late-onset Alzheimer's disease (AD) and Huntington's disease (HD). We identified networks of differentially co-expressed (DC) gene pairs that either gained or lost correlation in disease cases relative to the control group, with the former dominant for both AD and HD and both patterns replicating in independent human cohorts of AD and aging. When aligning networks of DC patterns and physical interactions, we identified a 242-gene subnetwork enriched for independent AD/HD signatures. This subnetwork revealed a surprising dichotomy of gained/lost correlations among two inter-connected processes, chromatin organization and neural differentiation, and included DNA methyltransferases, DNMT1 and DNMT3A, of which we predicted the former but not latter as a key regulator. To validate the inter-connection of these two processes and our key regulator prediction, we generated two brain-specific knockout (KO) mice and show that Dnmt1 KO signature significantly overlaps with the subnetwork (P = 3.1 × 10(-12)), while Dnmt3a KO signature does not (P = 0.017).
Subject(s)
Alzheimer Disease/genetics , Gene Regulatory Networks , Huntington Disease/genetics , Prefrontal Cortex/metabolism , Alzheimer Disease/pathology , Animals , Autopsy , Case-Control Studies , Chromatin/metabolism , DNA (Cytosine-5-)-Methyltransferase 1 , DNA (Cytosine-5-)-Methyltransferases/genetics , DNA Methyltransferase 3A , Dementia/pathology , Gene Expression Profiling , Gene Expression Regulation , Humans , Huntington Disease/pathology , Mice , Mice, Knockout , Prefrontal Cortex/pathology , Reproducibility of ResultsABSTRACT
Elevated triglycerides and non-high-density lipoprotein cholesterol (HDL-C) are risk factors for atherosclerotic cardiovascular disease (ASCVD). ARO-ANG3 is an RNA interference therapy that targets angiopoietin-like protein 3 (ANGPTL3), a regulator of lipoprotein metabolism. This first-in-human, phase 1, randomized, placebo-controlled, open-label trial investigated single and repeat ARO-ANG3 doses in four cohorts of fifty-two healthy participants and one cohort of nine participants with hepatic steatosis, part of a basket trial. Safety (primary objective) and pharmacokinetics (in healthy participants) and pharmacodynamics (secondary objectives) of ARO-ANG3 were evaluated. ARO-ANG3 was generally well tolerated, with similar frequencies of treatment-emergent adverse events in active and placebo groups. Systemic absorption of ARO-ANG3 in healthy participants was rapid and sustained, with a mean Tmax of 6.0-10.5 h and clearance from plasma within 24-48 h after dosing with a mean t½ of 3.9-6.6 h. In healthy participants, ARO-ANG3 treatment reduced ANGPTL3 (mean -45% to -78%) 85 days after dose. Reductions in triglyceride (median -34% to -54%) and non-HDL-C (mean -18% to -29%) (exploratory endpoints) concentrations occurred with the three highest doses. These early-phase data support ANGPTL3 as a potential therapeutic target for ASCVD treatment. ClinicalTrials.gov identifier: NCT03747224.
