Cytotoxicity of Anchusa arvensis Against HepG-2 Cell Lines: Mechanistic and Computational Approaches

Abstract

Background: Liver cancer is a devastating cancer with increasing incidence and mortality rates worldwide. Plants possess numerous therapeutic properties, therefore the search for novel, naturally occurring cytotoxic compounds is urgently needed. Methods: The anticancer activity of plant extracts and isolated compounds from Anchusa arvensis (A. arvensis) were studied against the cell culture of HepG-2 (human hepatocellular carcinoma cell lines) using 3-(4,5-Dimethylthiazol-yl)-diphenyl tetrazoliumbromide (MTT) assay. Apoptosis was investigated by performing Acridine orange –ethidium bromide staining, styox green assay and DNA interaction study. We also used tools for computational chemistry studies of isolated compounds with the tyrosine kinase. Results: In MTT assay, the crude extract caused a significant cytotoxic effect with IC50 of 34.14 ± 0.9 μg/ml against HepG-2 cell lines. Upon fractionation, chloroform fraction (Aa.Chm) exhibited the highest antiproliferative activity with IC50 6.55 ± 1.2 μg/ml followed by ethyl acetate (Aa.Et) fraction (IC50, 24.59 ± 0.85 μg/ml) and n-hexane (Aa.Hex) fraction (IC50 29.53 ± 1.5μg/ml). However, the aqueous (Aa.Aq) fraction did not show any anti-proliferative activity. Bioactivity-guided isolation led to the isolation of two compounds which were characterized as para–methoxycatechol (1) and decane (2) through various spectroscopic techniques. Against HepG-2 cells, compound 1 showed marked potency with IC50 6.03 ± 0.75 μg/ml followed by 2 with IC50 18.52 ± 1.9 μg/ml. DMSO was used as a negative control and doxorubicin as a reference standard (IC50 1.3 ± 0.21 μg/ml). It was observed that compounds 1-2 caused apoptotic cell death evaluated by Acridine orange –ethidium bromide staining, styox green assay and DNA interaction study, therefore both compounds were tested for molecular docking studies against tyrosine kinase to support cytotoxic activity. Conclusion: This study revealed that the plant extracts and isolated compounds possess promising antiproliferative activity against HepG-2 cell lines via apoptotic cell death.