Mid-to-Late Life Healthy Lifestyle Modifies Genetic Risk for Longitudinal Cognitive Aging among Asymptomatic Individuals.

IMPORTANCE: Genetic and lifestyle factors contribute to an individual's risk of developing Alzheimer's disease. However, it is unknown whether and how adherence to healthy lifestyles can mitigate the genetic risk of Alzheimer's. OBJECTIVE: The aim of this study is to investigate whether adherence to healthy lifestyles can modify the impact of genetic predisposition to Alzheimer's disease on later-life cognitive decline. DESIGN SETTING AND PARTICIPANTS: This prospective cohort study included 891 adults of European ancestry, aged 40 to 65, who were without dementia and had complete healthy-lifestyle and cognition data during the follow-up. Participants joined the Wisconsin Registry for Alzheimer's Prevention (WRAP) beginning in 2001. We conducted replication analyses using a subsample with similar baseline age range from the Health and Retirement Study (HRS). EXPOSURES: We assessed participants' exposures using a continuous non-APOE polygenic risk score for Alzheimer's, a binary indicator for APOE-ε4 carrier status, and a weighted healthy-lifestyle score, including factors such as no current smoking, regular physical activity, healthy diet, light to moderate alcohol consumption, and frequent cognitive activities. MAIN OUTCOMES AND MEASURES: We z-standardized cognitive scores for global (Preclinical Alzheimer's Cognitive Composite score 3 - PACC3) and domain-specific assessments (delayed recall and immediate learning). RESULTS: We followed 891 individuals for up to 10 years (mean [SD] baseline age, 58 [6] years, 31% male, 38% APOE-ε4 carriers). After false discovery rate (FDR) correction, we found statistically significant PRS × lifestyle × age interactions on preclinical cognitive decline but the evidence is stronger among APOE-ε4 carriers. Among APOE-ε4 carriers, PRS-related differences in overall and memory-related domains between people scoring 0-1 and 4-5 regarding healthy lifestyles became evident around age 67 after FDR correction. These findings were robust across several sensitivity analyses and were replicated in the population-based HRS. CONCLUSION: A favorable lifestyle can mitigate the genetic risk associated with current known non-APOE genetic variants for longitudinal cognitive decline, and these protective effects are particularly pronounced among APOE-ε4 carriers.

Assessing the Biological Mechanisms Linking Smoking Behavior and Cognitive Function: A Mediation Analysis of Untargeted Metabolomics.

(1) Smoking is the most significant preventable health hazard in the modern world. It increases the risk of vascular problems, which are also risk factors for dementia. In addition, toxins in cigarettes increase oxidative stress and inflammation, which have both been linked to the development of Alzheimer's disease and related dementias (ADRD). This study identified potential mechanisms of the smoking-cognitive function relationship using metabolomics data from the longitudinal Wisconsin Registry for Alzheimer's Prevention (WRAP). (2) 1266 WRAP participants were included to assess the association between smoking status and four cognitive composite scores. Next, untargeted metabolomic data were used to assess the relationships between smoking and metabolites. Metabolites significantly associated with smoking were then tested for association with cognitive composite scores. Total effect models and mediation models were used to explore the role of metabolites in smoking-cognitive function pathways. (3) Plasma N-acetylneuraminate was associated with smoking status Preclinical Alzheimer Cognitive Composite 3 (PACC3) and Immediate Learning (IMM). N-acetylneuraminate mediated 12% of the smoking-PACC3 relationship and 13% of the smoking-IMM relationship. (4) These findings provide links between previous studies that can enhance our understanding of potential biological pathways between smoking and cognitive function.

CSF metabolites associated with biomarkers of Alzheimer's disease pathology.

Introduction: Metabolomics technology facilitates studying associations between small molecules and disease processes. Correlating metabolites in cerebrospinal fluid (CSF) with Alzheimer's disease (AD) CSF biomarkers may elucidate additional changes that are associated with early AD pathology and enhance our knowledge of the disease. Methods: The relative abundance of untargeted metabolites was assessed in 161 individuals from the Wisconsin Registry for Alzheimer's Prevention. A metabolome-wide association study (MWAS) was conducted between 269 CSF metabolites and protein biomarkers reflecting brain amyloidosis, tau pathology, neuronal and synaptic degeneration, and astrocyte or microglial activation and neuroinflammation. Linear mixed-effects regression analyses were performed with random intercepts for sample relatedness and repeated measurements and fixed effects for age, sex, and years of education. The metabolome-wide significance was determined by a false discovery rate threshold of 0.05. The significant metabolites were replicated in 154 independent individuals from then Wisconsin Alzheimer's Disease Research Center. Mendelian randomization was performed using genome-wide significant single nucleotide polymorphisms from a CSF metabolites genome-wide association study. Results: Metabolome-wide association study results showed several significantly associated metabolites for all the biomarkers except Aß42/40 and IL-6. Genetic variants associated with metabolites and Mendelian randomization analysis provided evidence for a causal association of metabolites for soluble triggering receptor expressed on myeloid cells 2 (sTREM2), amyloid ß (Aß40), α-synuclein, total tau, phosphorylated tau, and neurogranin, for example, palmitoyl sphingomyelin (d18:1/16:0) for sTREM2, and erythritol for Aß40 and α-synuclein. Discussion: This study provides evidence that CSF metabolites are associated with AD-related pathology, and many of these associations may be causal.

