Flavonol isolated from ethanolic leaf extract of Thuja occidentalis arrests the cell cycle at G2‐M and induces ROS ‐independent apoptosis in A549 cells, targeting nuclear DNA

Objectives The K‐ras gene mutation commonly found in lung adenocarcinomas contributes to their non‐invasive expansion. Our main objective here was to develop a chemopreventive agent against K‐ras ‐mutated lung adenocarcinoma cell line like‐A549. Materials and methods We isolated flavonol from ethanolic leaf extract of Thuja occidentalis, and evaluated its apoptotic potentials on A549 cells. They were treated with 1–10 μg/ml of flavonol and viability was tested retaining normal lung cells L‐132 as control. We performed assays such as TUNEL , annexin V, cell‐cycle and mitochondrial membrane potentials, by FACS analysis. ROS ‐mediated oxidative stress and drug– DNA interactions were analysed along with gene expression studies for p53 , Bax‐Bcl2 , cytochrome c , the caspase cascade genes and PARP . Results Flavonol reduced A549 cell viability in a dose‐ and time‐dependent manner (IC 50 value = 7.6 ± 0.05 μg/ml following 48 h incubation) sparing normal L‐132 cells. It effected G2‐M phase cell cycle arrest and apoptosis, as indicated by progressive increase in the sub‐G1, annexin V and TUNEL ‐positive cell populations. Apoptotic effects appeared to be mitochondria‐dependent, caspase‐3‐mediated, but ROS ‐independent. Analysis of circular dichroism data revealed that flavonol intercalated with nuclear DNA . In vivo studies on non small cell lung carcinoma ( NSCLC )‐induced mice confirmed anti‐cancer potential of flavonol. Conclusion Flavonol‐induced apoptosis apparently resulted from intercalation of cells' nuclear DNA . Flavonol inhibited growth of induced lung tumours in the mice, indicating its potential as an effective agent against NSCLC .