To our knowledge, this is the first dense GWAS of longevity and aging traits in a community-based sample of adults from two generations of the same families. Over 1300 men and women have detailed longevity and aging-related phenotypes and 100K SNP genotyping results available on the web. This resource has the potential to detect novel susceptibility genes for human longevity and aging and to examine the relevance of promising candidate gene associations reported in animal models to human aging. We describe several strategies to prioritize SNP associations in this unique resource to enhance the discovery of various genes and pathways that contribute to the control of human longevity. Furthermore, FHS investigators are part of the NIA sponsored Longevity Consortium http://www.longevityconsortium.org
which offers the opportunity of collaboration with other investigators to replicate important findings in additional cohorts.
In our untargeted approach of ranking SNP associations by the strength of the p-value, 2 intronic FOXO1a
SNPs were associated with age at death. One of these SNPs (rs4943794) also was associated with biologic age by OSS in our a priori evaluation of select candidate genes. FOXO (forkhead box group O) transcription factors are targets of insulin-like signaling and are involved in a diverse set of physiological functions including DNA repair and resistance to oxidative stress [33
]. Further, FOXO plays a role in lifespan extension in C. elegans
]. Studies of this gene in humans are limited; two case-control studies have not identified an association between FOXO1a
and longevity [36
]. However, the prospective population-based Leiden 85-plus Study found that FOXO1a
was associated with increased mortality attributable to diabetes related deaths in participants aged 85 years and older [38
]. The Leiden 85-plus Study also reported that genetic variation causing a reduction in insulin/IGF-1 signaling resulted in improved old age survival among women [20
]. However, that report examined other genes in the insulin/insulin-like signaling pathway and did not specifically examine FOXO1a
. Finally, the untargeted approach to SNP selection also identified a SNP near FOXO3a
associated with age at natural menopause. This gene has been implicated in oocyte death, depletion of functioning ovarian follicles, and infertility in mice [39
] and thus represents a plausible candidate gene for menopause. Most positive common gene variant-disease association studies have failed replication [41
] including reports on exceptional longevity. Haplotype-based fine mapping of the region on chromosome 4 linked to human longevity initially suggested the MTP
gene, a gene important in lipoprotein synthesis, was associated with longevity [21
]. However, this association failed replication in a French cohort of long-lived individuals and subsequent case-control studies of nonagenarians [22
]. Beekman, et al
] found neither linkage to chromosome 4 nor association with the MTP gene and longevity among nonagenarians in the Leiden Longevity Study. Meta-analyses implicated admixture of the control sample in the original report as an explanation for the presumed false-positive association. Thus, our findings are hypothesis generating and their importance can not be determined without evidence of consistent replication in other populations.
We examined pleiotropic effects by identifying SNP associations across two pairs of related traits. One SNP near PON1
emerged as associated with both age at death and morbidity-free survival. Surprisingly, there were relatively few SNPs associated with both traits; prior work had suggested that longevity per se and healthy aging may share common genetic pathways [11
]. However, morbidity-free survival was measured at age 65 years, it is possible that as our participants age morbidity-free survival defined at age 75 or 85 years will share additional SNP associations with our longevity trait, age at death. A SNP near SOX5
, a gene potentially related to musculoskeletal function was associated with both biologic age by OSS and walking speed.
Our strategy of selecting SNPs in candidate genes and regions previously reported to be associated with longevity yielded interesting findings. For age at death, we identified SNPs in or near several genes including KL, LEPR, PON1, SOD2
, and WRN
. Defects in the WRN
gene are the cause of Werner Syndrome, an autosomal recessive disorder characterized by premature aging. A longitudinal study of ageing Danish twins recently reported a possible association between a successful aging trait and 3 SNPs in the WRN
]. We were unable to determine if our SNP (rs2725369) was in LD with the SNPs in the prior report because the SNPs were not included in HapMap. Mutations in the KL
(Klotho) gene in the mouse lead to a syndrome resembling human aging [45
]. There has been one report linking a functional variant of the KL
gene to human longevity [15
]. Thus, results from this GWAS may direct resources to the most relevant candidate genes and pathways for further investigation in humans.
Several important limitations merit comment. First, we acknowledge that there may be a survival bias as participants in this sample had to survive to provide DNA (first systematic DNA collection began 1995) and hence are likely healthier than the full FHS sample. To ameliorate this issue, we adjusted for covariates using the full Framingham sample, and used the residual traits for the subset of individuals genotyped using the 100K Affymetrix GeneChip to test for association with the SNPs using linear regression models. Residual traits from Cox and logistic models typically are not ideally distributed for linear regression models, but our adjustment method using the full sample precludes the testing of SNP associations with age at death and morbidity-free survival using Cox and logistic models. Second, the 100K Affymetrix GeneChip provides limited coverage of the genome; many of our a priori candidate genes did not have any SNP coverage on the chip. For example, several genes that have been studied in model organisms or even in humans such as ACE, Lamin A, SIRT2 and SIRT3, had no SNPs within 60 kb of the gene on the 100K Affymetrix GeneChip. However, genotyping is near-complete for the NHLBI funded 550 K genome-wide scan on all FHS participants. This will enable deeper exploration of our initial 100K SNP associations in a larger sample with denser coverage of the genome. Third, in this analysis we did not examine epistasis or gene-environment interactions which may modify the associations in this study. Importantly, this study is hypothesis generating. Our findings need to be replicated in other samples.