Antioxidative and ROS-dependent apoptotic effects of Cuscuta reflexa Roxb. stem against human lung cancer: network pharmacology and in vitro experimental validation

Lung cancer remains a formidable global health challenge, necessitating the exploration of novel therapeutic approaches. This study investigates the potential of Cuscuta reflexa Roxb. stem extract as an anticancer agent against human lung cancer, focusing on its antioxidative and ROS-dependent apoptotic effects. Utilizing a combination of network pharmacology and in-vitro experimental validation, we delineate the multifaceted molecular mechanisms underlying the observed effects. The antioxidant potential of C. reflexa stem extract was evaluated by the 2,2-diphenyl-1-picrylhydrazyl (DPPH^•), 2,2-azinobis (3-ethyl-benzothiazoline-6-sulfonic acid) (ABTS^•+) and ferric reducing/antioxidant power (FRAP), hydroxyl free radical scavenging, reactive nitrogen oxide scavenging and super oxide anion radical scavenging assays. Furthermore, the antiproliferative and proapoptotic effect of C. reflexa stem extract was evaluated against A549 lung adenocarcinoma cell line using the consecrated sulforhodamine B (SBR) and Annexin V-PI assays. Additionally, the mitochondrial membrane potential (MMP) and the total reactive oxygen species (ROS) estimation assays were performed. As a result, network pharmacology analysis revealed a complex interaction network between the bioactive constituents of C. reflexa and key proteins implicated in lung cancer progression. The C. reflexa stem extract showed dose-dependent antioxidant activity against DPPH^• (IC₅₀ - 87.38 µg/mL), reactive nitrogen oxide (IC₅₀ - 318.34 µg/mL), FRAP (IC₅₀ - 359.96 µg/mL), hydroxy free radicals (IC₅₀ - 526.12 µg/mL) than ABTS^●+ (IC₅₀ - 698.45 µg/mL) and super oxide anion (IC₅₀ - 892.71 µg/mL) as well as cytotoxic activity against A549 cells (IC₅₀ - 436.80 µg/mL). Observations of morphological features in treated cells have revealed hallmark of apoptosis properties. Furthermore, as a result of treatment with C. reflexa stem extract, ROS generation and mitochondrial depolarization were increased in A549 cells, suggesting that this treatment has significant apoptotic properties. . These findings highlight the potential utility of this natural extract as an innovative therapeutic strategy for lung cancer treatment. The integration of network pharmacology and experimental validation enhances our understanding of the underlying mechanisms and provide the way for further translational research.Communicated by Ramaswamy H. Sarma.