Bio-guided isolation and bioinformatic studies of cytotoxic phytosterols from Acanthospermum hispidum DC against breast (MCF7) and colorectal (HT29) cancer cells

Acanthospermum hispidum DC (Asteraceae), a traditional medicinal plant, plays a role as an alternative remedy for various diseases, such as bacterial and viral infections, jaundice, malaria, fever, gastrointestinal disorders, headache, convulsions, and snake bites. Flavonoids, alkaloids, saponins, tannins, terpenes, steroids, and cardioactive glycosides are the distinct classes of metabolites in the plant. Although A. hispidum was suggested as a promising antitumor phytomedicine, no studies identified its potential cytotoxic components. In this study, the cytotoxic compounds of A. hispidum were isolated using chromatographic techniques guided by in vitro MTT cytotoxicity assay against selected cancer cell lines; breast cancer (MCF7), colorectal adenocarcinoma (HT29), and hepatoblastoma (HepG2). The selective index (SI) was assessed on MRC5 (Normal human fetal lung fibroblast) cell line. The dichloromethane fraction (DCM) showed remarkable cytotoxic activity against MCF7 and HT29 (%Cell viability; 46.15 and 60.5, respectively). Hence, the main bioactive fraction from DCM was purified to afford two phytosterols; stigmasterol (1) and β-sitosterol (2), which were identified by 1-D and 2-D NMR spectroscopy. Cytotoxic evaluation of 1 and 2 revealed that β-sitosterol showed better selective cytotoxicity against MCF7 and HT29 (IC₅₀ 4.07 μg/mL, SI 2.63; IC₅₀ 4.52 μg/mL, SI 2.37) compared to stigmasterol (IC₅₀ 5.43 μg/mL, SI 1.38; IC₅₀ 4.21 μg/mL, SI 1.78), respectively. Bioinformatic assessments of drug-likeness and ADMET properties demonstrated that most criteria were obeyed by the investigated compounds except for their poor solubility, which recommended the preparation of special dosage forms, such as nanoformulation to enhance their oral bioavailability. Swiss Target prediction indicated that nuclear receptors represent the main target class (40%). Whereas caspase-3 stimulant activity (a key enzyme in apoptosis) was predicted by the PASS prediction tool as a potential anticancer mechanism. Our study suggests A. hispidum as a potential source of bioactive phytosterols and as a chemopreventive medicinal plant.