Subject(s)
Angiopoietin-Like Protein 3 , Atherosclerosis , Humans , Triglycerides , RNA Interference , Cholesterol , Atherosclerosis/drug therapy , Atherosclerosis/geneticsABSTRACT
BACKGROUND: Apolipoprotein C-III (APOC3) inhibits triglyceride clearance by reducing lipoprotein lipasemediated hydrolysis and hepatocyte uptake of triglyceride-rich lipoproteins. ARO-APOC3, a hepatocyte-targeting RNA interference therapeutic, inhibits APOC3 messenger ribonucleic acid expression, lowering triglyceride levels. The objective of this trial was to assess the safety, pharmacodynamic variables, and pharmacokinetic variables of ARO-APOC3 treatment. METHODS: Healthy participants and adults with hypertriglyceridemia were randomly assigned to receive escalating single (day 1) or repeat (days 1 and 29) doses, respectively, of subcutaneous injections of ARO-APOC3 10, 25, 50, or 100 mg or placebo; they were followed up until day 113. Additional cohorts of healthy participants and adults with chylomicronemia received repeat doses of open-label ARO-APOC3. The primary objective was to evaluate the safety and side effect profile of ARO-APOC3. Key secondary and exploratory objectives included pharmacokinetic variables and changes in serum APOC3, triglyceride, and cholesterol levels. RESULTS: Eighty-eight participants received ARO-APOC3 and 24 participants received placebo across double-blind and open-label cohorts. Treatment-emergent adverse events (AEs) of transient, mild to moderate liver transaminase changes occurred in 10 participants: 1 patient receiving ARO-APOC3 25 mg, 5 patients receiving ARO-APOC3 50 mg, and 4 participants receiving ARO-APOC3 100 mg (1 healthy participant and 3 patients with hypertriglyceridemia). These events were asymptomatic, and transaminase levels returned to near baseline by the end of the trial. No AEs related to thrombocytopenia or platelet declines were reported. In the hypertriglyceridemia cohorts, the day 113 mean changes from baseline in APOC3 at the 10-, 25-, 50-, and 100-mg doses were −62.0%, −81.7%, −90.1%, and −94.4%, respectively, compared with −1.6% with placebo. This corresponded to median changes in triglyceride levels of −65.6%, −69.9%, −81.2%, and −81.0% compared with −2.8% with placebo. CONCLUSIONS: In this small trial of short duration, ARO-APOC3 was associated with few AEs and reduced serum levels of APOC3 and triglycerides in healthy participants and patients with hypertriglyceridemia. (Funded by Arrowhead Pharmaceuticals, Inc.; ClinicalTrials.gov number, NCT03783377.)
Subject(s)
Hypertriglyceridemia , Humans , Apolipoprotein C-III/genetics , RNA Interference , Hypertriglyceridemia/complicationsABSTRACT
Frontotemporal dementia (FTD) is the second most common cause of dementia in people under the age of 65 years. A large proportion of FTD patients (35-50%) have a family history of dementia, consistent with a strong genetic component to the disease. In 1998, mutations in the gene encoding the microtubule-associated protein tau (MAPT) were shown to cause familial FTD with parkinsonism linked to chromosome 17q21 (FTDP-17). The neuropathology of patients with defined MAPT mutations is characterized by cytoplasmic neurofibrillary inclusions composed of hyperphosphorylated tau. However, in multiple FTD families with significant evidence for linkage to the same region on chromosome 17q21 (D17S1787-D17S806), mutations in MAPT have not been found and the patients consistently lack tau-immunoreactive inclusion pathology. In contrast, these patients have ubiquitin (ub)-immunoreactive neuronal cytoplasmic inclusions and characteristic lentiform ub-immunoreactive neuronal intranuclear inclusions. Here we demonstrate that in these families, FTD is caused by mutations in progranulin (PGRN) that are likely to create null alleles. PGRN is located 1.7 Mb centromeric of MAPT on chromosome 17q21.31 and encodes a 68.5-kDa secreted growth factor involved in the regulation of multiple processes including development, wound repair and inflammation. PGRN has also been strongly linked to tumorigenesis. Moreover, PGRN expression is increased in activated microglia in many neurodegenerative diseases including Creutzfeldt-Jakob disease, motor neuron disease and Alzheimer's disease. Our results identify mutations in PGRN as a cause of neurodegenerative disease and indicate the importance of PGRN function for neuronal survival.