Identification of plasma metabolites associated with modifiable risk factors and endophenotypes reflecting Alzheimer's disease pathology.

Modifiable factors can influence the risk for Alzheimer's disease (AD) and serve as targets for intervention; however, the biological mechanisms linking these factors to AD are unknown. This study aims to identify plasma metabolites associated with modifiable factors for AD, including MIND diet, physical activity, smoking, and caffeine intake, and test their association with AD endophenotypes to identify their potential roles in pathophysiological mechanisms. The association between each of the 757 plasma metabolites and four modifiable factors was tested in the wisconsin registry for Alzheimer's prevention cohort of initially cognitively unimpaired, asymptomatic middle-aged adults. After Bonferroni correction, the significant plasma metabolites were tested for association with each of the AD endophenotypes, including twelve cerebrospinal fluid (CSF) biomarkers, reflecting key pathophysiologies for AD, and four cognitive composite scores. Finally, causal mediation analyses were conducted to evaluate possible mediation effects. Analyses were performed using linear mixed-effects regression. A total of 27, 3, 23, and 24 metabolites were associated with MIND diet, physical activity, smoking, and caffeine intake, respectively. Potential mediation effects include beta-cryptoxanthin in the association between MIND diet and preclinical Alzheimer cognitive composite score, hippurate between MIND diet and immediate learning, glutamate between physical activity and CSF neurofilament light, and beta-cryptoxanthin between smoking and immediate learning. Our study identified several plasma metabolites that are associated with modifiable factors. These metabolites can be employed as biomarkers for tracking these factors, and they provide a potential biological pathway of how modifiable factors influence the human body and AD risk.

Multi-method investigation of factors influencing amyloid onset and impairment in three cohorts.

Alzheimer's disease biomarkers are becoming increasingly important for characterizing the longitudinal course of disease, predicting the timing of clinical and cognitive symptoms, and for recruitment and treatment monitoring in clinical trials. In this work, we develop and evaluate three methods for modelling the longitudinal course of amyloid accumulation in three cohorts using amyloid PET imaging. We then use these novel approaches to investigate factors that influence the timing of amyloid onset and the timing from amyloid onset to impairment onset in the Alzheimer's disease continuum. Data were acquired from the Alzheimer's Disease Neuroimaging Initiative (ADNI), the Baltimore Longitudinal Study of Aging (BLSA) and the Wisconsin Registry for Alzheimer's Prevention (WRAP). Amyloid PET was used to assess global amyloid burden. Three methods were evaluated for modelling amyloid accumulation using 10-fold cross-validation and holdout validation where applicable. Estimated amyloid onset age was compared across all three modelling methods and cohorts. Cox regression and accelerated failure time models were used to investigate whether sex, apolipoprotein E genotype and e4 carriage were associated with amyloid onset age in all cohorts. Cox regression was used to investigate whether apolipoprotein E (e4 carriage and e3e3, e3e4, e4e4 genotypes), sex or age of amyloid onset were associated with the time from amyloid onset to impairment onset (global clinical dementia rating ≥1) in a subset of 595 ADNI participants that were not impaired before amyloid onset. Model prediction and estimated amyloid onset age were similar across all three amyloid modelling methods. Sex and apolipoprotein E e4 carriage were not associated with PET-measured amyloid accumulation rates. Apolipoprotein E genotype and e4 carriage, but not sex, were associated with amyloid onset age such that e4 carriers became amyloid positive at an earlier age compared to non-carriers, and greater e4 dosage was associated with an earlier amyloid onset age. In the ADNI, e4 carriage, being female and a later amyloid onset age were all associated with a shorter time from amyloid onset to impairment onset. The risk of impairment onset due to age of amyloid onset was non-linear and accelerated for amyloid onset age >65. These findings demonstrate the feasibility of modelling longitudinal amyloid accumulation to enable individualized estimates of amyloid onset age from amyloid PET imaging. These estimates provide a more direct way to investigate the role of amyloid and other factors that influence the timing of clinical impairment in Alzheimer's disease.

