Screening methanolic extracts of Sutherlandia spp as anti-tumor agents and their effects on anti-apoptotic genes

Abstract

Cervical cancer is the most common malignancy after breast cancer in women worldwide. It accounts for 83% of all new cases and 85% cancer death in developing countries. In South Africa, cervical cancer is the common cancer in women with an annual crude incidence rate of 30.2 per 100,000 women and the highest rate were found to be in women between the ages of 66-69 years. Approximately 6800 women in S.A face new case of cervical cancer while accounting for 3700 cancer death annually. Because of unequal access to the health facilities, socio-economic differences, HPV and HIV infection, the rate of cervical cancer in black women is higher (42.1%) compared to the low rate in white women. Because of the name “cancer bush’ given to Sutherlandia Frutescence(S.F) plant by the traditional healers as well as Xhosas, Zulu, Sotho and cape Dutch for its anti-cancer activity, the plant was in this study to confirm its cytotoxic effect on the cervical cancer cell lines. Aim of the study: to evaluate the methanolic extracts of Sutherlandia Frutescens on cervical cancer cell lines. Materials and Methods: The MTT assay was performed to evaluate SiHa cell lines treated with methanolic extract of S.F (50μg/ml, 100μg/ml, 150μg/ml and 200μg/ml). The three compounds (Canavanine, GABA and Pinitol) were also evaluated for its anti-tumour activity. The cell growth was then showed in real time using Xcilligence. Flow cytometry was employed to determine the mode of action. Caspase 3/7 assay was performed to confirm if cell death was via caspase-dependent or independent and ATP was done to assess the ATP level in S.F treated cells. Results: MTT shows a significant decline in cells treated with 50μg/ml, 100μg/ml and 200μg/ml of the extract and 50μg/ml was concluded to be the concentration at which 50% of the cells die. The ATP results are inconsistent with MTT result; the ATP level increased in S.F treated cells. Cell index which represents the quantitative measure of cell growth in real time decline upon treatment with 50μg/ml. Flow cytometry showed cells are dying by apoptosis and the cell cycle arrest is mostly in the S phase. The cell death was caspase-dependent as it shows an increased luminescence which is proportional to the number of caspase. The concentrations of the compounds used, Canavanine (1000μM, 1500μM and 2500μM), GABA (100μM, 300μM and 500μM) and Pinitol (30μM, 90μM and 120μM) induce cell death and cell death shows to decrease after the maximum concentration. Conclusion: Sutherlandia Frutescence has proven with number of research that it induces cell death via apoptosis. After evaluating its cytotoxicity, the plant shows to be a promising anti-tumor agent that needs to be clinically proven.