Trifolin induces apoptosis via extrinsic and intrinsic pathways in the NCI-H460 human non-small cell lung-cancer cell line

Background: Trifolin (kaempferol-3-O-galactoside), which is a galactose-conjugated flavonol, exhibits Antifungal and Anticancer effects. However, the mechanisms underlying its Anticancer activities have not yet been examined.

Purpose: In this study, the Anticancer effects of trifolin were examined in human lung Cancer cells.

Methods: Cytotoxicity was determined by evaluating cell viability. Apoptosis was analyzed through flow cytometry and western blotting analysis. Death receptors and inhibitors of Apoptosis were evaluated through RT-PCR.

Results: Trifolin induced Apoptosis in NCI-H460 human non-small cell lung Cancer (NSCLC) cells by inhibiting the survival pathway and inducing the intrinsic and extrinsic Apoptosis pathways. Trifolin decreased levels of Akt/p-Akt, whereas levels of expression of phosphatidylinositide 3-kinase (PI3K), cyclin D1, cyclin E, and cyclin A were not altered. Trifolin initiated cytochrome c release by inducing mitochondrial outer membrane permeabilization (MOMP). Trifolin increased Bcl-2-associated X protein (Bax) levels and decreased b-cell lymphoma 2 (Bcl-2) levels, while the levels of Bcl-xL were not altered. In addition, trifolin increased the levels of the death receptor involving the Fas/Fas ligand (FasL) and Fas-associated protein with the death domain (FADD), which consequently activated Caspase-8, caspase-9, Caspase-3, and the proteolytic cleavage of poly (ADP-ribose) polymerase (PARP).

Conclusion: These results suggested that trifolin induced Apoptosis via death receptor-dependent and mitochondria-dependent pathways and that trifolin can be used as a therapeutic agent in human lung Cancer.