METHODS AND RESULTS: The leaves extracts were analysed for its antiproliferative effect on breast cancer (MCF7) cells and normal epithelial breast (MCF 10A) cells using Sulforhodamine B (SRB) assay. The selective extract was evaluated for its ability to induce apoptosis using Annexin V-FITC apoptosis staining and the expression of molecular genes using qualitative reverse transcription-polymerase chain reaction (RT-PCR) against MCF7 cells. Gas chromatography-mass spectrometry (GC-MS) was used to identify the compounds from the selective extract. The findings showed that dichloromethane fraction (CV-Dcm) extract had high antiproliferative effect against MCF7 cells (IC50 = 24 µg/mL, selective index (SI) = 8.17). The percentages of apoptosis cells in CV-Dcm-treated MCF7 cells was 58.8%. The CV-Dcm extract induced downregulation of PCNA level. The apoptotic genes were also triggered in both extrinsic and intrinsic signaling pathways, affecting a 1.5-fold increase in BAX, 1.4-fold increase in cytochrome c, 1.3-fold increase in caspase-8, 1.7-fold increase in caspase-3 and 0.5-fold-decrease in BCL-2. Treated MCF7 cells also activated P53-dependent apoptotic death pathway.
CONCLUSIONS: The present work strongly suggests that high efficacy of CV-Dcm extract was attributed to its antiproliferative and apoptosis-inducing activation in MCF7 cells, most likely due to its favourable compounds.
OBJECTIVE: This study investigates the chemical constituents, anti-proliferative, and apoptotic properties of C. nutans root extracts.
MATERIALS AND METHODS: The roots were subjected to solvent extraction using methanol and ethyl acetate. The anti-proliferative effects of root extracts were tested at the concentrations of 10 to 50 μg/mL on MCF-7 and HeLa by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay for 72 h. Morphological changes were observed under light microscope. Pro-apoptotic effects of root extracts were examined using flow cytometric analysis and RT-PCR. The chemical compositions of root extracts were detected using GC-MS.
RESULTS: The proliferation of MCF-7 cells was inhibited with the IC50 values of 35 and 30 μg/mL, respectively, for methanol and ethyl acetate root extracts. The average inhibition of HeLa cells was ∼25%. Induction of apoptosis in MCF-7 was supported by chromatin condensation, down-regulation of BCL2 and unaltered expression of BAX. However, only ethyl acetate extract caused the loss of mitochondrial membrane potential. GC-MS analysis revealed the roots extracts were rich with terpenoids and phytosterols.
DISCUSSION AND CONCLUSIONS: The results demonstrated that root extracts promote apoptosis by suppressing BCL2 via mitochondria-dependent or independent manner. The identified compounds might work solely or cooperatively in regulating apoptosis. However, further studies are required to address this.
PURPOSE: Here, we investigated whether OSCC cells were sensitive towards zerumbone treatment and further determined the molecular pathways involved in the mechanism of action.
METHODS: Cytotoxicity, anti-proliferative, anti-migratory and anti-invasive effects of zerumbone were tested on a panel of OSCC cell lines. The mechanism of action of zerumbone was investigated by analysing the effects on the CXCR4-RhoA and PI3K-mTOR pathways by western blotting.
RESULTS: Our panel of OSCC cells was broadly sensitive towards zerumbone with IC50 values of less than 5 µM whereas normal keratinocyte cells were less responsive with IC50 values of more than 25 µM. Representative OSCC cells revealed that zerumbone inhibited OSCC proliferation and induced cell cycle arrest and apoptosis. In addition, zerumbone treatment inhibited migration and invasion of OSCC cells, with concurrent suppression of endogenous CXCR4 protein expression in a time and dose-dependent manner. RhoA-pull down assay showed reduction in the expression of RhoA-GTP, suggesting the inactivation of RhoA by zerumbone. In association with this, zerumbone also inhibited the PI3K-mTOR pathway through the inactivation of Akt and S6 proteins.
CONCLUSION: We provide evidence that zerumbone could inhibit the activation of CXCR4-RhoA and PI3K-mTOR signaling pathways leading to the reduced cell viability of OSCC cells. Our results suggest that zerumbone is a promising phytoagent for development of new therapeutics for OSCC treatment.