Preclinical evaluation of 5-aza-2'-deoxycytidine, an inhibitor of DNA methylation, for therapy of breast cancer.

摘要

Breast cancer is the second most common cause of cancer death among women in the United States. Chemotherapy and hormonal therapy have proven to be effective in early breast cancers, but most women with metastatic disease that does not respond to hormonal therapy have only a limited life expectancy with the current conventional therapy. There is an urgent need to develop new approaches for therapy of advanced breast cancer.; The identification of many tumor suppressor and cancer-related genes that are silenced by aberrant methylation in breast cancer make it interesting to investigate the potential of therapy with demethylating agents, such as 5-Aza-2′-deoxycytidine (5-AZA-CdR) for this disease. Aberrant DNA methylation of promoter regions of cancer-related genes leads to silencing of their expression. Demethylation may restore the tumor suppressor functions and arrest tumorigenesis.; We investigated the in vitro antineoplastic activity of 5-AZA-CdR, an analog of 2′-deoxycytidine, on the human MDA-MB-231 breast carcinoma cell line. We found that this analog was able to inhibit cell growth and clonogenic potential of these tumor cells. The concentrations used in our assays were in the same range of the plasma levels of 5-AZA-CdR used in pilot clinical studies with responses in patients with cancer.; An interesting approach to increase the therapeutic efficacy of an anticancer drug and to avoid drug resistance, is to use a combination of agents whose mechanisms of action are different. We investigated the combination of 5-AZA-CdR with a new class of antineoplastic drugs, the histone deacetylase inhibitors (HDIs), such as trichostatin A (TSA). These latter agents maintain histone acetylation, leading to less compact, transcriptionally active chromatin. HDIs also induce terminal cellular differentiation. We investigated the antineoplastic action of combination of 5-AZA-CdR and TSA on the MDA-MB231 breast carcinoma cell line. Our results show a synergistic effect of the two drugs on the inhibition of clonogenic potential of these tumor cells.; In order to understand the mechanisms underlying this interaction, we studied the effect of 5-AZA-CdR and TSA on the expression of two tumor suppressor genes, the retinoic acid receptor β (RARβ) and the estrogen receptor (ER) gene. Both genes have been extensively reported to be silenced by aberrant methylation in breast cancer. We showed by RT-PCR that exposure of MDA-MB-231 tumor cells to 5-AZA restored expression of these two genes by demethylation of their promoter regions. TSA is able to restore expression of RARβ, but not that of ER, suggesting the existence of different mechanisms of epigenetic silencing for different genes. Combination of the two agents appeared to induce a more extensive reactivation of expression of the two genes studied.; This study supports future investigations on the use of 5-AZA-CdR in the therapy of breast cancer. The efficacy of this cytosine nucleoside analog can be increased by the combination of other agents, such as HDIs, to induce a higher expression of cancer-related genes. This new approach to breast cancer therapy may be effective for women with advanced breast cancer. In addition, this therapy has the potential to re-express the ER gene and possibly reestablish responsiveness to antiestrogen therapy with tamoxifen.