Subject(s)
Chromosomes, Human, Pair 17/genetics , Dementia/genetics , Frontal Lobe/physiopathology , Intercellular Signaling Peptides and Proteins/genetics , Mutation/genetics , Protein Precursors/genetics , Temporal Lobe/physiopathology , Cell Survival , Codon, Terminator/genetics , Dementia/physiopathology , Frontal Lobe/metabolism , Genetic Linkage/genetics , Humans , Intercellular Signaling Peptides and Proteins/metabolism , Neurons/metabolism , Neurons/pathology , Physical Chromosome Mapping , Progranulins , Protein Precursors/metabolism , RNA Stability , RNA, Messenger/genetics , RNA, Messenger/metabolism , Temporal Lobe/metabolism , tau Proteins/deficiency , tau Proteins/geneticsABSTRACT
Compelling evidence supports a causal role for lipoprotein(a) (Lp(a)) in cardiovascular disease. No pharmacotherapies directly targeting Lp(a) are currently available for clinical use. Here we report the discovery and development of olpasiran, a first-in-class, synthetic, double-stranded, N-acetylgalactosamine-conjugated small interfering RNA (siRNA) designed to directly inhibit LPA messenger RNA translation in hepatocytes and potently reduce plasma Lp(a) concentration. Olpasiran reduced Lp(a) concentrations in transgenic mice and cynomolgus monkeys in a dose-responsive manner, achieving up to over 80% reduction from baseline for 5-8 weeks after administration of a single dose. In a phase 1 dose-escalation trial of olpasiran (ClinicalTrials.gov: NCT03626662 ), the primary outcome was safety and tolerability, and the secondary outcomes were the change in Lp(a) concentrations and olpasiran pharmacokinetic parameters. Participants tolerated single doses of olpasiran well and experienced a 71-97% reduction in Lp(a) concentration with effects persisting for several months after administration of doses of 9 mg or higher. Serum concentrations of olpasiran increased approximately dose proportionally. Collectively, these results validate the approach of using hepatocyte-targeted siRNA to potently lower Lp(a) in individuals with elevated plasma Lp(a) concentration.
Subject(s)
Lipoprotein(a) , RNA, Small Interfering , Adult , Animals , Female , Humans , Male , Mice , Middle Aged , Hyperlipidemias/drug therapy , Lipoprotein(a)/blood , Lipoprotein(a)/genetics , Macaca fascicularis , Mice, Transgenic , RNA, Small Interfering/geneticsABSTRACT
The apolipoprotein E (APOE) epsilon4 allele is the best established genetic risk factor for late-onset Alzheimer's disease (LOAD). We conducted genome-wide surveys of 502,627 single-nucleotide polymorphisms (SNPs) to characterize and confirm other LOAD susceptibility genes. In epsilon4 carriers from neuropathologically verified discovery, neuropathologically verified replication, and clinically characterized replication cohorts of 1411 cases and controls, LOAD was associated with six SNPs from the GRB-associated binding protein 2 (GAB2) gene and a common haplotype encompassing the entire GAB2 gene. SNP rs2373115 (p = 9 x 10(-11)) was associated with an odds ratio of 4.06 (confidence interval 2.81-14.69), which interacts with APOE epsilon4 to further modify risk. GAB2 was overexpressed in pathologically vulnerable neurons; the Gab2 protein was detected in neurons, tangle-bearing neurons, and dystrophic neuritis; and interference with GAB2 gene expression increased tau phosphorylation. Our findings suggest that GAB2 modifies LOAD risk in APOE epsilon4 carriers and influences Alzheimer's neuropathology.