The Survey of the Health of Wisconsin (SHOW) Program: An Infrastructure for Advancing Population Health.

Introduction: The Survey of the Health of Wisconsin (SHOW) was established in 2008 by the University of Wisconsin (UW) School of Medicine and Public Health (SMPH) with the goals of (1) providing a timely and accurate picture of the health of the state residents; and (2) serving as an agile resource infrastructure for ancillary studies. Today, the SHOW program continues to serve as a unique and vital population health research infrastructure for advancing public health. Methods: SHOW currently includes 5,846 adult and 980 minor participants recruited between 2008 and 2019 in four primary waves. WAVE I (2008-2013) includes annual statewide representative samples of 3,380 adults ages 21 to 74 years. WAVE II (2014-2016) is a triannual statewide sample of 1,957 adults (age ≥18 years) and 645 children (age 0-17). WAVE III (2017) consists of follow-up of 725 adults from the WAVE I and baseline surveys of 222 children in selected households. WAVEs II and III include stool samples collected as part of an ancillary study in a subset of 784 individuals. WAVE IV consists of 517 adults and 113 children recruited from traditionally under-represented populations in biomedical research including African Americans and Hispanics in Milwaukee, Wisconsin. Findings to Date: The SHOW resource provides unique spatially granular and timely data to examine the intersectionality of multiple social determinants and population health. SHOW includes a large biorepository and extensive health data collected in a geographically diverse urban and rural population. Over 60 studies have been published covering a broad range of topics including, urban and rural disparities in cardio-metabolic disease and cancer, objective physical activity, sleep, green-space and mental health, transcriptomics, the gut microbiome, antibiotic resistance, air pollution, concentrated animal feeding operations and heavy metal exposures. Discussion: The SHOW cohort and resource is available for continued follow-up and ancillary studies including longitudinal public health monitoring, translational biomedical research, environmental health, aging, microbiome and COVID-19 research.

Exploring common genetic contributors to neuroprotection from amyloid pathology.

Preclinical Alzheimer's disease describes some individuals who harbour Alzheimer's pathologies but are asymptomatic. For this study, we hypothesized that genetic variation may help protect some individuals from Alzheimer's-related neurodegeneration. We therefore conducted a genome-wide association study using 5 891 064 common variants to assess whether genetic variation modifies the association between baseline beta-amyloid, as measured by both cerebrospinal fluid and positron emission tomography, and neurodegeneration defined using MRI measures of hippocampal volume. We combined and jointly analysed genotype, biomarker and neuroimaging data from non-Hispanic white individuals who were enrolled in four longitudinal ageing studies (n = 1065). Using regression models, we examined the interaction between common genetic variants (Minor Allele Frequency >0.01), including APOE-ɛ4 and APOE-ɛ2, and baseline cerebrospinal levels of amyloid (CSF Aß42) on baseline hippocampal volume and the longitudinal rate of hippocampal atrophy. For targeted replication of top findings, we analysed an independent dataset (n = 808) where amyloid burden was assessed by Pittsburgh Compound B ([11C]-PiB) positron emission tomography. In this study, we found that APOE-ɛ4 modified the association between baseline CSF Aß42 and hippocampal volume such that APOE-ɛ4 carriers showed more rapid atrophy, particularly in the presence of enhanced amyloidosis. We also identified a novel locus on chromosome 3 that interacted with baseline CSF Aß42. Minor allele carriers of rs62263260, an expression quantitative trait locus for the SEMA5B gene (P = 1.46 × 10-8; 3:122675327) had more rapid neurodegeneration when amyloid burden was high and slower neurodegeneration when amyloid was low. The rs62263260 × amyloid interaction on longitudinal change in hippocampal volume was replicated in an independent dataset (P = 0.0112) where amyloid burden was assessed by positron emission tomography. In addition to supporting the established interaction between APOE and amyloid on neurodegeneration, our study identifies a novel locus that modifies the association between beta-amyloid and hippocampal atrophy. Annotation results may implicate SEMA5B, a gene involved in synaptic pruning and axonal guidance, as a high-quality candidate for functional confirmation and future mechanistic analysis.

Crosswalk study on blood collection-tube types for Alzheimer's disease biomarkers.

