Contents lists are available at the Mechanisms of Ageing and Development journal homepage: www.elsevier.com/locate/mechagedev. 

A genome-wide association study conducted by Almut Nebel et al. confirmed that APOE is the major gene influencing survival in long-lived individuals. The study compared autosomal SNPs in 763 long-lived individuals (LLI; mean age: 99.7 years) and 1085 controls (age: 60.2 years) from Germany. Only one association, namely that of SNP rs4420638 near the gene, achieved genome-wide significance (allele-based P = 1.8 x 10^-10). However, logistic analysis revealed that this association, which was replicated in an independent German population, is fully explicable by linkage disequilibrium with the APOE allele e4, which has been identified as the only major genetic determinant of survival into old age.

Their GWAS failed to identify any autosomal susceptibility genes. One explanation for this lack of success in their study would be limited statistical power for a polygenic phenotype with loci of small effect such as human longevity. A recent GWAS in Dutch LLI independently confirmed the APOE-longevity association, thus strengthening the conclusion that this locus is a very, if not the most, important genetic influence on longevity.

Introduction: The APOE e4 variant has been identified as a major genetic determinant for both Alzheimer's and cardiovascular diseases (Corder et al., 1993; Panza et al., 2004). It is also reportedly associated with a large adverse effect upon survival at advanced age (Schachter et al., 1994), and this association has been replicated in several populations (Christensen et al., 2006). In contrast, variation in the human forkhead box O3A gene (FOXO3A) has been found to be associated with the ability to live long, an effect corroborated by studies in Japanese, German, Italian, US-American, Jewish, Chinese and Danish populations (Anselmi et al., 2009; Flachsbart et al., 2009; Li et al., 2009; Pawlikowska et al., 2009; Soerensen et al., 2010; Willcox et al., 2008). More recently, the exonuclease 1 (EXO1) has been identified as a potential novel longevity gene (Nebel et al., 2009). All rights reserved. (2011 Elsevier Ireland Ltd.)Three genes were detected through candidate-gene approaches. Approximately 25-30% of the variation in adult lifespan is attributable to genetic factors that become more important with increasing age and exert their strongest effects in nonagenarians and centenarians (Gogele et al., 2010; Hjelmborg et al., 2006). However, only a few genetic variants have been found consistently to influence longevity. The first to be discovered was the e4 allele of the apolipoprotein E (APOE) gene, a mortality factor. A few genome-wide association studies (GWAS) for the longevity phenotype have been reported so far. In 2007, a genome-wide screening for longevity-related traits in a community-based population was published, but independent replication of its results is still pending (Lunetta et al., 2007). A meta-analysis of four prospective cohort studies of individuals reaching 90 years and older observed an association of SNP rs9664222 near the gene MINPP1, which did not reach genome-wide significance but was replicated independently (Newman et al., 2010). A case-control GWAS, including a large sample of US-American centenarians, reported 33 replicated genome-wide significant SNPs (Sebastiani et al., in press), but the validity of the results is still critically discussed (Editorial Expression of Concern; http://www.science-mag.org/content/330/6006/912.2.full). More recently, a GWAS in Dutch nonagenarians from the Leiden Longevity Study identified a SNP that tagged the deleterious effects of the APOE e4 allele (Deelen et al., in press).

TaqMan SNP Genotyping Assays and the SNPlex Genotyping System Biosystems, Foster City, USA) on an automated platform (Hampe et al., 2001). Data storage, quality control, and association analysis were performed using a customized database, R v2.7.1 (R Development Team, 2008) (http://www.R-project.org) and PLINK v1.04 (Purcell et al., 2007). Only autosomal SNPs with high-quality genotypes were included in subsequent analyses: SNPs were included if they had (a) a call rate ≥95% in cases and controls and (b) a minor allele frequency (MAF) ≥0.02 in controls or if jMAF(controls) - MAF(cases)j > 0.02 (see Fig. 2 for Manhattan-plot and Supplementary Fig. 1 for QQ-plot) and (c) P value of Hardy Weinberg equilibrium (HWE) in controls >0.01. A total of 664,472 SNPs passed quality control. To reduce the rate of false-positive results, the genotyping quality of the 1600 most significantly associated SNPs (with P values of up to 9.6 x 10^-6) was investigated by visual inspection of the corresponding cluster plots (Supplementary Figs. 2 and 3). More detailed quality control for GWAS and follow-up is described in detail in Supplementary methods.

Genotype-phenotype associations were assessed for statistical significance using a Cochran-Armitage trend (CAT) test and Fisher's exact test, with the latter applied to both an allele- (CCA) and a genotype-based (CCG) case-control comparison. Population stratification was low (genomic inflation (GC) factor (Devlin and Roeder, 1999) for stage 1: 1.064 and stage 2: 1.06). GC-adjusted P values were calculated according to Devlin and Roeder (1999). Logistic regression analysis of rs4420638 and APOE e4 status as influence variables was conducted with and without interaction using R v2.7.1 (R Development Team, 2008). To this end, genotypes were encoded by the number of the major allele of rs4420638 or the e4 allele (APOE), respectively. All tests were two-sided.