Subject(s)
Adaptor Proteins, Signal Transducing/genetics , Alzheimer Disease/genetics , Alzheimer Disease/metabolism , Apolipoprotein E4/genetics , Genetic Predisposition to Disease/genetics , Polymorphism, Single Nucleotide/genetics , Brain/metabolism , Brain/pathology , Brain/physiopathology , Brain Chemistry/genetics , Cell Line, Tumor , DNA Mutational Analysis , Gene Expression Regulation/genetics , Gene Frequency , Genetic Markers/genetics , Genetic Testing , Haplotypes/genetics , Humans , Mutation , Neurofibrillary Tangles/genetics , Neurofibrillary Tangles/metabolism , Neurofibrillary Tangles/pathology , Phosphorylation , Risk Factors , tau Proteins/metabolismABSTRACT
Late-onset Alzheimer's disease (LOAD) is the most common neurodegenerative disorder, and has a complex etiology. Recently an intronic polymorphism in the ubiquilin 1 gene (UBQLN1) and a particular haplotype was reported to be associated with LOAD. We investigated whether variants in UBQLN1 confer a risk for the disease in 407 Polish LOAD patients and 407 controls. We observed a weak association with the rs2781002 polymorphism, however, contrary to the initial reports, in our group the association was with the A allele. Risk estimation for AA versus GG genotypes showed that the AA genotype is a weak risk factor for AD (OR = 1.8, 95% CI = 1.1-3.1, p = 0.025). This effect was stronger in a group of LOAD patients without APOE4 allele. Haplotype analyses indicate that there is an increase of haplotypes with an A allele in the case group. Also, the specific haplotypes with the A allele that increase AD risk differ between the APOE4-positive and APOE4-negative pools. However, the association observed seems to be driven mostly by rare (<5%) haplotypes. Results suggest a need for additional association studies and in silico analysis of the UBQLN1 locus.
Subject(s)
Alzheimer Disease/epidemiology , Alzheimer Disease/genetics , Carrier Proteins/genetics , Cell Cycle Proteins/genetics , Polymorphism, Single Nucleotide , Adaptor Proteins, Signal Transducing , Age of Onset , Aged , Apolipoprotein E4/genetics , Autophagy-Related Proteins , Female , Genetic Predisposition to Disease/epidemiology , Haplotypes , Humans , Male , Poland/epidemiology , Risk FactorsABSTRACT
BACKGROUND: The objective of this study was to analyze factors influencing the risk and timing of Alzheimer's disease (AD) in central Norway. The APOE epsilon4 allele is the only consistently identified risk factor for late onset Alzheimer's disease (LOAD). We have described the allele frequencies of the apolipoprotein E gene (APOE) in a large population of patients with AD compared to the frequencies in a cognitively-normal control group, and estimated the effect of the APOE epsilon4 allele on the risk and the age at onset of AD in this population. METHODS: 376 patients diagnosed with AD and 561 cognitively-normal control individuals with no known first degree relatives with dementia were genotyped for the APOE alleles. Allele frequencies and genotypes in patients and control individuals were compared. Odds Ratio for developing AD in different genotypes was calculated. RESULTS: Odds Ratio (OR) for developing AD was significantly increased in carriers of the APOE epsilon4 allele compared to individuals with the APOE epsilon3/epsilon3 genotype. Individuals carrying APOE epsilon4/epsilon4 had OR of 12.9 for developing AD, while carriers of APOE epsilon2/epsilon4 and APOE epsilon3/epsilon4 had OR of 3.2 and 4.2 respectively. The effect of the APOE epsilon4 allele was weaker with increasing age. Carrying the APOE epsilon2 allele showed no significant protective effect against AD and did not influence age at onset of the disease. Onset in LOAD patients was significantly reduced in a dose dependent manner from 78.4 years in patients without the APOE epsilon4 allele, to 75.3 in carriers of one APOE epsilon4 allele and 72.9 in carriers of two APOE epsilon4 alleles. Age at onset in early onset AD (EOAD) was not influenced by APOE epsilon4 alleles. CONCLUSION: APOE epsilon4 is a very strong risk factor for AD in the population of central Norway, and lowers age at onset of LOAD significantly.
