Abstract The biological origin of life expectancy remains a fundamental and unanswered scientific question with important ramifications for human health, especially as the bulk of burden of human healthcare shifts from infectious to age-related diseases. The striking variability in life-span among animals occupying similar ecological niches 1 and the numerous mutations that have been shown to increase lifespan in model organisms 2–5 point to a considerable genetic contribution. Using mammalian comparative genomics, we correlate lifespan phenotypes with relative evolutionary rates, a measure of evolutionary selective pressure 6 . Our analysis demonstrates that many genes and pathways are under increased evolutionary constraint in both Long-Lived Large-bodied mammals (3L) and mammals Exceptionally Long-Lived given their size (ELL), suggesting that these genes and pathways contribute to the maintenance of both traits. For 3L species, we find strong evolutionary constraint on multiple pathways involved in controlling carcinogenesis, including cell cycle, apoptosis, and immune pathways. These findings provide additional perspective on the well-known Peto’s Paradox that large animals with large numbers of cells do not get cancer at higher rates than smaller animals with fewer cells 7 . For the ELL phenotype, our analysis strongly implicates pathways involved in DNA repair, further supporting the importance of DNA repair processes in aging 8–12 . Moreover, these correlations with lifespan phenotypes are consistent across the entire mammalian phylogeny, suggesting that additional constraint on these pathways is a universal requirement for long lifespan.