INTRODUCTION: Blood-based Alzheimer's disease (AD) biomarkers show promise, but pre-analytical protocol differences may pose problems. We examined seven AD blood biomarkers (amyloid beta [ A ß ] 42 , A ß 40 , phosphorylated tau [ p - ta u 181 , total tau [t-tau], neurofilament light chain [NfL], A ß 42 40 , and p - ta u 181 A ß 42 ) in three collection tube types (ethylenediaminetetraacetic acid [EDTA] plasma, heparin plasma, serum). METHODS: Plasma and serum were obtained from cerebrospinal fluid or amyloid positron emission tomography-positive and -negative participants (N = 38) in the Wisconsin Registry for Alzheimer's Prevention. We modeled AD biomarker values observed in EDTA plasma versus heparin plasma and serum, and assessed correspondence with brain amyloidosis. RESULTS: Results suggested bias due to tube type, but crosswalks are possible for some analytes, with excellent model fit for NfL ( R 2 = 0.94), adequate for amyloid ( R 2 = 0.40-0.69), and weaker for t-tau ( R 2 = 0.04-0.42) and p - ta u 181 ( R 2 = 0.22-0.29). Brain amyloidosis differentiated several measures, especially EDTA plasma pTa u 181 A ß 42 ( d = 1.29). DISCUSSION: AD biomarker concentrations vary by tube type. However, correlations for some biomarkers support harmonization across types, suggesting cautious optimism for use in banked blood.

The Survey of the Health of Wisconsin (SHOW) Program: An infrastructure for Advancing Population Health Sciences.

PURPOSE: The Survey of the Health of Wisconsin (SHOW) was established in 2008 by the University of Wisconsin (UW) School of Medicine and Public Health (SMPH) with the goals of 1) providing a timely and accurate picture of the health of the state residents; and 2) serving as an agile resource infrastructure for ancillary studies. Today SHOW continues to serve as a vital population health research infrastructure. PARTICIPANTS: SHOW currently includes 5,846 adult and 980 minor participants recruited between 2008-2019 in four primary waves. WAVE I (2008-2013) includes annual statewide representative samples of 3,380 adults ages 21 to 74 years. WAVE II (2014-2016) is a triannual statewide sample of 1957 adults (age ≥18 years) and 645 children. WAVE III (2017) consists of follow-up of 725 adults from the WAVE I and baseline surveys of 222 children in selected households. WAVEs II and III include stool samples collected as part of an ancillary study in a subset of 784 individuals. WAVE IV consist of 517 adults and 113 children recruited from traditionally under-represented populations in biomedical research including African Americans and Hispanics in Milwaukee county, WI. FINDINGS TO DATE: The SHOW provides extensive data to examine the intersectionality of multiple social determinants and population health. SHOW includes a large biorepository and extensive health data collected in a geographically diverse urban and rural population. Over 60 studies have been published covering a broad range of topics including, urban and rural disparities in cardio-metabolic disease and cancer, objective physical activity, sleep, green-space and mental health, transcriptomics, the gut microbiome, antibiotic resistance, air pollution, concentrated animal feeding operations and heavy metal exposures. FUTURE PLANS: The SHOW cohort is available for continued longitudinal follow-up and ancillary studies including genetic, multi-omic and translational environmental health, aging, microbiome and COVID-19 research. ARTICLE SUMMARY: Strengths and limitations: The Survey of the Health of Wisconsin (SHOW) is an infrastructure to advance population health sciences including biological sample collection and broader data on individual and neighborhood social and environmental determinants of health.The extensive data from diverse urban and rural populations offers a unique study sample to compare how socio-economic gradients shape health outcomes in different contexts.The objective health data supports novel interdisciplinary research initiatives and is especially suited for research in causes and consequences of environmental exposures (physical, chemical, social) across the life course on cardiometabolic health, immunity, and aging related conditions.The extensive biorepository supports novel omics research into common biological mechanisms underlying numerous complex chronic conditions including inflammation, oxidative stress, metabolomics, and epigenetic modulation.Ancillary studies, such as the Wisconsin Microbiome Study, have expanded the utility of the study to examine human susceptibility to environmental exposures and opportunities for investigations of the role of microbiome in health and disease.Long-standing partnerships and recent participation among traditionally under-represented populations in biomedical research offer numerous opportunities to support community-driven health equity work.No biological samples were collected among children.The statewide sampling frame may limit generalizability to other regions in the United States.

Metabolites Associated with Early Cognitive Changes Implicated in Alzheimer's Disease.