Subject(s)
Alzheimer Disease/epidemiology , Alzheimer Disease/genetics , Apolipoprotein E4/genetics , Gene Frequency/genetics , Age of Onset , Aged , Aged, 80 and over , Alleles , Case-Control Studies , Female , Humans , Male , Norway/epidemiology , Odds Ratio , Risk FactorsABSTRACT
OBJECTIVE: To estimate the effect of education on the risk of Alzheimer's disease (AD). METHODS: 373 patients diagnosed with AD and 559 healthy control individuals without first degree relatives with known dementia, were included in a case-control study (2003-2006). All individuals were genotyped for APOE alleles. Odds ratio (OR) for developing AD was calculated by binary logistic regression, with the number of APOE epsilon 4 alleles and educational level as covariates. Analyses were carried out separately for men and women and for different age groups. RESULTS: Carriers of one APOE epsilon 4 allele had OR of 4.2, and carriers of two APOE epsilon 4 alleles OR of 12.4 for developing AD. When adjusted for the number of APOE epsilon 4 alleles, OR for developing AD was significantly reduced in participants with 8-9 years of education compared to those with only 6-7 years, and was reduced further for those with 10-18 years of education. These findings were obtained for all the age groups studied and for both men and women. CONCLUSIONS: Education had a consistently protective effect on the risk of developing clinical AD in a dose-dependent manner in both men and women, and in all age groups, also when adjusting for the number of APOE epsilon 4 alleles. Male gender was protective, probably at least in part because of a higher educational level.
Subject(s)
Alzheimer Disease/genetics , Alzheimer Disease/prevention & control , Apolipoprotein E4/genetics , Educational Status , Age of Onset , Aged , Aged, 80 and over , Alleles , Case-Control Studies , Female , Gene Frequency , Genetic Predisposition to Disease/genetics , Humans , Logistic Models , Male , Odds Ratio , Risk Assessment , Sex DistributionABSTRACT
Over 30 different mutations have now been identified in MAPt that cause frontotemporal dementia (FTD). However, there are several families with FTD that show definite linkage to the region on chromosome 17 that contains MAPt, in which no mutation(s) has been identified. Although these families could have a complex mutation of the MAPt locus that has evaded detection it is also possible that another gene in this region is associated with FTD. This possibility is supported by neuropathological findings in these families, which consist of neuronal inclusions that are immunoreactive for ubiquitin (ub-ir) but not for tau. In addition to neuronal cytoplasmic inclusions, several chromosome 17-linked families are reported to have ub-ir neuronal intranuclear inclusions (NII); a finding which is uncommon in sporadic FTD. Here, we describe detailed clinical and neuropathological findings in a new large, multigenerational family with autosomal dominant FTD and autopsy proven tau-negative, ub-ir neuronal cytoplasmic and intranuclear inclusions. We have demonstrated that this family is linked to a 19.06 cM region of chromosome 17q21 with a maximum multipoint LOD score of 3.911 containing MAPt. By combining the results of our genetic analysis with those previously published for other families with similar pathology, we have further refined the minimal region to a 3.53 cM region of chromosome 17q21. We did not identify point mutations in MAPt by direct sequencing or any gross MAPt gene alterations using fluorescent in situ hybridization. In addition, tau protein extracted from members of this family was unremarkable in size and quantity as assessed by western blotting. Neuropathological characterization of the ub-ir NII in this family shows that they are positive for promyelocytic leukaemia protein (PML) and SUMO-1 that suggests that these inclusions form in the nuclear body and suggests a possible mechanism of neurodegeneration in tau-negative FTD linked to chromosome 17q21.
Subject(s)
Chromosomes, Human, Pair 17/genetics , Dementia/genetics , Intranuclear Inclusion Bodies/pathology , Aged , Blotting, Western , Dementia/metabolism , Dementia/pathology , Female , Genetic Linkage , Genetic Predisposition to Disease , Humans , In Situ Hybridization, Fluorescence , Male , Middle Aged , Neurons/ultrastructure , Pedigree , Ubiquitin/analysis , tau Proteins/analysis , tau Proteins/geneticsABSTRACT
Neurofibrillary tangles (NFTs) made up of aggregated tau protein have been identified as the pathologic hallmark of several neurodegenerative diseases including Alzheimer's disease. In vivo detection of NFTs using PET imaging represents a unique opportunity to develop a pharmacodynamic tool to accelerate the discovery of new disease modifying therapeutics targeting tau pathology. Herein, we present the discovery of 6-(fluoro-(18)F)-3-(1H-pyrrolo[2,3-c]pyridin-1-yl)isoquinolin-5-amine, 6 ([(18)F]-MK-6240), as a novel PET tracer for detecting NFTs. 6 exhibits high specificity and selectivity for binding to NFTs, with suitable physicochemical properties and in vivo pharmacokinetics.
