In vitro anti-cancer properties of a commercially available polyherbal nutraceutical (Vernolac) capsule on cancer stem cell-like (NTERA-2 cl.D1) cells.

Vernolac is a commercially available polyherbal nutraceutical formulation comprising Vernonia zeylanica aerial parts, Nigella sativa seeds, Hemidesmus indica roots, Smilax glabra rhizome, and Leucas zeylanica aerial parts. Previous studies have demonstrated anti-cancer activities of phytochemicals derived from these individual plant components. However, the anti-cancer properties of the supercritical CO2 extract of Vernolac remain unexplored against cancer stem-like cell populations. The current study is focused on the anti-cancer potential of Vernolac extract on cancer stem-like (CSCL) NTERA-2 cl.D1 cells, a human embryonal carcinoma-derived pluripotent cell line. Several in vitro assays evaluated the anti-cancer properties of the Vernolac extract. Cytotoxicity was assessed using the Sulforhodamine B assay, and apoptosis induction was determined by Acridine Orange/Ethidium Bromide staining and the caspase 3/7 activity assay. Scratch assay was used to evaluate cell migration and quantitative Reverse Transcription Polymerase Chain Reaction (RT-PCR) was performed to analyze the expression levels of apoptosis-related genes (TP53, Survivin) and the autophagy-related gene mTOR. Further, the free-radical scavenging activity of Vernolac extract was assessed using 2,2-diphenyl-1-picrylhydrazyl (DPPH), and the reactive oxygen species (ROS) levels in NTERA-2 cl.D1 cells were quantified using the nitroblue tetrazolium assay. Results demonstrated that Vernolac extract exhibits significant anti-proliferative activity in NTERA-2 cl.D1 cells, with an IC50 of 41.12 μg/mL at 48 h, while exerting minimal effects on non-cancerous MCF-10A cells (IC50 > 1000 μg/mL). Fluorescence microscopy and caspase 3/7 assay showed that Vernolac extract leads to early apoptosis in NTERA-2 cl.D1 cells. Quantitative RT-PCR revealed the upregulation of tumor suppressor protein TP53 while downregulating Survivin and mTOR genes. Additionally, Vernolac extract inhibited the migration rate of NTERA-2 cl.D1 cells and elevated intracellular reactive oxygen species (ROS) levels. These findings suggest that the supercritical CO2 extract of Vernolac exerts potent anticancer properties against NTERA-2 cl.D1 cancer stem-like cells, highlighting its therapeutic potential for targeting cancer stem cells.