BACKGROUND: Understanding metabolic mechanisms associated with cognitive changes preceding an Alzheimer's disease (AD) diagnosis could advance our understanding of AD progression and inform preventive methods. OBJECTIVE: We investigated the metabolomics of the early changes in executive function and delayed recall, the earliest aspects of cognitive function to change in the course of AD development, in order to better understand mechanisms that could contribute to early stages and progression of this disease. METHODS: This investigation used longitudinal plasma samples from the Wisconsin Registry for Alzheimer's Prevention (WRAP), a cohort of participants who were dementia free at enrollment and enriched with a parental history of AD. Metabolomic profiles were quantified for 2,324 fasting plasma samples among 1,200 participants, each with up to three study visits, which occurred every two years. Metabolites were individually tested for association with executive function and delayed recall trajectories across age. RESULTS: Of 1,097 metabolites tested, levels of seven were associated with executive function trajectories, including an amino acid cysteine S-sulfate and three fatty acids, including erucate (22 : 1n9), while none were associated with delayed recall trajectories. Replication was attempted for four of these metabolites that were present in the Vietnam Era Twin Study of Aging (VETSA). Although none reached statistical significance, three of these associations showed consistent effectdirections. CONCLUSION: Our results suggest potential metabolomic mechanisms that could contribute to the earliest signs of cognitive decline. In particular, fatty acids may be associated with cognition in a manner that is more complex than previously suspected.

Association Between Common Variants in RBFOX1, an RNA-Binding Protein, and Brain Amyloidosis in Early and Preclinical Alzheimer Disease.

Importance: Genetic studies of Alzheimer disease have focused on the clinical or pathologic diagnosis as the primary outcome, but little is known about the genetic basis of the preclinical phase of the disease. Objective: To examine the underlying genetic basis for brain amyloidosis in the preclinical phase of Alzheimer disease. Design, Setting, and Participants: In the first stage of this genetic association study, a meta-analysis was conducted using genetic and imaging data acquired from 6 multicenter cohort studies of healthy older individuals between 1994 and 2019: the Anti-Amyloid Treatment in Asymptomatic Alzheimer Disease Study, the Berkeley Aging Cohort Study, the Wisconsin Registry for Alzheimer's Prevention, the Biomarkers of Cognitive Decline Among Normal Individuals cohort, the Baltimore Longitudinal Study of Aging, and the Alzheimer Disease Neuroimaging Initiative, which included Alzheimer disease and mild cognitive impairment. The second stage was designed to validate genetic observations using pathologic and clinical data from the Religious Orders Study and Rush Memory and Aging Project. Participants older than 50 years with amyloid positron emission tomographic (PET) imaging data and DNA from the 6 cohorts were included. The largest cohort, the Anti-Amyloid Treatment in Asymptomatic Alzheimer Disease Study (n = 3154), was the PET screening cohort used for a secondary prevention trial designed to slow cognitive decline associated with brain amyloidosis. Six smaller, longitudinal cohort studies (n = 1160) provided additional amyloid PET imaging data with existing genetic data. The present study was conducted from March 29, 2019, to February 19, 2020. Main Outcomes and Measures: A genome-wide association study of PET imaging amyloid levels. Results: From the 4314 analyzed participants (age, 52-96 years; 2478 participants [57%] were women), a novel locus for amyloidosis was noted within RBFOX1 (ß = 0.61, P = 3 × 10-9) in addition to APOE. The RBFOX1 protein localized around plaques, and reduced expression of RBFOX1 was correlated with higher amyloid-ß burden (ß = -0.008, P = .002) and worse cognition (ß = 0.007, P = .006) during life in the Religious Orders Study and Rush Memory and Aging Project cohort. Conclusions and Relevance: RBFOX1 encodes a neuronal RNA-binding protein known to be expressed in neuronal tissues and may play a role in neuronal development. The findings of this study suggest that RBFOX1 is a novel locus that may be involved in the pathogenesis of Alzheimer disease.

Association between vitamin D status and age-related macular degeneration by genetic risk.