Subject(s)
Drug Discovery , Isoquinolines/chemistry , Molecular Imaging , Neurofibrillary Tangles/pathology , Positron-Emission Tomography , Fluorine Radioisotopes/chemistry , Humans , Isoquinolines/chemical synthesis , Isoquinolines/pharmacokinetics , Molecular Structure , Neurofibrillary Tangles/metabolismABSTRACT
The development of common age-related neurodegenerative disorders as Parkinson's disease and Alzheimer's disease (AD) are influenced by genetic factors. Recently, pathogenic mutations in the leucine-rich repeat kinase 2 (LRRK2) gene have been identified in familial Parkinsonism. Individuals in some of these families developed symptoms of dementia with Lewy-bodies and AD. The LRRK2 gene is also located within a locus on chromosome 12 reported in late-onset AD, and is therefore a good candidate gene for dementia. A series of 242 patients from Norway diagnosed clinically with dementia were included in the study, the majority were diagnosed with AD. Individuals were screened for the presence of seven known pathogenic mutations previously reported in the LRRK2 gene. We did not identify LRRK2 mutations in our series of dementia patients, indicating that known pathogenic mutations are not common in patients clinically diagnosed with AD. However, these results do not exclude a possible role of other genetic variants within the LRRK2 gene in AD or other forms of dementia.
Subject(s)
Alzheimer Disease/genetics , Mutation , Protein Serine-Threonine Kinases/genetics , Aged , Aged, 80 and over , Alzheimer Disease/physiopathology , Chromosomes, Human, Pair 12 , Female , Humans , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2 , Male , Middle Aged , Neuropsychological Tests , Parkinson Disease/genetics , Polymorphism, Genetic , Protein Serine-Threonine Kinases/metabolismABSTRACT
In plants, transcribed inverted repeats trigger RNA interference (RNAi) and DNA methylation of identical sequences. RNAi is caused by processing of the double-stranded RNA (dsRNA) transcript into small RNAs that promote degradation of complementary RNA sequences. However, the signals for DNA methylation remain to be fully elucidated. The Arabidopsis tryptophan biosynthetic PAI genes provide an endogenous inverted repeat that triggers DNA methylation of PAI-identical sequences. In the Wassilewskija strain, two PAI genes are arranged as a tail-to-tail inverted repeat and transcribed from an unmethylated upstream promoter. This locus directs its own methylation, as well as methylation of two unlinked singlet PAI genes. Previously, we showed that the locus is likely to make an RNA signal for methylation because suppressed transcription of the inverted repeat leads to reduced PAI methylation. Here we characterize a central rearrangement in the inverted repeat that also confers reduced PAI methylation. The rearrangement creates a premature polyadenylation signal and suppresses readthrough transcription into palindromic PAI sequences. Thus, a likely explanation for the methylation defect of the mutant locus is a failure to produce readthrough dsRNA methylation triggers.
