Metabolic syndrome, including low HDL cholesterol, has been associated with an increased breast cancer risk, whereas little is known of the relationship with total cholesterol. Cox proportional hazards regression models were performed to evaluate the association between self-reported total serum cholesterol, cholesterol-lowering drugs, and risk of breast cancer in 69 088 women from the French E3N cohort study. A total of 2932 cases of primary invasive breast cancer were reported during 12 years of follow-up. Compared with women with low/normal serum cholesterol (<6.6 mol/l), users of cholesterol-lowering drugs had a significantly decreased breast cancer risk [hazard ratio (HR): 0.79, 95% confidence interval (CI): 0.68, 0.93]. There was no variation in HRs according to the menopausal status. In strata defined by the hormone receptor status of the tumor, the risk reached statistical significance only for the estrogen-positive and progesterone-positive receptor subtype (HR: 0.64, 95% CI: 0.50, 0.82). A high cholesterol without cholesterol-lowering drug use was not associated with breast cancer risk (HR: 0.99, 95% CI: 0.85, 1.15) in the entire population. Our result concerning cholesterol-lowering drugs is consistent with studies showing that hypolipidemic molecules are effective in inhibiting cancer cell growth proliferation. Further studies should investigate whether these findings apply to all classes of cholesterol-lowering drugs.