IMPORTANCE: Deficient 25-hydroxyvitamin D (25[OH]D) concentrations have been associated with increased odds of age-related macular degeneration (AMD). OBJECTIVE: To examine whether this association is modified by genetic risk for AMD and whether there is an association between AMD and single-nucleotide polymorphisms of genes involved in vitamin D transport, metabolism, and genomic function. DESIGN, SETTING, AND PARTICIPANTS: Postmenopausal women (N = 913) who were participants of the Carotenoids in Age-Related Eye Disease Study (CAREDS) (aged 54 to <75 years) with available serum 25(OH)D concentrations (assessed October 1, 1993, to December 31, 1998), genetic data, and measures of AMD (n = 142) assessed at CAREDS baseline from May 14, 2001, through January 31, 2004, were studied. MAIN OUTCOMES AND MEASURES: Prevalent early or late AMD was determined from graded, stereoscopic fundus photographs. Logistic regression was used to estimate odds ratios (ORs) and 95% CIs for AMD by the joint effects of 25(OH)D (<12, ≥12 to <20, ≥20 to <30, and ≥30 ng/mL) and risk genotype (noncarrier, 1 risk allele, or 2 risk alleles). The referent group was noncarriers with adequate vitamin D status (≥30 ng/mL). Joint effect ORs were adjusted for age, smoking, iris pigmentation, self-reported cardiovascular disease, self-reported diabetes status, and hormone use. Additive and multiplicative interactions were assessed using the synergy index (SI) and an interaction term, respectively. To examine the association between AMD and variants in vitamin D-related genes, age-adjusted ORs and 95% CIs were estimated using logistic regression. RESULTS: Among the 913 women, 550 had adequate levels of vitamin D (≥20 ng/mL), 275 had inadequate levels (≥12 to <20 mg/mL), and 88 had deficient levels (<12 ng/mL). A 6.7-fold increased odds of AMD (95% CI, 1.6-28.2) was observed among women with deficient vitamin D status (25[OH]D <12 ng/mL) and 2 risk alleles for CFH Y402H (SI for additive interaction, 1.4; 95% CI, 1.1-1.7; P for multiplicative interaction = .25). Significant additive (SI, 1.4; 95% CI, 1.1-1.7) and multiplicative interactions (P = .02) were observed for deficient women with 2 high-risk CFI (rs10033900) alleles (OR, 6.3; 95% CI, 1.6-24.2). The odds of AMD did not differ by genotype of candidate vitamin D genes. CONCLUSIONS AND RELEVANCE: In this study, the odds of AMD were highest in those with deficient vitamin D status and 2 risk alleles for the CFH and CFI genotypes, suggesting a synergistic effect between vitamin D status and complement cascade protein function. Limited sample size led to wide CIs. Findings may be due to chance or explained by residual confounding.

Reproductive windows, genetic loci, and breast cancer risk.

PURPOSE: The reproductive windows between age at menarche and age at first birth (standardized age at first birth) and from menarche to menopause (reproductive lifespan) may interact with genetic variants in association with breast cancer risk. METHODS: We assessed this hypothesis in 6131 breast cancer cases and 7274 controls who participated in the population-based Collaborative Breast Cancer Study. Risk factor information was collected through telephone interviews, and DNA samples were collected on a subsample (N= 1484 cases, 1307 controls) to genotype for 13 genome-wide association study-identified loci. Adjusted odds ratios (ORs) and 95% confidence intervals (CIs) were calculated, and P values for the interaction between reproductive windows and genotypes were obtained by adding cross-product terms to statistical models. RESULTS: For standardized age at first birth, the OR was 1.52 (CI, 1.36-1.71) comparing the highest quintile with the lowest quintile. Carrier status for rs10941679 (5p12) and rs10483813 (RAD51B) appeared to modify this relationship (P = .04 and P = .02, respectively). For reproductive lifespan, the OR comparing the highest quintile with the lowest quintiles was 1.62 (CI, 1.35-1.95). No interactions were detected between genotype and reproductive lifespan (all P > .05). All results were similar regardless of ductal versus lobular breast cancer subtype. CONCLUSIONS: Our results suggest that the reproductive windows are associated with breast cancer risk and that associations may vary by genetic variants.

The association of telomere length with colorectal cancer differs by the age of cancer onset.

OBJECTIVES: Telomeres are nucleoprotein structures that cap the end of chromosomes and shorten with sequential cell divisions in normal aging. Short telomeres are also implicated in the incidence of many cancers, but the evidence is not conclusive for colorectal cancer (CRC). Therefore, the aim of this study was to assess the association of CRC and telomere length. METHODS: In this case-control study, we measured relative telomere length from peripheral blood leukocytes (PBLs) DNA with quantitative PCR in 598 CRC patients and 2,212 healthy controls. RESULTS: Multivariate analysis indicated that telomere length was associated with risk for CRC, and this association varied in an age-related manner; younger individuals (≤50 years of age) with longer telomeres (80-99 percentiles) had a 2-6 times higher risk of CRC, while older individuals (>50 years of age) with shortened telomeres (1-10 percentiles) had 2-12 times the risk for CRC. The risk for CRC varies with extremes in telomere length in an age-associated manner. CONCLUSIONS: Younger individuals with longer telomeres or older individuals with shorter telomeres are at higher risk for CRC. These findings indicate that the association of PBL telomere length varies according to the age of cancer onset and that CRC is likely associated with at minimum two different mechanisms of telomere dynamics.

Genetic evidence for role of carotenoids in age-related macular degeneration in the Carotenoids in Age-Related Eye Disease Study (CAREDS).