Subject(s)
Arabidopsis/genetics , Arabidopsis/metabolism , Arabidopsis Proteins/genetics , Base Sequence , DNA Methylation , DNA, Plant/chemistry , DNA, Plant/genetics , Gene Rearrangement , Gene Silencing , Genes, Plant , Models, Genetic , Molecular Sequence Data , Mutation , Phenotype , RNA Processing, Post-Transcriptional , RNA, Plant/genetics , RNA, Plant/metabolism , Repetitive Sequences, Nucleic AcidABSTRACT
For late onset Alzheimer's disease (LOAD), the only confirmed, genetic association is with the apolipoprotein E (APOE) locus on chromosome 19. Meta-analysis is often employed to sort the true associations from the false positives. LOAD research has the advantage of a continuously updated meta-analysis of candidate gene association studies in the web-based AlzGene database. The top 30 AlzGene loci on May 1(st), 2007 were investigated in our whole genome association data set consisting of 1411 LOAD cases and neuropathoiogicaiiy verified controls genotyped at 312,316 SNPs using the Affymetrix 500K Mapping Platform. Of the 30 "top AlzGenes", 32 SNPs in 24 genes had odds ratios (OR) whose 95% confidence intervals that did not include 1. Of these 32 SNPs, six were part of the Affymetrix 500K Mapping panel and another ten had proxies on the Affymetrix array that had >80% power to detect an association with α=0.001. Two of these 16 SNPs showed significant association with LOAD in our sample series. One was rs4420638 at the APOE locus (uncorrected p-value=4.58E-37) and the other was rs4293, located in the angiotensin converting enzyme (ACE) locus (uncorrected p-value=0.014). Since this result was nominally significant, but did not survive multiple testing correction for 16 independent tests, this association at rs4293 was verified in a geographically distinct German cohort (p-value=0.03). We present the results of our ACE replication aiongwith a discussion of the statistical limitations of multiple test corrections in whole genome studies.
ABSTRACT
We recently reported evidence for an association between the individual variation in normal human episodic memory and a common variant of the KIBRA gene, KIBRA rs17070145 (T-allele). Since memory impairment is a cardinal clinical feature of Alzheimer's disease (AD), we investigated the possibility of an association between the KIBRA gene and AD using data from neuronal gene expression, brain imaging studies, and genetic association tests. KIBRA was significantly over-expressed and three of its four known binding partners under-expressed in AD-affected hippocampal, posterior cingulate and temporal cortex regions (P<0.010, corrected) in a study of laser-capture microdissected neurons. Using positron emission tomography in a cohort of cognitively normal, late-middle-aged persons genotyped for KIBRA rs17070145, KIBRA T non-carriers exhibited lower glucose metabolism than did carriers in posterior cingulate and precuneus brain regions (P<0.001, uncorrected). Lastly, non-carriers of the KIBRA rs17070145 T-allele had increased risk of late-onset AD in an association study of 702 neuropathologically verified expired subjects (P=0.034; OR=1.29) and in a combined analysis of 1026 additional living and expired subjects (P=0.039; OR=1.26). Our findings suggest that KIBRA is associated with both individual variation in normal episodic memory and predisposition to AD.
Subject(s)
Alzheimer Disease/genetics , Alzheimer Disease/pathology , Brain/pathology , Genetic Predisposition to Disease , Proteins/genetics , Aged , Aged, 80 and over , Alzheimer Disease/complications , Alzheimer Disease/diagnostic imaging , Apolipoproteins E/genetics , Brain/diagnostic imaging , Brain/enzymology , Brain Mapping , Cognition Disorders/etiology , Cognition Disorders/genetics , Female , Gene Expression Profiling/methods , Genome-Wide Association Study/methods , Genotype , Glial Fibrillary Acidic Protein/metabolism , Humans , Intracellular Signaling Peptides and Proteins , Male , Neurons/metabolism , Neurons/pathology , Neuropsychological Tests , Oligonucleotide Array Sequence Analysis/methods , Phosphoproteins , Polymorphism, Single Nucleotide/genetics , Positron-Emission Tomography/methodsABSTRACT
We report the development and validation of experimental methods, study designs, and analysis software for pooling-based genomewide association (GWA) studies that use high-throughput single-nucleotide-polymorphism (SNP) genotyping microarrays. We first describe a theoretical framework for establishing the effectiveness of pooling genomic DNA as a low-cost alternative to individually genotyping thousands of samples on high-density SNP microarrays. Next, we describe software called "GenePool," which directly analyzes SNP microarray probe intensity data and ranks SNPs by increased likelihood of being genetically associated with a trait or disorder. Finally, we apply these methods to experimental case-control data and demonstrate successful identification of published genetic susceptibility loci for a rare monogenic disease (sudden infant death with dysgenesis of the testes syndrome), a rare complex disease (progressive supranuclear palsy), and a common complex disease (Alzheimer disease) across multiple SNP genotyping platforms. On the basis of these theoretical calculations and their experimental validation, our results suggest that pooling-based GWA studies are a logical first step for determining whether major genetic associations exist in diseases with high heritability.
