Medulloblastoma is the most common malignant cancer of the central nervous system in children. AKT kinases are part of a survival pathway that has been found to be significantly elevated in medulloblastoma. This pathway is a point of convergence for many growth factors and controls cellular processes that are critical for tumor cell survival and proliferation. The alkyl-phospholipid perifosine [octadecyl-(1,1-dimethyl-4-piperidylio) phosphate] is a small molecule inhibitor in clinical trials in peripheral cancers which acts as a competitive inhibitor of AKT kinases. Medulloblastoma cell cultures were used to study the effects of perifosine response in preclinical studies in vitro. Perifosine treatment led to the rapid induction of cell death in medulloblastoma cell lines, with pronounced suppression of phosphorylated AKT in a time-dependent and concentration-dependent manner. LD50 concentrations were established using viability assays for perifosine, cisplatin, and etoposide. LD50 treatment of medulloblastoma cells with perifosine led to the cleavage of caspase 9, caspase 7, caspase 3, and poly-ADP ribosylation protein, although caspase 8 was not detectable. Combination single-dose treatment regimens of perifosine with sublethal doses of etoposide or irradiation showed a greater than additive effect in medulloblastoma cells. Lower perifosine concentrations induced cell cycle arrest at the G1 and G2 cell cycle checkpoints, accompanied by increased expression of the cell cycle inhibitor p21cip1/waf1. Treatment with p21 small interfering RNA prevented perifosine-induced cell cycle arrest. These findings indicate that perifosine, either alone or in combination with other chemotherapeutic drugs, might be an effective therapeutic agent for the treatment of medulloblastoma.