Sirolimus, and its antiproliferative capacity, was studied in vivo in three different syngenic rat tumours in the liver. Sirolimus is an inhibitor of the cytosolic mTOR-kinase, associated with the phosphoinositide-3-kinase/Akt pathway. After one week of daily sirolimus treatment, initiated on the day of tumour-cell inoculation, a dose-response relationship was shown at doses between 0.01 mg/kg/day and 1 mg/kg/day, decreasing tumour weight from 0.5+/-0.1 g in control rats (n=9) to 0.09+/-0.04 g for sirolimus 1 mg/kg (n=9). Treating established liver adenocarcinoma (n=15), sirolimus halved the tumour weight (1.4+/-0.2 g vs 0.7+/-0.1 g, p=0.005). Trough concentration in blood was 6.4+/-0.2 ng/ml after five days of daily treatment with 1 mg/kg sirolimus intraperitoneally. At this dose, there was no decrease in food consumption or rat weight, but decrease in weight of spleen, and increase in weight of liver (p<0.01). The three tumours studied, an nitrosoguanidin-induced adenocarcinoma, a Leydig cell sarcoma and a hepatoma, all responded, establishing sirolimus as a promising anticancer drug.