Molecular correlates of the action of bis(ethyl)polyamines in breast cancer cell growth inhibition and apoptosis

Abstract

Polyamines are known to be involved in cell growth regulation in breast cancer. To evaluate the efficacy of bis(ethyl)polyamine analogs for breast cancer therapy and to understand their mechanism of action we measured the effects of a series of polyamine analogs on cell growth, activities of enzymes involved in polyamine metabolism, intracellular polyamine levels, and the uptake of putrescine and spermidine using MCF-7 breast cancer cells. The IC50values for cell growth inhibition of three of the compounds, N1,N12-bis(ethyl)spermine, N1,N11- bis(ethyl)norspermine, and N1,N14-bis(ethyl)homospermine, were in the range of 1-2 µM. Another group of three compounds showed antiproliferative activity at about 5 µM level. These compounds are also capable of suppressing colony formation in soft agar assay and inducing apoptosis of MCF-7 cells. The highly effective growth inhibitory agents altered the activity of polyamine biosynthetic and catabolic enzymes and down-regulated the transport of natural polyamines, although each compound produced a unique pattern of alterations in these parameters. HPLC analysis showed that cellular uptake of bis(ethyl)polyamines was highest for bis(ethyl)spermine. We also analyzed polyamine analog conformations and their binding to DNA minor or major grooves by molecular modelling and molecular dynamics simulations. Results of these analyses indicate that tetramine analogs fit well in the minor groove of DNA whereas, larger compounds extend out of the minor groove. Although major groove binding was also possible for the short tetramine analogs, this interaction led to a predominantly bent conformation. Our studies show growth inhibitory activities of several potentially important analogs on breast cancer cells and indicate that multiple sites are involved in the mechanism of action of these analogs. While the activity of an analog may depend on the sum of these different effects, molecular modelling studies indicate a correlation between antiproliferative activity and stable interactions of the analogs with major or minor grooves of DNA.Key words: polyamine analogs, breast cancer cells, apoptosis, molecular modelling.

Regulation of estrogenic and nuclear factor κB functions by polyamines and their role in polyamine analog-induced apoptosis of breast cancer cells

“…Previous studies by us and other investigators have shown a delayed apoptotic response by polyamine analogs in several cancer cell lines (Faaland et al, 2000;Kramer et al, 1997). Therefore, we performed experiments after 72 and 96 h of treatment with polyamine analogs.…”

“…Thus, these polyamine analogs were active in inhibiting both ERE-and NRE-promoters, with some structure-dependent di erences in their e cacy. Previous studies from our laboratory and others have shown that these polyamine analogs inhibit cell growth and induce apoptosis over the course of 2 ± 4 days (Davidson et al, 1993;Faaland et al, 2000;Hu and Pegg, 1997). Thus inhibition of transactivation observed at 24 h preceded e ects on cell growth or cell death.…”

“…Thus, the increased association of p300/CBP with ERa and NF-kB in the presence of spermine may help transcriptional activation of multiple genes. Polyamine analogs, by inhibiting these protein-protein associations and decreasing cellular polyamine levels (Faaland et al, 2000), may disrupt the formation of the transcription initiation complex.…”

“…Polyamine levels are higher in malignant cells and tumors compared to normal cells and tissues (Canizares et al, 1999;Leveque et al, 1999). Synthetic polyamine analogs, such as bis(ethyl)polyamines, have been shown to disrupt the natural polyamine pathway and lead to cell/tumor growth inhibition and apoptosis (Porter and Bergeron, 1988;Faaland et al, 2000;Marton and Pegg, 1995;McCloskey et al, 1996). In certain cell types, polyamine analogs cause a superinduction of spermidine/spermine N 1 -acetyltransferase (SSAT), a polyamine catabolizing enzyme Casero and Pegg, 1993;Ha et al, 1997).…”

“…In certain cell types, polyamine analogs cause a superinduction of spermidine/spermine N 1 -acetyltransferase (SSAT), a polyamine catabolizing enzyme Casero and Pegg, 1993;Ha et al, 1997). However, SSAT was not superinduced in MCF-7 breast cancer cells (Davidson et al, 1993;Faaland et al, 2000). Thus, the molecular mechanism(s) of the action of both natural polyamines and their synthetic analogs in cell growth and apoptosis remains to be elucidated.…”