PURPOSE: We tested variants in genes related to lutein and zeaxanthin status for association with age-related macular degeneration (AMD) in the Carotenoids in Age-Related Eye Disease Study (CAREDS). METHODS: Of 2005 CAREDS participants, 1663 were graded for AMD from fundus photography and genotyped for 424 single nucleotide polymorphisms (SNPs) from 24 candidate genes for carotenoid status. Of 337 AMD cases 91% had early or intermediate AMD. The SNPs were tested individually for association with AMD using logistic regression. A carotenoid-related genetic risk model was built using backward selection and compared to existing AMD risk factors using the area under the receiver operating characteristic curve (AUC). RESULTS: A total of 24 variants from five genes (BCMO1, BCO2, NPCL1L1, ABCG8, and FADS2) not previously related to AMD and four genes related to AMD in previous studies (SCARB1, ABCA1, APOE, and ALDH3A2) were associated independently with AMD, after adjusting for age and ancestry. Variants in all genes (not always the identical SNPs) were associated with lutein and zeaxanthin in serum and/or macula, in this or other samples, except for BCO2 and FADS2. A genetic risk score including nine variants significantly (P = 0.002) discriminated between AMD cases and controls beyond age, smoking, CFH Y402H, and ARMS2 A69S. The odds ratio (95% confidence interval) for AMD among women in the highest versus lowest quintile for the risk score was 3.1 (2.0-4.9). CONCLUSIONS: Variants in genes related to lutein and zeaxanthin status were associated with AMD in CAREDS, adding to the body of evidence supporting a protective role of lutein and zeaxanthin in risk of AMD.

Breast cancer susceptibility loci in association with age at menarche, age at natural menopause and the reproductive lifespan.

BACKGROUND: Genome-wide association studies have identified single nucleotide polymorphisms (SNPs) associated with breast cancer risk. Some of these loci have unknown functional significance and may mediate the effects of hormonal exposures on breast cancer risk. We examined relationships between breast cancer susceptibility variants and menstrual/reproductive factors using data from two population-based studies. METHODS: The first analysis was based on a sample of 1328 women age 20-74 who participated as controls in a case-control study of breast cancer conducted in three U.S. states. We evaluated the associations between age at menarche, age at natural menopause and the reproductive lifespan with 13 previously identified breast cancer variants. Associations were also examined with a genetic score created as the sum of at-risk alleles across the 13 variants. For validation, significant results were evaluated in a second dataset comprised 1353 women age 43-86 recruited as part of a cohort study in Wisconsin. RESULTS: Neither the genetic score nor any of the 13 variants considered individually were associated with age at menarche or reproductive lifespan. Two SNPs were associated with age at natural menopause; every increase in the minor allele (A) of rs17468277 (CASP8) was associated with a 1.12 year decrease in menopause age (p=0.02). The minor allele (G) of rs10941679 (5p12) was associated with a 1.01 year increase in age at natural menopause (p=0.01). The results were not replicated in the validation cohort (B=-0.61, p=0.14 and B=-0.01, p=.0.98, respectively). CONCLUSIONS: The evaluated variants and reproductive experiences may work through separate pathways to influence breast cancer risk.

Correlation of chromosomal instability, telomere length and telomere maintenance in microsatellite stable rectal cancer: a molecular subclass of rectal cancer.

INTRODUCTION: Colorectal cancer (CRC) tumor DNA is characterized by chromosomal damage termed chromosomal instability (CIN) and excessively shortened telomeres. Up to 80% of CRC is microsatellite stable (MSS) and is historically considered to be chromosomally unstable (CIN+). However, tumor phenotyping depicts some MSS CRC with little or no genetic changes, thus being chromosomally stable (CIN-). MSS CIN- tumors have not been assessed for telomere attrition. EXPERIMENTAL DESIGN: MSS rectal cancers from patients ≤50 years old with Stage II (B2 or higher) or Stage III disease were assessed for CIN, telomere length and telomere maintenance mechanism (telomerase activation [TA]; alternative lengthening of telomeres [ALT]). Relative telomere length was measured by qPCR in somatic epithelial and cancer DNA. TA was measured with the TRAPeze assay, and tumors were evaluated for the presence of C-circles indicative of ALT. p53 mutation status was assessed in all available samples. DNA copy number changes were evaluated with Spectral Genomics aCGH. RESULTS: Tumors were classified as chromosomally stable (CIN-) and chromosomally instable (CIN+) by degree of DNA copy number changes. CIN- tumors (35%; n=6) had fewer copy number changes (<17% of their clones with DNA copy number changes) than CIN+ tumors (65%; n=13) which had high levels of copy number changes in 20% to 49% of clones. Telomere lengths were longer in CIN- compared to CIN+ tumors (p=0.0066) and in those in which telomerase was not activated (p=0.004). Tumors exhibiting activation of telomerase had shorter tumor telomeres (p=0.0040); and tended to be CIN+ (p=0.0949). CONCLUSIONS: MSS rectal cancer appears to represent a heterogeneous group of tumors that may be categorized both on the basis of CIN status and telomere maintenance mechanism. MSS CIN- rectal cancers appear to have longer telomeres than those of MSS CIN+ rectal cancers and to utilize ALT rather than activation of telomerase.

