Several studies suggest an important role of Interleukin-27 in the development of atherosclerosis. The aim of this study was to establish whether the gene polymorphisms are associated with premature coronary artery disease and/or other cardiovascular risk factors. Four gene polymorphisms were selected and genotyped in 1162 premature coronary artery disease cases and 1107 controls. rs26528 and rs40837 alleles were significantly associated with a lower risk of premature coronary artery disease under different inheritance models (P = 0.046; P = 0.002; P = 0.007 for rs26528T; P = 0.008 and P = 0.031 for rs40837). The rs40837 allele was also associated with a lower risk of insulin resistance, in cases (P = 0.037) and controls (P = 0.008; P = 0.047; P = 0.014; P = 0.006), while the rs26528 allele was associated with a lower risk of insulin resistance only in the control group (P = 0.016; P = 0.021). Interleukin-27 plasma levels were measured in 450 controls and 450 cases, and were significantly higher in cases compared to controls (P = 0.004). However, Interleukin-27 plasma levels were not associated with polymorphisms. Luciferase assays showed that co-transfection of the rs40837 allele and miR-379-5p significantly decreased luciferase gene expression. Our study shows for the first time, that IL gene polymorphisms are associated with premature coronary artery disease and with some metabolic parameters. The rs40837 allele in presence of miR-379-5p significantly decreased luciferase gene expression.