P P P OR 95% C.I., Bonferroni adjusted P GC P value SNP call rate (LLI) SNP call rate (controls) CAT CCA CCG CCA 
0.006 0.007 0.021 
0.82 0.71 
0.94 
0.084 
0.02 
0.95 
0.98 
0.496 
0.527 
0.469 
1.06 
0.90-1-25 
1 
032-2(cid:3)10(cid:3)07 
0209-2(cid:3)10(cid:3)05 
044-2(cid:3)10(cid:3)07 
0205-4(cid:3)10(cid:3)07 
0411-1(cid:3)10(cid:3)11 
0233-7(cid:3)10(cid:3)08 
0922-2(cid:3)10(cid:3)06 
1 
0.98 
0.185 
0.621 
0382-1(cid:3)10(cid:3)07 
0280-7(cid:3)10(cid:3)12 
0910-2(cid:3)10(cid:3)06 

Bonferroni adjustment was made for 13 tests (see Supplementary methods). For details see legend to Table 2. Table 4 for SNPs included in the 5000 most significantly associated haplotypes yielded only 19 marker pairs with high-quality genotypes for both SNPs. Of these, two haplotypes including the APOE locus and four redundant haplotypes from a region of strong LD on chromosome 6 were not considered for follow-up. Thus, 13 haplotype pairs were chosen for validation in the stage 2 samples. However, none of them showed aConsidered, 51,912 (7.7%) showed a nominally significant association with longevity in a score test assuming an additive risk model for haplotypes (P < 0.05). Visual inspection of the scatter plots scores. 

Discussion: Our study confirmed the known association between APOE and survival at advanced age but failed to identify any additional susceptibility loci for longevity. This result is in marked contrast to other complex phenotypes for which GWAS have successfully uncovered multiple genetic risk factors at unprecedented speed and with high levels of reproducibility (Esparza-Gordillo et al., 2009; Franke et al., 2008; Hofmann et al., 2008). One explanation for this lack of success in our study would be that GWAS provides only limited statistical power for a polygenic phenotype with loci of small effect such as human longevity. GWAS is more suitable for the detection of genetic variants with large effects, which presumably explain only a small fraction of the total variation (Barrett et al., 2008). It is assumed that many causal gene variants, each with a weak to moderate contribution, influence our ability to become long-lived (Christensen et al., 2006). A recent study supports this notion and concludes that the alleles with small individual effects may jointly influence lifespan so that the resulting impact can be both substantial and significant (Yashin et al., 2010).

Table 5: Association results of SNPs identified in the GWAS of Sebastiani et al. in the German GWAS dataset. Markers not genotyped on the Affymetrix 6.0 Chip were:

Chr. Position B36 ID MAF (controls) AF (LLI) P value INFO score 

1 243871287 rs10924270 0.461 0.467 0.738 0.795 
1 246927264 rs4916176 0.125 0.122 0.820 
2 5210976 rs1377638 0.132 0.123 0.380 0.979 
2 171530712 rs4668356 0.054 0.065 0.185 0.991 
2 173328711 rs6433379 0.123 0.131 0.527 0.914 
2 234432806 rs579327 0.177 0.159 0.152 
2 235521853 rs2042831 0.076 0.080 0.488 0.940 
3 14874782 rs294636 0.183 0.189 0.566 0.963 
3 97267562 rs4393926 0.341 0.330 0.543 (in LD with rs4390941, a r2=0.99) 
4 80183611 rs1455311 0.177 0.172 0.767 0.991 
5 55450713 rs415407 0.467 0.471 0.834 (in LD with rs9899404, a r2=0.99) 
5 65819465 rs10069397 0.484, [space here] [remove hyphen]   [space here]   [remove hyphen]   [space here]   [remove hyphen]   [space here]   [remove hyphen]   [space here]   [remove hyphen]   [space here]   [remove hyphen]   [space here]   [remove hyphen]    [remove comma]    [add space after comma]    [add space after semicolon] 0.055 0.993 
5 83892667 rs7717527 0.182 0.196 0.443 0.967 
6 31237686 rs2073724 0.086 0.085 0.861 
6 33198814 rs4713607 0.495 0.519 0.197 1.000 
8 58468572 rs954295 0.212 0.207 0.764 0.992 
8 58940672 rs1436013 0.214 0.219 0.714 0.885 
8 79864431 rs2717536 NA NA NA 
8 110758554 rs3133926 0.215, [space here] [remove comma]   [remove space after comma]   [remove hyphen]   [space here]   [remove hyphen]   [space here]   [remove hyphen]   [space here]   [remove hyphen]   [space here]   [remove hyphen]   [space here]  One point to note is the detection of the longevity gene FOXO3A (Anselmi et al., 2009; Flachsbart et al., 2009; Li et al., 2009; Pawlikowska et al., 2009; Soerensen et al., 2010; Willcox et al., 2008) and the candidate EXO1 (Nebel et al., 2009) in our GWAS. The P values obtained were 0.007 and CCA 0.035, respectively. However, these P values were too large to qualify the corresponding SNPs for follow-up in stage 2, suggesting that our study was likely to miss most genetic factors of similarly small effect (i.e. OR   1.4). Furthermore, we could not detect male-specific effects due to the small number of men in the replication study. The German and French centenarian/LLI samples used in this study were among the largest to-date, but they may not have been powerful enough to detect small-effect variants in a longevity GWAS. For example, the German sample used in stage 1 had a power of only 3.2% to detect an association of an OR of 1.4 (minor allele frequency of 50% and applying a significance level of 7.5 x 10^8 (Supplementary Fig. 5)).

Acknowledgements
We would like to acknowledge the excellent technical support provided by all laboratory staff at the Institute of Clinical Molecular Biology, the Popgen biobank, and the Biological Resource Center of the Fondation Jean Dausset-CEPH. This study was supported by the German Federal Ministry of Education and Research (BMBF) through the German National Genome Research Network (NGFN-2 and NGFN-plus), the DFG Excellence Cluster Inflammation at Interfaces, the Helmholtz Center Munich German Research Center for Environmental Health, the Munich Center of Health Sciences (MC Health) as part of LMUinnovativ, and the French Ministère de l'Enseignement Supérieur et de la Recherche.

Appendix A. Supplementary data
Supplementary data associated with this article can be found in the online version at doi:10.1016/j.mad.2011.06.008.