Plumbagin Induces ROS-Independent DNA Damage Leading to Apoptosis in Non-Small Cell Lung Cancer

Abstract

Lung cancer remains the leading cause of cancer-related deaths worldwide. Although chemotherapy is a primary treatment, its efficacy is limited by toxicity and drug resistance. Phytocompounds such as Plumbagin, derived from Plumbago zeylanica, have shown promising anticancer activity, but their precise mechanisms remain unclear. Plumbagin is known to induce reactive oxygen species (ROS), leading to DNA damage and apoptosis; however, whether it directly binds to DNA and induces ROS-independent damage is unknown. To address this, we evaluated the cytotoxic and DNA-damaging potential of Plumbagin in non-small cell lung cancer cells. DAPI, AO/EtBr, and γ-H2AX assays demonstrated significant DNA damage even after ROS inhibition with N-acetylcysteine (NAC). Gene expression analysis showed alteration in DNA damage response genes. DNA laddering assay confirmed fragmentation despite ROS inhibition, suggesting a ROS-independent mechanism. To assess direct DNA binding, we employed UV–VIS spectroscopy, differential scanning calorimetry (DSC), circular dichroism (CD), and isothermal titration calorimetry (ITC). These revealed that Plumbagin interacts weakly with DNA, insufficient to induce direct damage. Molecular dynamics (MD) simulations further indicated that the Plumbagin–DNA complex is stable but not disruptive to DNA structure. Overall, our results suggest that Plumbagin induces DNA damage in lung cancer cells via both ROS-dependent and ROS-independent mechanisms, with weak direct DNA interaction, indicating the involvement of alternate molecular targets contributing to its genotoxic potential.