2-Methoxyestradiol inhibits hepatocellular carcinoma cell growth by inhibiting Cdc25 and inducing cell cycle arrest and apoptosis

Abstract

2-Methoxyestradiol (2-ME) is a physiological metabolite of estrogen, which can inhibit growth of many types of tumor cells, including hepatocellular carcinoma, both in vitro and in vivo. The exact mechanisms of its action are still unclear. We have studied the mechanisms of growth inhibition of several of human and rat hepatoma and normal liver cells by 2-ME. Human (Hep3B, HepG2, PLC/PRF5) and rat (McA-RH7777, JM-1) hepatoma and normal rat (CRL-1439) and human (CRL-11233) liver cell lines were cultured in vitro, in presence of 2-ME, and its IC50s were determined. Cell cycle arrest, Cdc25 phosphatase inhibition and apoptosis induction were studied. Finally, the effect of 2-ME on the growth of JM-1 rat hepatoma cells in rat liver was determined in vivo. The IC50 range for growth inhibition of hepatoma cells was found to be between 0.5 and 3 microM. In contrast, normal rat hepatocytes and liver cell lines were resistant to 2-ME up to 20 microM. JM-1 cells were arrested in the G2/M phase of the cell cycle. Cdc25A and Cdc25B, cell cycle controlling phosphatases, activities were inhibited in vitro and 2-ME was found to likely bind to their catalytic site cysteines. As a consequence, their cellular substrates Cdk1 and Cdk2 were tyrosine phosphorylated. Caspase-3 was cleaved suggesting apoptotic cell death. Moreover, growth of JM-1 tumors, which were transplanted into rat liver, was also inhibited by treatment with 2-ME in vivo. 2-Methoxyestradiol is a selective, potent and relatively non-toxic hepatoma growth inhibitor both in vitro and in vivo. Cell cycle arrest of hepatoma cells was likely mediated by binding and inactivation of the Cdc25 phosphatases and induction of apoptosis.