Abstract 779: Investigation of the anti-breast cancer activity of five traditionally used Sudanese medicinal plants

Abstract

Abstract Five Sudanese medicinal plants were selected on the basis of their traditional use for treatment of breast cancer in Sudan. After collection and authentication, plants were dried, ground and extracted with methanol. Growth inhibitory properties of extracts were then examined by MTT assay against 4 breast carcinoma, 1 pancreatic carcinoma, a normal non- transformed fibroblast and normal endothelial cell line. Two extracts (6 and 7 from Indigofera astragalina) demonstrated the highest anticancer activity. Viable cell counts were performed after treatment of MDA468 cells with extract 6 and cell cycle perturbations examined. Cells undergoing early apoptosis were detected by flow cytometry (annexin V), simultaneous cell cycle/annexin V staining revealed from which phase apoptotic populations emerged, finally, detection of cleaved PARP by Western blot corroborated apoptosis. Extract 6 was then fractioned by liquid extraction, flash chromatography and HPLC, and MTT assays conducted to verify the activity of the fractions. Extract 6 showed selective activity against breast cancer cell lines (GI50: 12-31µg/ml for breast cell lines, 62 µg/ml for pancreas cell line, 69 µg/ml for the fibroblast, and 33 µg/ml for the endothelial cells). Extract 6 decreased the number of viable MDA468 cells in a dose- and time-dependent manner. Total preG1 and G2/M cell cycle events increased slightly (4-9% and 35-42% respectively) after 48h 50-100 µg/ml treatment, and Annexin V staining/PI exclusion revealed early apoptotic cells (40-46%) after 24h treatment (50-100 µg/ml). Simultaneous cell cycle/apoptosis examination exposed apoptotic populations emerging from each cell cycle compartment. The intensity of cleaved PARP proteins detected after treatment (50-100 µg/ml; 24h, 48h) increased in a dose- and time-dependent manner. Three fractions were successively partitioned from water into hexane and ethyl acetate/chloroform. The hexane fraction was inactive against MDA468, the aqueous fraction showed weak activity, whereas the ethyl acetate/chloroform fraction showed the highest activity (GI50: 23 µg/ml). The latter gave 16 sub-fractions by normal phase silica gel flash chromatography, of which fractions 9, 10, 12 and 13 demonstrated the highest growth inhibitory activity (GI50 5, 9, 10 and 7 μg/ ml respectively) against MDA 468 cells. Fraction 9 (yield = 0.066% of original methanol extract) was less potent against fibroblast cells (32 μg/ml). Analytical RP-HPLC fractionation of fraction 9 resulted in 3 main components. In conclusion, extract 6 possesses growth inhibitory and pro-apoptotic properties against human MDA468 breast cancer cells. To date no investigations of the pharmacological activity of I. astragalina have been reported. Isolation and identification of active compounds are underway. Keywords: Sudan, traditional medicinal plants, Indigofera astragalina, breast cancer, apoptotic activity. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 779.