Euphorbia abyssinica Gmel. has been traditionally used in Ethiopia to treat various diseases, including swelling and cancer. This study assessed the cytotoxic effects of the ethanolic crude extract and its isolates from the aerial parts of E. abyssinica using the MTT assay method and molecular docking studies with the AutoDock Vina 4.2 program. E. abyssinica showed strong cytotoxicity against HeLa cells (IC 50= 18.6±0.7 μg/mL) and weak cytotoxicity against PC3 cells, with percent inhibition of 55.2% and 23.7% at 30.0 μg/mL, respectively. Among the fractions, the Dichloromethane fraction exhibited strong cytotoxicity against PC3 cells (IC 50= 21.7±0.9 μg/mL) and weak cytotoxicity against Hela cells, with 78.4% and 29.98% growth inhibition at 30.0 μg/mL, respectively. Dichloromethane fraction yielded β-sitosterol (1), friedelin (2) epifriedelanol (3), taraxerol (4) and glutinol (5). Friedelin (2) demonstrated strong cytotoxic activity against HeLa and PC3 cells. Similarly, epifriedelanol (3) showed strong cytotoxicity against PC3 cells (IC 50= 28.4±0.7 µM) and notable growth inhibition of HeLa cells (38.5% at 30 µM). Friedelin (2) showed better binding affinity and inhibition constant towards Human prostate specific antigen (PDB: 2ZCH)(-9.01 kcal/mol and 0.25 µM) and Alpha-actinin-4 (PDB: 6O31)(-7.61 kcal/mol and 2.64 µM).

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