Novel herbal flavonoids promote apoptosis but differentially induce cell cycle arrest in human colon cancer cell

Formononetin is a novel herbal isoflavonoid isolated from Astragalus membranaceus, a medicinal plant that possesses antitumorigenic property. We attempted to compare the anticarcinogenic mechanism of formononetin with that of the known proapoptotic flavonoid isoliquiritigenin (ISL) in human cancer cells. We first evaluated the effects of formononetin and ISL on HCT 116 colon cancer cell viability. Immunofluorescence staining was then performed to observe the morphological changes of cancer cells undergoing apoptosis, which had been substantiated using Annexin V-FITC/propidium iodide staining. Western immunoblotting and flow cytometry were also employed to study parameters associated with apoptosis and cell proliferation. Our data show that formononetin and ISL both inhibited the growth of colon cancer cells and promoted apoptosis. These processes were accompanied by caspase activation and downregulation of the antiapoptotic proteins Bcl-2 and Bcl-x_L. Besides, the novel proapoptotic protein NSAID-activated gene (NAG-1) and its upstream regulator were overexpressed in drug-treated cells. Nevertheless, only ISL was found to induce a G2 arrest. These findings exemplify that both formononetin and ISL could cause growth inhibition and facilitate apoptosis in colon cancer cells, while only ISL is capable of inducing phase-specific cell cycle arrest. This suggests that the anticarcinogenic activities of different herbal flavonoids may involve both common and differential mechanisms of action, which could be developed as potential anticancer drugs.