BACKGROUND: APOE is a known genetic contributor to cardiovascular disease, but the differential role APOE alleles play in subclinical atherosclerosis remains unclear. METHODS: The PESA (Progression of Early Subclinical Atherosclerosis) is an observational cohort study that recruited 4184 middle-aged asymptomatic individuals to be screened for cardiovascular risk and multiterritorial subclinical atherosclerosis. Participants were APOE -genotyped, and omics data were additionally evaluated. RESULTS: In the PESA study, the frequencies for APOE -ε2, -ε3, and -ε4 alleles were 0.060, 0.844, and 0.096, respectively. This study included a subcohort of 3887 participants (45.8±4.3 years of age; 62% males). As expected, APOE -ε4 carriers were at the highest risk for cardiovascular disease and had significantly greater odds of having subclinical atherosclerosis compared with ε3/ε3 carriers, which was mainly explained by their higher levels of low-density lipoprotein (LDL)-cholesterol. In turn, APOE -ε2 carriers were at the lowest risk for cardiovascular disease and had significantly lower odds of having subclinical atherosclerosis in several vascular territories (carotids: 0.62 [95% CI, 0.47–0.81]; P =0.00043; femorals: 0.60 [0.47–0.78]; P =9.96×10 ⁻⁵ ; coronaries: 0.53 [0.39–0.74]; P =0.00013; and increased PESA score: 0.58 [0.48–0.71]; P =3.16×10 ⁻⁸ ). This APOE -ε2 atheroprotective effect was mostly independent of the associated lower LDL-cholesterol levels and other cardiovascular risk factors. The protection conferred by the ε2 allele was greater with age (50–54 years: 0.49 [95% CI, 0.32–0.73]; P =0.00045), and normal (<150 mg/dL) levels of triglycerides (0.54 [0.44–0.66]; P =4.70×10 ⁻⁹ versus 0.90 [0.57–1.43]; P =0.67 if ≥150 mg/dL). Omics analysis revealed an enrichment of several canonical pathways associated with anti-inflammatory mechanisms together with the modulation of erythrocyte homeostasis, coagulation, and complement activation in ε2 carriers that might play a relevant role in the ε2’s atheroprotective effect. CONCLUSIONS: This work sheds light on the role of APOE in cardiovascular disease development with important therapeutic and prevention implications on cardiovascular health, especially in early midlife. REGISTRATION: URL: https://www.clinicaltrials.gov : NCT01410318.