Methanol extract of Pergularia daemia (Forssk.) Chiov. leaves induce apoptosis in triple-negative breast cancer through intrinsic pathway

Abstract

Most traditional anticancer medications are ineffective in treating triple negative breast cancer (TNBC), which has enhanced resistance to chemotherapeutic agents. Herbs continue to be a key element in the discovery of anticancer medications. In the present study, we assessed the cytotoxicity of methanol extract of leaves of Pergularia daemia (MLPD) in MDA-MB-231 triple-negative breast cancer (TNBC) cell line. The cytotoxicity of MLPD was assessed using MTT assay which revealed significant concentration-dependent cytotoxicity with an IC₅₀ value of 35.95±3.57 µg/mL. Morphological evaluation of MDA-MB-231 cells treated with MLPD showed cytotoxic changes like vacuole formation, altered morphology, and reduction in the number of cells with apoptotic bodies formation. Staining techniques, such as acridine orange and ethidium bromide (AO/EB) showed early apoptosis characterized by yellow-green fluorescence and crescent-shaped nucleus. Hoechst 33258 staining of MDA-MB-231 cells treated with MLPD showed nuclear marginalization indicative of nuclear apoptotic changes. To assess the mitochondrial membrane potential (MMP), we did JC-1 staining and the results revealed green fluorescence indicative of mitochondrial dependent apoptotic pathway. Comet assay confirmed significant DNA damaging property in MDA-MB-231 cells treated with MLPD. Western blotting analysis showed significant downregulation of Bcl-2 by 0.52±0.03 folds with no significant change in caspase-8 expression. Hence, the present study demonstrated that MLPD possessed potent cytotoxic effect against MDA-MB-231 TNBC cells through mitochondria mediated intrinsic pathway of apoptosis.