Hyperforin-mediated anticancer mechanisms in MDA-MB-231 cell line: insights into apoptotic mediator modulation and caspase activation

Abstract

Breast cancer has been recorded as a frequent malignant cancer. The side effects of current chemotherapy drugs are still a severe problem. Hence, naturally occurring phytochemicals can be a possible solution to overcome this issue. Therefore, naturally occurring hyperforin was used to assess its anticancer effect against triple-negative breast cancer cells (MDA-MB-231) and its effectiveness against taxol (clinically used drug). Both compounds’ anticancer activity and mechanism of action were evaluated by analysing various approaches such as ELISA, flow cytometry and PCR microarray. Our results showed a lower IC50 of hyperforin (7.18 µg/mL) compared to taxol (7.91 µg/mL, P < 0.05) and hyperforin has almost similar anticancer effects as taxol against MDA-MB-231 cells. PCR microarray results showed that hyperforin induced the expression of caspases activator leading to a caspases-dependent cell death pathway whereas taxol uses TNF-mediated cell death. The results suggest that hyperforin may act as a good cell death-inducing agent. ABBREVIATIONS: TNBC: Triple negative breast cancer; DNA: Deoxyribonucleic acid; RNA: Ribonucleic acid; ROS: Reactive oxygen species; 5-FU: Fluorouracil; TRAIL: TNF-related apoptosis-inducing ligand; AKT: Protein kinase B; COX-2: Cyclooxygenase-2; PCR: Polymerase chain reaction; Bax: Bcl-2-associated X protein; BCL-2: B-cell lymphoma 2; MTT: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide; mRNA: Messenger ribonucleic acid; MCL1: Myeloid-cell leukemia 1; LDH: Lactate dehydrogenase; ELISA: Enzyme-linked immunosorbent assay; MMP: Mitochondrial membrane potential; MCP-1: Monocyte chemoattractant protein-1; ATCC: American Type Culture Collection; RPMI: Roswell Park Memorial Institute; XIAP: X-linked inhibitor of apoptosis protein; FBS: Fetal bovine serum; MCL-1: Myeloid-cell leukemia 1; DMEM: Dulbecco’s Modified Eagle Medium; DMSO: Dimethyl sulfoxide; CASP3: Caspase 3; RPMI: Roswell Park Memorial Institute; DMEM: Dulbecco’s Modified Eagle Medium; FBS: Fetal bovine serum; h: Hour; TNF-α: Tumour necrosis factor alpha; P53: Tumor protein P53; min: Minutes; cps: Count presecond; OD: Optical density; rpm: Revolutions per minute; PI: Propidium iodide; SD: Standard deviation; DDR: Death Domoin Receptor; TRADD: Tumor necrosis factor receptor type 1-associated DEATH domain; HRK: Harakiri, BCL2 Interacting Protein; RIPK2: Receptor-interacting serine/threonine-protein kinase 2; BIK: BCL2 interacting killer; DCF: 2’, 7’ –dichlorofluorescein; DCF-DA: Dichlorofluorescin Diacetate; FADD: Fas Associated Via Death Domain; TNFRSF: Tumor necrosis factor receptor superfamily; FITC: Fluorescein isothiocyanate; DMSO: Dimethyl sulfoxide; CFLAR: CASP8 And FADD Like Apoptosis Regulator; BID: BH3-interacting domain death agonist; BIRC: Baculoviral IAP repeat-containing protein; CIDEA: Cell death activator CIDE-A; CIDEB: Cell Death Inducing DFFA Like Effector B; BrdU: Bromodeoxyuridine; RFU: Relative fluorescence unit; NF-kB: Nuclear Factor kappa-light-chain-enhancer of activated B cells; %: Percentage; µL: microliter; µM: Micromolar; °C: degree Celcius; ng/mL: Nanograms per millimeter; mg/mL: Milligrams per milliliter; µg/mL: Micrograms per milliliter; ΔΨm: Mitochondrial membrane potential; TNFα: Tumour necrosis factor α; Bcl-xL: B-cell lymphoma-extra large; MDR1: Multidrug Resistance Mutation; AIB1: Amplified in breast cancer 1; LTBR: Lymphotoxin Beta Receptor Highlights Utilization of hyperforin as anticancer agent against triple negative breast cancer. Hyperforin exhibited a lower IC50 compared to taxol in inhibiting growth of MDA-MB-231 cells. Hyperforin shows similar potential anticancer activity as taxol. Hyperforin uses mitochondrial death pathway to induce cell death in MDA-MB-231 cells.