Manganese and an isoflavone in combination treatment inhibits breast cancer cell proliferation

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Breast cancer is the most frequently diagnosed form of cancer in women and is governed by both genetic and epigenetic factors. The genetic factors account for about 1-2% of breast cancer cases whereas diet, an epigenetic factor, is one major factor that can influence breast cancer incidence in a population. Clinical & in vitro studies indicate breast cancer tissues and cell lines exhibit higher levels of reactive oxygen species (ROS) compared to normal breast tissues and cells. Consumption of diet rich in fruits, green leafy vegetables and soy-based products correlates with lesser incidence of breast cancer in certain populations. Since both biochanin-A (Bio-A, a soy isoflavone with antioxidant properties) and manganese (Mn, a trace element) are present in such diets, we hypothesized that treatment of breast cancer cells with a combination of Bio-A and Mn results in reduced breast cancer cell survival. To test our hypothesis, we carried out cell survival analysis with Bio-A and Mn alone and in combination. Treatment of SKBr3 cancer cells with combinations of Bio-A and Mn (Bio-A+Mn; 50+0.5µM→81.679±1.48, p<0.0001; 80+0.5µM→32.904±1.15, p<0.0001) led to increased inhibition of cell survival as compared to that of Bio-A and Mn alone. Increased lactate dehydrogenase released by SKBr3 cells in the medium upon combination treatment compared to Bio-A (50µM) or Mn (0.5µM) treatment alone suggests necrosis as one cell death mechanism. Western blot analysis to determine Akt/p-Akt protein expression indicates a decrease in phosphorylation/activation of pro-survival Akt pathway with combinational treatments of Bio-A (50 & 80µM) & Mn (0.5µM) after 72 hrs. Since ROS can alter cellular proliferation, we investigated effect of Bio-A and Mn on mechanisms that control intracellular level of ROS and found increased manganese superoxide dismutase (MnSOD) level upon treatment with the combination of Bio-A (50µM) & Mn (0.5µM). Elevated MnSOD could result in increased intracellular H2O2 levels and stimulate cell death pathways as indicated by increased level of LDH release by the cells. Our data suggest Bio-A- and Mn-mediated increased inhibition of cell survival may be partly due to necrosis, inhibition of anti-apoptotic Akt pathway & change in intracellular redox levels from increased expression of MnSOD. These observations suggest treatment with Bio-A combined with Mn may provide an enhanced anticancer effect. (This study was supported by FRC Grant # 681-507-01)