Discussion

The CHARGE Consortium collaboration allowed us to conduct a meta-analysis of GWAS for longevity in a sample of long-lived individuals and a corresponding comparison group derived from the same longitudinal community-based cohort studies. Although none of the SNP associations for longevity in the first discovery phase achieved prespecified level of genome-wide significance, a polymorphism associ- ated with the MINPP1 genes was among the strongest as- sociations observed in our sample, with effect sizes that were similar within the four cohorts. The finding related to the MINPP1 gene was strengthened after including two ad- ditional cohorts in a second discovery phase but did not reach genome-wide significance. Among the top 10 associa- tions in the initial meta-analysis, additional SNP associa- tions of potential interest in longevity include SNPs in or near LASS3, ACCN1, IL20RB, and PAPPA2. These SNPs

Figure 3. Study-specific odds ratios (ORs) and 95% confidence intervals for MINPP1 (rs9664222) longevity association. Note: AGES-Reykjavik = Age, Gene/Environment Susceptibility-Reykjavik Study; CHS = Cardiovascular Health Study; FHS = Framingham Heart Study.

studies in a joint meta-analysis, with the p value decreasing about 10-fold, from 1.61 × 10−5 to 6.77 × 10−7 and corre- sponding OR of 0.82. This SNP, rs9664222, is ~25 kb from the MINPP1 gene (Figure 4). In the CHARGE analysis, the minor allele was associated with a lower odds of survival past age 90 (OR = 0.77). The Leiden study yielded a similar effect estimate (OR = 0.76, p = .0014), whereas the Danish study showed a nonsignificant trend in the same direction

1s9664222 region

Observed (-logP)1s9664222 regionrs9664222@ P=1.608e-05GLUD1 FAM22A PAPSS2 ATADI KILLINFAM3SA 89000 89300 89600Chromosome 10 position (ha18) (kb)(QW/W9) ayes uo}eUIquiodeYy

Figure 4. Regional plot for rs9664222 near MINPP1.

the likelihood that longevity is a complex process, in that there were no variants with large effects, supporting the hypothesis that there may be many genes with small effects that contribute to longevity.

are near genes that have not previously been reported to be associated with longevity in human populations but are in- teresting because these genes are conserved in basic bio- logic pathways.

The MINPP1 gene codes multiple inositol polyphosphate phosphatases, which are compartmentalized to the endo- plasmic reticulum lumen. MINPP1-deficient mice have no obvious defects, though targeted deletion in vitro is associ- ated with slowed cellular proliferation (46). There is no evi- dence that this SNP is functional; furthermore, its distance from the gene shows that it is not in strong linkage disequi- librium with SNPs in MINPP1 (47). However, it is well known that important regulatory elements are found outside of genes. This SNP is within 50 kb of two copy number variants. The finding of an SNP near a gene regulating pro- liferation is intriguing because of the higher rate of cancer death in the comparison group.

The strengths of this study include the community-based prospective design and the long-term follow-up of these co- horts. In all cases, vital status was confirmed using death certificates and hospital records. Another strength was our ability to use controls that were equally well characterized and were drawn from within the same cohorts. The number of long-lived individuals reported here is very large relative to other studies in the literature, allowing greater ability to identify SNPs with small effects. The cohorts were rela- tively homogeneous with respect to ancestry, limited to Caucasians of European decent. Our top associations were homogeneous across cohorts. Screening for latent popula- tion substructure also supported ethnic homogeneity. Thus, the findings reported are less likely to be due to population stratification.

from http://biomedgerontology.oxfordjournals.org/ at National Sun Yat-sen University on August 22, 2014

The initial finding in the LASS3 gene region was of inter- est because of the historical association of its homologue with longevity in yeast (46). The LASS gene family contains a group of highly conserved genes that are found in all eu- karyotic species. LASS isoforms are mammalian homo- logues of the yeast longevity assurance gene 1, which encodes a protein that regulates life span (48). The strongest association was noted for rs8029244; this SNP is in the in- tronic enhancer region of the LASS3 gene. LASS3 is a mem- ber of the ceramide synthase family, which is important in sphingolipid metabolism, cell differentiation, cell cycling, and apoptosis (46). LASS3 may be involved in sphingolipid synthesis or its regulation (49).

There are important aspects of the study that need to be kept in mind when interpreting the results. The differences in causes of death in the longevous individuals versus the comparison groups are expected as death from cancer tends to occur earlier in life than death from heart disease or de- mentia. Many of the long-lived people are still alive and we do not yet know what their ultimate cause of death will be, but it is likely that cancer will be underrepresented among persons achieving longevity. Power remains a limitation. Thus, future GWAS aiming to identify variants for this phe- notype will have to consider small effect sizes and target a sample size larger than our nearly 2,000 long-lived persons. DNA collection in cohort studies is a recent enough phe- nomenon that relatively few cohort members who had DNA collected have had the opportunity to survive to age 90 years. Continuous study of these and other similarly designed cohorts will allow us to extend this study to larger numbers and to older ages.