The hypothesis that dietary fat acts as a promotional agent for the development of breast cancer by influencing sex hormone levels was tested in a dietary intervention study. Thirty-three women in good health were randomly allocated to commence either a standard diet (deriving 40% of their energy from fat) or a low-fat diet (deriving 20% of their energy from fat). After 2 months, the women were crossed over to the alternative diet for another 2 months. Serum hormone and lipid levels were measured in the middle and at the end of each dietary period. In premenopausal women, the low-fat diet appeared to decrease levels of both non-protein-bound estradiol (1.48 down to 1.27%; P = .07) and non-protein-bound testosterone (1.06 down to 0.86%; P = .11). Cholesterol levels were lowered by the low-fat diet and were significantly associated with estradiol, testosterone, and dehydroepiandrosterone. High-density lipoprotein (HDL) cholesterol was associated with estradiol and prolactin. For the postmenopausal women, the low-fat diet lowered cholesterol and HDL cholesterol levels, but there were not the same associations with the hormones. These findings add weight to the concept that attention to diet may be a means of reducing the incidence of breast cancer in our community.