Apoptotic and Antiproliferative Effects of <i>Asparagus racemosus</i> Willd. via Oxidative Stress and Caspase Activation in HCT‐116 Cells

Cancer remains a global health challenge, with colorectal cancer ranking third in prevalence and second in cancer-related deaths worldwide. Conventional treatments have limitations, leading to increased interest in phytotherapy as a potential alternative. Asparagus racemosus Willd., known for its pharmacological properties, remains scientifically underexplored. Our previous study demonstrated that the ethanolic extract of A. racemosus Willd. exhibited potent cytotoxic effects on HCT-116 colorectal cancer cells, prompting further investigation of its therapeutic potential. In this study, A. racemosus Willd. ethanolic extract (ARE)'s apoptotic effects on HCT-116 cells were explored through various molecular techniques. DAPI (4'-6'-diamidino-2-phenylindole) staining showed clear signs of apoptosis, and DCFH-DA (2',7'-dichlorodihydrofluorescein diacetate) staining indicated an ROS-dependent apoptotic pathway. Rhodamine staining revealed mitochondrial membrane potential disruption, suggesting the involvement of the intrinsic pathway. Annexin V assays showed a significant increase in apoptotic cells, and DNA fragmentation confirmed apoptosis. Cell cycle analysis indicated cell cycle arrest, whereas Caspase-3/7 activity assays and Western blotting confirmed apoptosis induction via a Caspase-3-dependent pathway. These findings suggest that ARE is a potent natural anticancer agent targeting colorectal cancer through interconnected apoptotic pathways. This research highlights the potential of ARE for developing a nontoxic, cost-effective anticancer treatment, offering hope for colorectal cancer patients.