Digitoxin activates EGR1 and synergizes with paclitaxel on human breast cancer cells
Linda Saxe Einbond1, Hsan-Au Wu, Tao Su
1Columbia University College of Physicians and Surgeons, New York, NY, USA.
Background:
Numerous studies have suggested that digitalis derivatives promise to be superior to existing adjuvant therapy for breast cancer as to effects and side-effects. In the present study, we have used gene expression analysis to determine the molecular action of digitoxin on breast cancer cells and assessed digitoxin's ability to synergize with the chemotherapy agent paclitaxel with respect to inhibition of cell proliferation
Materials And Methods:
We treated (Her2 overexpressing, ER low) MDA-MB-453 human breast cancer cells with digitoxin at four doses {20 ng/ml (26 nM) to 1 μg/ml} and collected RNA at 6 h and 24 h for gene expression analysis. To examine the effects on ER positive cells, we treated MCF7 cells with digitoxin at 1 μg/ml and collected RNA for RT-PCR analysis. In addition, we assayed the growth inhibitory effect of low doses of digitoxin combined with paclitaxel and determined combination index values.
Results:
To reveal primary effects, we examined digitoxin's effect 6 h post-treatment with the highest dose, 1μg/ml, and found upregulation of the stress response genes EGR-1 and NAB2, lipid biosynthetic genes and the tumor suppressor gene p21, and downregulation of the mitotic cell cycle gene CDC16 and the replication gene PolR3B. RT-PCR analysis validated effects on stress response, apoptotic and cell cycle genes on MDA-MB-453 and MCF7 cells. Western blot analysis confirmed induction of EGR1 protein at 1 h and ATF3 at 24 h. Paclitaxel, as well as digitoxin, inhibited the in vitro activity of the purified Na(+)-K(+)-ATPase; digitoxin enhanced the growth inhibitory effects of paclitaxel on Her2 overexpressing breast cancer cells.
Conclusions:
Our studies show the potential of digitoxin to prevent and treat breast cancer and indicate that the combination of digitoxin and paclitaxel is a promising treatment for ER negative breast cancer. These findings are the first to alert physicians to the possible dangers to patients who take a combination of digitoxin and paclitaxel. The potential dangers ensuing when paclitaxel and digitoxin are combined are dependent on the dose of digitoxin.
Insights
Digitoxin shows promise for breast cancer treatment, particularly when combined with paclitaxel for ER-negative types. Physicians should be aware of potential dose-dependent dangers when using this combination therapy.
Area of Science:
- Oncology
- Pharmacology
- Molecular Biology
Background:
- Digitalis derivatives, including digitoxin, are investigated as potential adjuvant therapies for breast cancer.
- Existing treatments have limitations, prompting research into novel agents like digitoxin.
- Gene expression analysis is employed to understand digitoxin's molecular mechanisms in breast cancer cells.
Purpose of the Study:
- To determine the molecular action of digitoxin on breast cancer cells using gene expression analysis.
- To assess the synergistic effect of digitoxin and paclitaxel in inhibiting breast cancer cell proliferation.
- To evaluate digitoxin's impact on both Her2-overexpressing and estrogen receptor (ER)-positive breast cancer cells.
Main Methods:
- Gene expression analysis (RNA sequencing, RT-PCR) of breast cancer cells (MDA-MB-453, MCF7) treated with digitoxin.
- Western blot analysis to confirm protein expression changes.
- Assay of growth inhibition and combination index determination for digitoxin and paclitaxel.
Main Results:
- Digitoxin upregulated stress response genes (EGR-1, NAB2), lipid biosynthetic genes, and p21, while downregulating cell cycle genes (CDC16, PolR3B).
- Digitoxin enhanced the growth inhibitory effects of paclitaxel on Her2-overexpressing breast cancer cells.
- Both agents inhibited purified Na(+)-K(+)-ATPase activity; digitoxin potentiated paclitaxel's effects.
Conclusions:
- Digitoxin demonstrates potential for breast cancer prevention and treatment.
- The combination of digitoxin and paclitaxel is a promising strategy for ER-negative breast cancer.
- Physicians must be alerted to potential dose-dependent risks associated with combining digitoxin and paclitaxel.
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