Selective Cytotoxicity and Pro-apoptotic Activity of Stem Bark of Wrightia tinctoria (Roxb.) R. Br. in Cancerous Cells.

Abstract

Background:Wrightia tinctoria (Roxb.) R. Br. is a widely available shrub in India used traditionally in various ailments, including cancer. However, the anticancer activity of the bioactive fractions has not been validated scientifically.Objective:To investigate the anticancer potential of stem bark of W. tinctoria and establish its phytochemical basis.Materials and Methods:The ethanol extract and subsequent fractions, petroleum ether, ethyl acetate, n-butanol, and aqueous were prepared by standard methods. In vitro cytotoxicity was determined in MCF-7 (breast) and HeLa (cervical) adenocarcinoma cells, and V79 (nontumor fibroblast) cells and apoptogenic activity in MCF-7 cells by acridine orange (AO)/ethidium bromide (EB) staining. Additionally, the antioxidant potential was evaluated using suitable methods. High-performance thin layer chromatography (HPTLC) analysis was performed for identification of active phytoconstituents.Results:Petroleum ether and ethyl acetate fractions were most potent with IC50 values of 37.78 and 29.69 μg/ml in HeLa and 31.56 and 32.63 μg/ml in MCF-7 cells respectively in the sulforhodamine B assay. Comparable results were obtained in HeLa cells in 3-(4,5-dimethylthiazolyl-2-yl)-2,5-diphenyl tetrazolium bromide assay and interestingly, the fractions were found to be safe to noncancerous fibroblast cells. Both fractions induced significant (P < 0.05) apoptotic morphological changes observed by AO/EB staining. Moreover, extract/fractions exhibited excellent inhibition of lipid peroxidation with the ethyl acetate fraction being most active (IC50:23.40 μg/ml). HPTLC confirmed the presence of two anti-cancer triterpenoids, lupeol, and β-sitosterol in active fractions.Conclusion:Extract/fractions of W. tinctoria exhibit selective cytotoxicity against cancerous cells that is mediated by apoptosis. Fractions are less toxic to noncancerous cells; hence, they can be developed as safer chemopreventive agents.SUMMARY Petroleum ether and ethyl acetate fractions were most active and exhibited dose-dependent cytotoxicity in HeLa and MCF-7 cells.Fractions were relatively less toxic to non-tumor fibroblast cells demonstrating its selectivity to cancer cells.Fractions exhibited pro-apoptotic activity in MCF-7 cells in AO/EB staining.Lupeol and β-sitosterol were identified as anticancer constituents by HPTLC.