Lanostanoid triterpenes isolated from Ganoderma amboinense were found to inhibit the growth of numerous cancer cell lines, and some of them inhibited the activities of topoisomerases I and IIalpha in vitro. Among the bioactive isolates, one of the most potent triterpene was identified to be 3 alpha-hydroxy-15 alpha-acetoxy-lanosta-7,9(11),24-trien-26-oic acid, ganoderic acid X (GAX). Treatment of human hepatoma HuH-7 cells with GAX caused immediate inhibition of DNA synthesis as well as activation of ERK and JNK mitogen-activated protein kinases, and cell apoptosis. Molecular events of apoptosis including degradation of chromosomal DNA, decrease in the level of Bcl-xL, the disruption of mitochondrial membrane, cytosolic release of cytochrome c and activation of caspase-3 were elucidated. The ability of GAX to inhibit topoisomerases and to sensitize the cancer cells toward apoptosis fulfills the feature of a potential anticancer drug.