Subject(s)
Genome, Human , Models, Genetic , Polymorphism, Single Nucleotide/genetics , Software , Alzheimer Disease/genetics , Case-Control Studies , Computer Simulation , Genetic Markers , Genotype , Gonadal Dysgenesis/genetics , Humans , Male , Oligonucleotide Array Sequence Analysis , Research Design , Supranuclear Palsy, Progressive/genetics , Syndrome , Testis/abnormalitiesABSTRACT
To date, only the H1 MAPT haplotype has been consistently associated with risk of developing the neurodegenerative disease progressive supranuclear palsy (PSP). We hypothesized that additional genetic loci may be involved in conferring risk of PSP that could be identified through a pooling-based genomewide association study of >500,000 SNPs. Candidate SNPs with large differences in allelic frequency were identified by ranking all SNPs by their probe-intensity difference between cohorts. The MAPT H1 haplotype was strongly detected by this methodology, as was a second major locus on chromosome 11p12-p11 that showed evidence of association at allelic (P<.001), genotypic (P<.001), and haplotypic (P<.001) levels and was narrowed to a single haplotype block containing the DNA damage-binding protein 2 (DDB2) and lysosomal acid phosphatase 2 (ACP2) genes. Since DNA damage and lysosomal dysfunction have been implicated in aging and neurodegenerative processes, both genes are viable candidates for conferring risk of disease.
Subject(s)
Chromosomes, Human, Pair 11/genetics , Genetic Predisposition to Disease , Polymorphism, Single Nucleotide/genetics , Supranuclear Palsy, Progressive/genetics , Acid Phosphatase/genetics , DNA-Binding Proteins/genetics , Genomics/methods , HumansABSTRACT
Null mutations in the progranulin gene (PGRN) were recently reported to cause tau-negative frontotemporal dementia linked to chromosome 17. We assessed the genetic contribution of PGRN mutations in an extended population of patients with frontotemporal lobar degeneration (FTLD) (N=378). Mutations were identified in 10% of the total FTLD population and 23% of patients with a positive family history. This mutation frequency dropped to 5% when analysis was restricted to an unbiased FTLD subpopulation (N=167) derived from patients referred to Alzheimer's Disease Research Centers (ADRC). Among the ADRC patients, PGRN mutations were equally frequent as mutations in the tau gene (MAPT). We identified 23 different pathogenic PGRN mutations, including a total of 21 nonsense, frameshift and splice-site mutations that cause premature termination of the coding sequence and degradation of the mutant RNA by nonsense-mediated decay. We also observed an unusual splice-site mutation in the exon 1 5' splice site, which leads to loss of the Kozac sequence, and a missense mutation in the hydrophobic core of the PGRN signal peptide. Both mutations revealed novel mechanisms that result in loss of functional PGRN. One mutation, c.1477C>T (p.Arg493X), was detected in eight independently ascertained familial FTLD patients who were shown to share a common extended haplotype over the PGRN genomic region. Clinical examination of patients with PGRN mutations revealed highly variable onset ages with language dysfunction as a common presenting symptom. Neuropathological examination showed FTLD with ubiquitin-positive cytoplasmic and intranuclear inclusions in all PGRN mutation carriers.