The PCSK1 gene is an insulin-processing enzyme that regulates insulin biosynthesis. Mutations in this gene are associated with obesity, tumorigenesis, and metastasis. The PAX4 gene is a member of the paired box family of transcription factors that mediates differentiation of insulin-producing beta-cells in the pancreas and plays critical roles during cancer growth. The EGFR gene is a transmembrane glycoprotein that functions as a tyrosine protein kinase. Mutations in this gene have oncogenic potential in the development of non-small cell lung cancer. The LYN gene encodes a tyrosine kinase that is essential in immunoglobulin-mediated signaling, particularly in establishing B cell tolerance.

The study acknowledges that genetic variation exists across ethnic groups and reports significant association of genes for human leukocyte antigen. Supplemental Data Tables S2 and S3 present additional results for genotype combinations composed of each candidate SNP pair for two genes along with numbers of cases and controls. The study found significant epistatic interactions between PDGFC and GYS2 for males older than 90 years and between TNFSF11 and CDK10 in the group adjusted by gender. These interactions showed a dose-response relationship with increasing copies of the target allele for each SNP.genes and SNPs were nominally associated with longevity in at least one of the age/gender groups studied. However, it is important to note that the study had limitations, such as the inability to control for environmental factors due to lack of information in the control group. Additionally, the small number of centenarians may have reduced the statistical power to detect genes with weak effects, particularly in gene-gene interactions. The interpretation of findings using the over-dominant model and a control group consisting of 12-14 year old middle school students also raises concerns.

Although none of the SNPs achieved genome-wide significance, the underlying LD structure among the markers reduced the number of effectively independent tests, and thus, the genome-wide threshold of significance may be too stringent, particularly in the context of a candidate gene approach. The theory of evolution suggests that differential survival reflects the effect of deleterious genotypes of age-related diseases. However, some longevity genes (e.g., klotho associated with low HDL) buffer the deleterious effect of age-related disease genes (e.g., lipoprotein, Lp(a), a susceptibility gene for vascular disease), which may explain an observed paradoxical increase of deleterious genotypes in centenarians.

In this study, 60 genes out of 179 candidates were nominally associated with longevity in at least one age/gender group. Among eleven genes that consistently yielded statistical evidence of allelic, genotypic and haplotypic association with lifespan, only four genes remained significant after Bonferroni correction for haplotypes. The protein encoded by one of these four genes drew a 10 ml fasting venous blood sample in an EDTA tube. The gender and age composition of study subjects are shown in Supplemental Data Table S1.

For genetic variants with strong effects, relatively small sample size is sufficient to achieve statistical power to detect association. However, the rarity of centenarians makes international collaboration essential to increase statistical power for tests of gene by gene or gene by environment interaction. Continuing efforts may be necessary to find a functional variant in the associated region.

Candidate genes and SNP selection were based on searches for their functions in public databases (OMIM, Gene, etc.) and previous literature reviews. SNP markers within or near candidate genes were identified from the literature, the dbSNP database, the HapMap database, and Vector NTI software. A total of 565 informative single nucleotide polymorphism (SNP) markers with high design scores provided by Illumina, Inc. and heterozygosity above 0.05 using 300 independent healthy Korean samples were included. Information for these 565 SNPs located in 194 genes is presented in Supplemental Data Table S2.

Overall, the study provides evidence for nominally associated candidate genes and SNPs with longevity in Korean populations, but further replication studies with larger sample sizes and international collaboration are necessary to increase statistical power and validate these findings.An allele of a potent SNP confers a greater likelihood of longevity in Koreans. Previous studies (Franceschi et al., 2000) have shown that the effects of gender mediate genetic association with the probability of achieving longevity, and the observed gender difference in our findings supports, in part, the gender-specific probability of achieving longevity. Interestingly, a few candidate gene variants with no or insignificant allelic effect revealed strong evidence for the allele with a large heterozygote effect on longevity (e.g. MGP, TNFSF11, and ABCC8).

DNA preparation and SNP genotyping:

Genomic DNA samples were prepared from peripheral blood using the QIAamp DNA Blood Maxi kit following the manufacturer's instructions (Qiagen, Inc., Valencia, CA). For each sample, we determined an average concentration of DNA from three repeated measures using a SpectraMax Plus 384 spectrophotometer (Molecular Devices, Inc., Sunnyvale, CA). A 250 ng aliquot of DNA was genotyped for SNP markers using Golden-Gate chemistry on Sentrix Array Matrices (Illumina, Inc.) at the Samsung Biomedical Research Institute. We confirmed genotypes showing significant statistical evidence for association with longevity using an ABI PRISM 3100 sequencer (Applied Biosystems, Inc., Foster City, CA).