HYDROXYTYROSOL AND PIPERLONGUMINE INHIBIT BREAST CANCER CELL PROLIFERATION BY INDUCING S AND G2 DELAY

Madeline Hart , Amanda L. Kalen, Jaimee C. Eckers, Ehab H. Sarsour, Prabhat C. Goswami

HYDROXYTYROSOL AND PIPERLONGUMINE INHIBIT BREAST CANCER CELL PROLIFERATION BY INDUCING S AND G2 DELAY

Cancer cells have a lower antioxidant capacity and tolerate less oxidative stress compared to normal cells.  Phytochemicals alter the redox state of cells.  The dietary phytochemical hydroxytyrosol (2-(3,4-dihydroxyphenyl) ethanol) increases hydrogen peroxide (H2O2) in cancer cells, causing inhibition of cell proliferation.  Another dietary phytochemical, piperlongumine (1-[(2E)-3-(3,4,5-Trimethoxyphenyl)prop-2-enoyl]-5,6-dihydropyridin-2(1H)-one), inhibits the mechanism that removes H2O2.  Therefore, possibly a combination of the two phytochemicals will increase oxidative stress and therefore increase cancer cell toxicity. 

To test this hypothesis, MB231 breast cancer cells were treated with hydroxytyrosol, piperlongumine or a combination and the doubling times were calculated.  Treated cells were also stained with propidium iodide and the DNA content analyzed using flow cytometry.  MB231 cells treated with hydroxytyrosol, piperlongumine or a combination were also analyzed using Q/RT-PCR in order to analyze the expression of MnSOD and catalase. Cells were additionally analyzed using an SOD and MnSOD activity assay to determine the activity of these two proteins.

MB231 cells treated with hydroxytyrosol and piperlongumine had greater doubling times than untreated cells.  Furthermore, cells treated with both chemicals had even greater doubling times.  Cells analyzed using flow cytometry revealed that hydroxytyrosol induced an early S-phase block, and piperlongumine created a G2 block.  In cells treated with both chemicals, the effects are synergized.  A combination of hydroxytyrosol and piperlongumine decreased superoxide dismutase activity in MB231 cells and perturbed catalase and MnSOD expression. 

 As the data demonstrate, piperlongumine and hydroxytyrosol inhibit the proliferation of MB231 cancer cells, alter the internal redox environment, and disturb the cell cycle.  This can be seen from piperlongumine and hydroxytyrosol altering SOD activity in MB231 cancer cells as well as influencing MnSOD and catalase expression.  Overall, piperlongumine and hydroxytyrosol exhibit anti-proliferative properties in MB231 cancer cells and have synergistic effects when used together in treatments.  Further experiments will investigate how the two phytochemicals alter the cell cycle of MB231 cells.

We thank funding from T34 GM 08718 and NIH R01 CA 111365 for supporting this work.

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