d-δ-Tocotrienol-mediated Cell Cycle Arrest and Apoptosis in Human Melanoma Cells

Background: The rate-limiting enzyme of the mevalonate pathway, 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase, provides essential intermediates for the prenylation or dolichylation of growth-related proteins. d-delta-Tocotrienol, a post-transcriptional down-regulator of HMG CoA reductase, suppresses the proliferation of murine B16 melanoma cells. Dietary d-delta-tocotrienol suppresses the growth of implanted B16 melanomas without toxicity to host mice. Materials and Methods: The proliferation of human A2058 and A375 melanoma cells following a 72 h incubation in 96-well plates was measured by CellTiter 96 (R) Aqueous One Solution. Cell cycle distribution was determined by flow cytometry. Fluorescence microscopy following acridine orange and ethidium bromide dual staining and procaspase-3 cleavage were used to detect apoptosis. Western-blot was employed to measure protein expression. Results: d-delta-Tocotrienol induced dose-dependent suppression of cell proliferation with 50% inhibitory concentrations (1050) of 37.5 +/- 1.4 (A2058) and 22.3 +/- 1.8 (A375) mu mol/l, respectively (data are reported as mean +/- standard deviation). d-delta-Tocotrienol-mediated cell cycle arrest at the G(1) phase was accompanied by reduced expression of cyclin-dependent kinase 4. Concomitantly, d-delta-tocotrienol induced caspase-3 activation and apoptosis. The impact of d-d-tocotrienol on A2058 cell proliferation was potentiated by lovastatin (IC50=3.1 +/- 0.5 mu mol/l), a competitive inhibitor of HMG CoA reductase. Conclusion: d-delta-Tocotrienol may have potential application in melanoma chemoprevention and/or therapy.