Telomere length varies by DNA extraction method: implications for epidemiologic research.

BACKGROUND: Both shorter and longer telomeres in peripheral blood leukocyte (PBL) DNA have been associated with cancer risk. However, associations remain inconsistent across studies of the same cancer type. This study compares DNA preparation methods to determine telomere length from patients with colorectal cancer. METHODS: We examined PBL relative telomere length (RTL) measured by quantitative PCR (qPCR) in 1,033 patients with colorectal cancer and 2,952 healthy controls. DNA was extracted with phenol/chloroform, PureGene, or QIAamp. RESULTS: We observed differences in RTL depending on DNA extraction method (P < 0.001). Phenol/chloroform-extracted DNA had a mean RTL (T/S ratio) of 0.78 (range 0.01-6.54) compared with PureGene-extracted DNA (mean RTL of 0.75; range 0.00-12.33). DNA extracted by QIAamp yielded a mean RTL of 0.38 (range 0.02-3.69). We subsequently compared RTL measured by qPCR from an independent set of 20 colorectal cancer cases and 24 normal controls in PBL DNA extracted by each of the three extraction methods. The range of RTL measured by qPCR from QIAamp-extracted DNA (0.17-0.58) was less than from either PureGene or phenol/chloroform (ranges, 0.04-2.67 and 0.32-2.81, respectively). CONCLUSIONS: RTL measured by qPCR from QIAamp-extracted DNA was less than from either PureGene or phenol/chloroform (P < 0.001). IMPACT: Differences in DNA extraction method may contribute to the discrepancies between studies seeking to find an association between the risk of cancer or other diseases and RTL.

The associations between a polygenic score, reproductive and menstrual risk factors and breast cancer risk.

We evaluated whether 13 single nucleotide polymorphisms (SNPs) identified in genome-wide association studies interact with one another and with reproductive and menstrual risk factors in association with breast cancer risk. DNA samples and information on parity, breastfeeding, age at menarche, age at first birth, and age at menopause were collected through structured interviews from 1,484 breast cancer cases and 1,307 controls who participated in a population-based case-control study conducted in three US states. A polygenic score was created as the sum of risk allele copies multiplied by the corresponding log odds estimate. Logistic regression was used to test the associations between SNPs, the score, reproductive and menstrual factors, and breast cancer risk. Nonlinearity of the score was assessed by the inclusion of a quadratic term for polygenic score. Interactions between the aforementioned variables were tested by including a cross-product term in models. We confirmed associations between rs13387042 (2q35), rs4973768 (SLC4A7), rs10941679 (5p12), rs2981582 (FGFR2), rs3817198 (LSP1), rs3803662 (TOX3), and rs6504950 (STXBP4) with breast cancer. Women in the score's highest quintile had 2.2-fold increased risk when compared to women in the lowest quintile (95 % confidence interval: 1.67-2.88). The quadratic polygenic score term was not significant in the model (p = 0.85), suggesting that the established breast cancer loci are not associated with increased risk more than the sum of risk alleles. Modifications of menstrual and reproductive risk factors associations with breast cancer risk by polygenic score were not observed. Our results suggest that the interactions between breast cancer susceptibility loci and reproductive factors are not strong contributors to breast cancer risk.

Detecting gene-by-smoking interactions in a genome-wide association study of early-onset coronary heart disease using random forests.

BACKGROUND: Genome-wide association studies are often limited in their ability to attain their full potential due to the sheer volume of information created. We sought to use the random forest algorithm to identify single-nucleotide polymorphisms (SNPs) that may be involved in gene-by-smoking interactions related to the early-onset of coronary heart disease. METHODS: Using data from the Framingham Heart Study, our analysis used a case-only design in which the outcome of interest was age of onset of early coronary heart disease. RESULTS: Smoking status was dichotomized as ever versus never. The single SNP with the highest importance score assigned by random forests was rs2011345. This SNP was not associated with age alone in the control subjects. Using generalized estimating equations to adjust for sex and account for familial correlation, there was evidence of an interaction between rs2011345 and smoking status. CONCLUSION: The results of this analysis suggest that random forests may be a useful tool for identifying SNPs taking part in gene-by-environment interactions in genome-wide association studies.