AbstractMammary tumorigenesis is augmented when C3H/Ou mice are fed diet ad libitum but delayed when calories are restricted by 40%. Three feeding experiments were done to evaluate the effect of ethanol on mammary tumorigenesis in isocalorically fed C3H/Ou mice: 1) ad libitum feeding of semipurified solid diet, with one group receiving 12% ethanol (15 g/kg/day) in the drinking water while controls received water alone; 2) isocaloric pair feeding of semipurified solid diet, with ethanol (4 g/kg/day) administered by gavage five times per week; and 3) isocaloric pair feeding of Lieber‐DeCarli liquid diet, with one group receiving 29% of calories as ethanol (20 g/kg/day) in the diet. Despite administration of ethanol to isocalorically fed C3H/Ou mice for 65 weeks by three different methods, mammary tumor development was not enhanced. In two of the three ethanol‐consuming groups, weight gain and mean body weight were less in the ethanol‐consuming mice than in the controls, despite equal total calorie consumption. In only one ethanol‐consuming group, where mice received ethanol as a 12% solution in the drinking water, was any difference noted in the tendency to develop mammary tumors. In this case, delay in tumorigenesis was apparent in the ethanol‐consuming animals (p = 0.03). These findings do not support the hypothesis that ethanol calories augment the risk for breast tumorigenesis among breast cancer‐prone mice consuming isocaloric diets. Instead, reductions in weight gain and body weight among ethanol‐consuming mice and an apparent reduction in mammary tumorigenesis in one of three experimental groups suggest that ethanol may decrease metabolic utilization of calories and hence contribute to lowered energy availability. This in turn could decrease tumorigenesis.