The role of the polyamine catabolic enzymes SSAT and SMO in the synergistic effects of standard chemotherapeutic agents with a polyamine analogue in human breast cancer cell lines.

INTRODUCTION: Polyamine analogues have demonstrated significant activity against human breast cancer cell lines as single agents as well as in combination with other cytotoxic drugs. This study evaluates the ability of a polyamine analogue N (1),N (11)-bis(ethyl)norspermine (BENSpm) to synergize with six standard chemotherapeutic agents, 5-fluorouracil (FU), fluorodeoxyuridine, cis-diaminechloroplatinum(II) (C-DDP), paclitaxel, docetaxel, and vinorelbine. MATERIALS AND METHODS: Four human breast cancer cell lines (MDA-MB-231, MCF-7, Hs578t, and T47D) and one immortalized, non-tumorigenic mammary epithelial cell line (MCF-10A) were used for in vitro combination studies with BENSpm and cytotoxic drugs. Xenograft mice models generated with MDA-MB-231 cells were used for in vivo studies with BENSpm and paclitaxel. RESULTS AND CONCLUSION: BENSpm exhibited synergistic inhibitory effect on cell proliferation in combination with 5-FU or paclitaxel in human breast cancer cell lines (MDA-MB-231 and MCF-7) and was either antagonistic or less effective in the non-tumorigenic MCF-10A cell line. Synergism was highest with 120 h concomitant treatment or pre-treatment with BENSpm for 24 h followed by concomitant treatment for 96 additional hours. Since the cytotoxic effects of many polyamine analogues and cytotoxic agents are believed to act, in part, through induction of the polyamine catabolic enzymes SSAT and SMO, the role of these enzymes on synergistic response was evaluated in MDA-MB-231 and MCF-7 treated with BENSpm and 5-FU or paclitaxel. Combination treatments of BENSpm with 5-FU or paclitaxel resulted in induction of SSAT mRNA and activity in both cell lines compared to either drug alone, while SMO mRNA and activity were increased only in MDA-MB-231 cells. Induction was greater with BENSpm/paclitaxel combination than BENSpm/5-FU. Further, RNAi studies demonstrated that both SSAT and SMO play a significant role in the response of MDA-MB-231 cells to treatment with BENSpm and 5-FU or paclitaxel. In MCF-7 cells, only SSAT appears to be involved in the response to these treatments. In an effort to translate combination studies from in vitro to in vivo, and to form a basis for clinical setting, the in vivo therapeutic efficacy of BENSpm alone and in combination with paclitaxel on tumor regression was evaluated in xenograft mice models generated with MDA-MB-231 cells. Intraperitoneal exposure to BENSpm or taxol singly and in combination for 4 weeks resulted in significant inhibition in tumor growth. These findings help elucidate the mechanisms involved in synergistic drug response and support combinations of polyamine analogues with chemotherapeutic agents which could potentially be used in the treatment of breast cancer.

Sulforaphane induces cell type-specific apoptosis in human breast cancer cell lines.

Sulforaphane, an isothiocyanate found in cruciferous vegetables, has been shown to induce phase 2 detoxication enzymes and inhibit the growth of chemically induced mammary tumors in rats, although the exact mechanisms of action of sulforaphane are not understood. In this study, we evaluated the effects of sulforaphane on cell growth and death in several human breast cancer cell lines and examined the hypothesis that sulforaphane acts as a histone deacetylase (HDAC) inhibitor in these cell lines. Sulforaphane treatment inhibited cell growth, induced a G(2)-M cell cycle block, increased expression of cyclin B1, and induced oligonucleosomal DNA fragmentation in the four human breast cancer cell lines examined, MDA-MB-231, MDA-MB-468, MCF-7, and T47D cells. Activation of apoptosis by sulforaphane in MDA-MB-231 cells seemed to be initiated through induction of Fas ligand, which resulted in activation of caspase-8, caspase-3, and poly(ADP-ribose) polymerase, whereas apoptosis in the other breast cancer cell lines was initiated by decreased Bcl-2 expression, release of cytochrome c into the cytosol, activation of caspase-9 and caspase-3, but not caspase-8, and poly(ADP-ribose) polymerase cleavage. Sulforaphane inhibited HDAC activity and decreased the expression of estrogen receptor-alpha, epidermal growth factor receptor, and human epidermal growth factor receptor-2 in each cell line, although no change in the acetylation of H3 or H4 was seen. These data suggest that sulforaphane inhibits cell growth, activates apoptosis, inhibits HDAC activity, and decreases the expression of key proteins involved in breast cancer proliferation in human breast cancer cells. These results support testing sulforaphane in vivo and warrant future studies examining the clinical potential of sulforaphane in human breast cancer.