G(0)/G(1) arrest and S phase inhibition of human cancer cell lines by inositol hexaphosphate (IP6)

Background: Inositol hexaphosphate (InsP(6) or IP6) has shown a striking an ti-cancer activity in both in vivo and in vitro models. In an attempt to el ucidate the mechanism(s) underlying the anti-neoplastic potential of IP6, w e investigated its effect on cell cycle progression of MCF-7 estrogen recep tor (ER)positive and MDA-MB 231 ER-negative human breast cancer cell lines and HT-29 human colon cancer cells. Methods: The anti-proliferative effect of IP6 was evaluated using dual-parameter flow cytometric measurements of D NA content, versus the incorporation of 5-bromo-2-deoxyuridine (BrdU) to de termine cells actively synthesizing DNA. Combined analysis of the expressio n of cell cycle-related proteins, proliferation marker Ki-67 and proliferat ing cell nuclear antigen (PCNA) versus DNA content were used to determine t he amount of proliferating cells in each phase, engaged in cell cycle trans it. Results: After 3 days of treatment with 5 mM IP6, S-phase, as estimated by BrdU uptake, was significantly decreased in all three cell lines (p = 0 .002). MCF-7 and HT-29 cells accumulated in the G(0)/G(1) range of DNA cont ents (p = 0.002 and p = 0.001, respectively). MDA MB-231 cells transiently accumulated in G(0)/G(1) only after 2 days (p = 0.01). There was a signific ant decrease in the percentage of Ki-67 expression in IP6-treated cells, fr om 82.8 +/- 3.0% to 66.8 +/- 4.2% in MCF-7 (p = 0.007), from 93.4 +/- 4.6% to 71.7 +/- 3.3% in MDA-MB 231 (p = 0.004), and from 95.2 +/- 1.2% to 73.5 +/- 2.5% in HT-29 cells (p = 0.002) respectively. PCNA expression levels we re also significantly decreased by IP6 in all three cell lines (MCF-7 p = 0 .0007; MDA-MB 231 p = 0.0006; HT-29 p = 0.0001). Conclusion: These results show that IP6 controls the progression of cells through the cycle by decrea sing S- phase and arresting cells in the G(0)/G(1) phase of the cell cycle. A significant decrease in the expression of proliferation markers indicate d that IP6 disengaged cells from actively cycling. Further investigations o f cell cycle regulators may lead its to a better understanding of the mecha nism (s) of the anti-neoplastic action of IP6.