Samples

Data from three studies were analyzed. Two are large cohort studies of older adults: SOF and CHS. The third study is a case– control study (AJC).

Study of Osteoporotic Fractures (SOF)

A total of 9704 ambulatory community-dwelling Caucasian women age 65 and older were recruited from September 1986 through October 1988 from four areas in the United States: Baltimore, MD: Monongahela Valley, PA; Minneapolis, MN; and Portland, OR (Cummings et al., 1990). The study protocol was approved by the appropriate institutional review committees and written informed consent was obtained from all par- ticipants. Self-reported ethnicity was recorded as Northern, Southern, or Central European, or mixed/other. The cohort has been followed for over 20 years with age and cause of death recorded for participants who have died. Eight hundred and ninety-six SOF participants were selected for this study. These included 293 long-lived cases (lifespan ‡ 92 years, mean ± SD = 95.3 ± 2.2 years) of whom approximately 56% were still living at the last study visit. We selected 603 controls who were the youngest to die in this cohort (lifespan £ 79 year, mean = 75.7 ± 2.6 year); however, the age of death in the con- trols is not substantially different compared to the general popu- lation, as all women had to be relatively healthy to enter the cohort (Cummings et al., 1990). Cause of death was adjudi- cated by an experienced internist based on review of death

Ashkenazi Jewish Centenarians (AJC)

Subjects were of self-reported Ashkenazi Jewish ancestry (Barzi- lai et al., 2003; Atzmon et al., 2006; Suh et al., 2008). Informed written consent was obtained in accordance with the policy of the Committee on Clinical Investigations of the Albert Einstein College of Medicine. The 383 cases (74.8% female) were required to be living independently at 95 years of age as a reflection of good health (mean age 97.7 year, range 95– 108 year). The 363 controls (mean age 79.5 years, age range 43–94 year, 57% female), all living at time of recruitment, were

Table 5 Association of genotype with cause of death in SOF and CHS

Cause of death Cohort Significance AKT1 rs3803304 FOXO3A rs1935949 FOXO3A rs4946935 Atherosclerosis SOF P 0.069 0.013 0.017 OR (95%CI) 1.33 (0.98–1.81) 0.68 (0.51–0.92) 0.69 (0.51–0.94) CHS P 0.040 0.49 0.73 OR (95%CI) 1.38 (1.02–1.89) 1.10 (0.84–1.45) 1.05 (0.79–1.40) Meta-analysis P 0.0062 0.25 0.17 OR (95%CI) 1.36 (1.09–1.69) 0.89 (0.73–1.09) 0.87 (0.70–1.07) Cancer SOF P 0.087 0.045 0.12 OR (95%CI) 1.28 (0.97–1.67) 0.76 (0.58–0.99) 0.81 (0.62–1.06) CHS P 0.37 0.38 0.16 OR (95%CI) 1.14 (0.85–1.53) 0.89 (0.70–1.15) 0.83 (0.64–1.08) Meta-analysis P 0.064 0.044 0.037 OR (95%CI) 1.21 (0.99–1.48) 0.83 (0.69–0.99) 0.82 (0.68–0.99) Other SOF P 0.10 0.32 0.48 OR (95%CI) 1.30 (0.95–1.78) 0.86 (0.64–1.16) 0.90 (0.66–1.21) CHS P 0.036 0.035 0.048 OR (95%CI) 1.34 (1.02–1.75) 0.76 (0.58–0.98) 0.76 (0.58–1.00) Meta-analysis P 0.0079 0.025 0.051 OR (95%CI) 1.32 (1.08–1.62) 0.80 (0.66–0.97) 0.82 (0.67–1.00)

For this analysis, the case group comprises participants who died of specific causes and the control group comprises long-lived participants. Results for an additive model are shown. Analyses in SOF are adjusted for self-reported ethnicity; analyses in CHS are adjusted for sex. Nominally significant associations (P < 0.05) are highlighted in bold.

of self-reported Ashkenazi Jewish descent without a family history of unusual longevity (parents’ age < 85 year). Partici- pants’ ages were defined by birth certificates or passports. Four percentage of controls and 76% of centenarians have died since recruitment.

using http://www.autoprimer.com, primer3 (http://frodo.wi. mit.edu/cgi-bin/primer3/primer3_www.cgi), and as previously described (Hsu & Kwok, 2003). Assay conditions are available upon request. Genotyping in AJC was performed at the Albert Einstein College of Medicine using the Sequenom platform and pyrosequencing.

Samples were genotyped by investigators blinded to pheno- type. All data were scored by at least two investigators. DNA samples were genotyped on 384-well plates with negative and positive controls (duplicates) included in each plate. There were 19 duplicates among the SOF and 267 among the CHS samples. Plates with fewer than 90% successful genotypes or with dupli- cate discrepancies were repeated. Plates where duplicate dis- crepancies persisted after re-genotyping were excluded. Overall genotyping accuracy was > 99.5%.

Gene and variant selection