The AGES-Reykjavik was funded by the National Insti- tute on Aging (NIA) and was designed to examine genetic susceptibility and environmental interactions as risk factors for disease and disability in old age. Detailed phenotyping of the cardiovascular, neurocognitive, musculoskeletal, and body composition and metabolism was conducted in 5,764 men and women enrolled in 2002–2006 who were sampled from the 11,549 survivors of the AGES-Reykjavik of 30,000 men and women sampled from the 1907–1935 birth cohort (31). The CHS is a National Heart Lung and Blood Institute (NHLBI) contract-funded cohort study designed to evaluate risk factors for coronary heart disease (CHD) and stroke in older adults (32). Participants (n = 5,201) were recruited in 1989–1990, with an additional 687 minorities recruited in 1992–1993. The FHS is an NHLBI contract-funded co- hort study initiated in 1948 to study determinants of CVD and other major illnesses. The original cohort comprised 5,209 men and women aged 28–62 years at enrollment who have undergone routine biennial examinations (33,34). In 1971, 5,124 offspring of the original cohort participants and offspring spouses, aged 5–70 years, were enrolled into the Framingham Offspring Study. Offspring participants have been examined approximately every 4–8 years (35,36). In the 1990s, DNA was obtained for genetic studies from sur- viving original cohort and offspring participants. The RS was planned and designed in the early 1990s as a longitudi- nal study investigating the incidence and progression of dis- eases in the elderly participants. From 1991 to 1995, all inhabitants of Ommoord, a district of Rotterdam in the Netherlands, who were aged 55 years or older, were invited to participate in this study (38). Of 10,275 eligible individu- als, 7,983 agreed to participate (78%). The participants in the CHARGE studies are Caucasian by self-report. In each CHARGE study, population structure was assessed using principal components analysis, and outliers were removed. Any remaining within-study structure was adjusted for us- ing appropriate methods (39). The details of each participat- ing cohort study’s genotyping platform, imputation algorithm, and quality control procedures used by each study are summarized in Supplementary Table 1. Each study was approved by the respective Institutional Review Board, and all participants provided consent.

Second Discovery Stage Genotyping

Among the top 24 independent regions with the strongest associations for longevity in the four-study meta-analysis (p < 10−4), we selected the 22 SNPs that had been tested in all four CHARGE cohorts in two additional Caucasian cohorts: the Leiden Longevity Study cohort and the Danish 1905 cohort. We excluded the two SNPs that could not be geno- typed or imputed in all four CHARGE cohorts. Of the 22 SNPs selected for genotyping, 2 could not be genotyped and 4 did not pass quality control procedures; thus, 16 SNPs were analyzed in the second stage.

In the “Leiden Longevity Study” (7,40), a total of 950 long-lived proband siblings (mean age 94 years, range 89– 104 years), 1,750 offspring (mean age 61 years, range 39– 81 years), and 758 partners of offspring (mean age 60 years, range 36–79 years) were included. The additional genotyp- ing of selected SNPs was undertaken in all 950 long-lived probands, and these were compared with the 744 partners of their offspring and an additional 680 blood bank donors

(60% men, mean age 31 years, range 18–40 years). All long-lived individuals and the comparison groups were from the Leiden area in the Netherlands and of European ancestry.

were meta-analyzed. The study-specific inflation factors (l ) were computed using the set of chi-square statistics GC used for the meta-analysis for each study. The inflation fac- tor is computed as the median of all chi-square statistics divided by the expected median of the statistics (approxi- mately 0.456) for a chi-square distribution with 1 df. We calculated a meta-analysis odds ratio (OR) for each SNP using a fixed-effects model that combined logistic regres- sion parameters and standard errors across the studies using inverse variance weights. The meta-analysis OR represents the increase in log-odds of surviving to age 90 years or older versus dying between ages 55 and 80 years for each addi- tional copy of the minor allele of the SNP. SNP associations were considered to be significant on a genome-wide level at p < 5 × 10−8. The 16 SNPs in the second discovery phase effort were analyzed in the two study samples using an ad- ditive model. The results were added to the previous meta- analysis using a fixed-effects model as described earlier. Finally, using the top 24 results, we conducted a pathway analysis with the Database for Annotation, Visualization and Integrated Discovery (http://david.abcc.ncifcrf.gov/).

Participants in the “Danish 1905 Cohort Survey” are from the Danish 1905 birth cohort ascertained in 1998 when they were aged 92–93 years (41). Of the 3,600 participants alive from that cohort, 2,262 participants enrolled in the study. Participants underwent a home-based interview on health and lifestyle parameters, physical and cognitive tests, and collection of biologic material. The current genetic study comprises a total of 1,644 participants from this survey, mean age 93 years (range 92–93 years), 28% men. A com- parison group included 2,007 Caucasians who were twins (one twin per pair) collected from all over Denmark, with a mean age of 57 years (range 46–68 years), 45% men.

Second Discovery Stage Genotyping Methods