Statins directly inhibit 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR) activity, while gamma-tocotrienol, an isoform of vitamin E, enhances the degradation and reduces cellular levels of HMGR in various tumor cell lines. Since treatment with statins or gamma-tocotrienol alone induced a dose-responsive inhibition, whereas combined treatment with subeffective doses of these agents resulted in a synergistic inhibition in +SA mammary tumor cell growth, studies were conducted to investigate the role of the HMGR pathway in mediating the antiproliferative effects of combined low dose statin and gamma-tocotrienol. Treatment with 8 microM simvastatin inhibited cell growth and isoprenylation of Rap1A and Rab6, and supplementation with 2 microM mevalonate reversed these effects. However, the growth inhibitory effects of 4 microM gamma-tocotrienol were not dependent upon suppression in mevalonate synthesis. Treatment with subeffective doses of simvastatin (0.25 microM), lovastatin (0.25 microM), mevastatin (0.25 microM), pravastatin (10 microM), or gamma-tocotrienol (2 muM) alone had no effect on protein prenylation or mitogenic signaling, whereas combined treatment with these agents resulted in a significant inhibition in +SA cell growth, and a corresponding decrease in total HMGR, Rap1A and Rab6 prenylation, and MAPK signaling, and mevalonate supplementation reversed these effects. These findings demonstrate that the synergistic antiproliferative effects of combined low dose statin and gamma-tocotrienol treatment are directly related to an inhibition in HMGR activity and subsequent suppression in mevalonate synthesis.