Leaf Extract from European Olive (Olea europaea L.) Post-Transcriptionally Suppresses the Epithelial-Mesenchymal Transition and Sensitizes Gastric Cancer Cells to Chemotherapy.

The overall survival of patients with the advanced and recurrent gastric cancer (GC) remains unfavorable. In particular, this is due to cancer spreading and resistance to chemotherapy associated with the epithelial-mesenchymal transition (EMT) of tumor cells. EMT can be identified by the transcriptome profiling of GC for EMT markers. Indeed, analysis of the TCGA and GTEx databases (n = 408) and a cohort of GC patients (n = 43) revealed that expression of the CDH2 gene was significantly decreased in the tumors vs. non-tumor tissues and correlated with the overall survival of GC patients. Expression of the EMT-promoting transcription factors SNAIL and ZEB1 was significantly increased in GC. These data suggest that targeting the EMT might be an attractive therapeutic approach for patients with GC. Previously, we demonstrated a potent anti-cancer activity of the olive leaf extract (OLE). However, its effect on the EMT regulation in GC remained unknown. Here, we showed that OLE efficiently potentiated the inhibitory effect of the chemotherapeutic agents 5-fluorouracil (5-FU) and cisplatin (Cis) on the EMT and their pro-apoptotic activity, as was demonstrated by changes in the expression of the EMT markers (E- and N-cadherins, vimentin, claudin-1) in GC cells treated with the aforementioned chemotherapeutic agents in the presence of OLE. Thus, culturing GC cells with 5-FU + OLE or Cis + OLE attenuated the invasive properties of cancer cells. Importantly, upregulation of expression of the apoptotic markers (PARP cleaved form) and increase in the number of cells undergoing apoptosis (annexin V-positive) were observed for GC cells treated with a combination of OLE and 5-FU or Cis. Collectively, our data illustrate that OLE efficiently interferes with the EMT in GC cells and potentiates the pro-apoptotic activity of certain chemotherapeutic agents used for GC therapy.

The Anticancer Effect of Inula viscosa Methanol Extract by miRNAs' Re-regulation: An in vitro Study on Human Malignant Melanoma Cells.

Alternative and natural therapies are needed for malignant melanoma (MM), the most deadly skin cancer type due to chemotherapy's limited effect. In the present study, we evaluated the anticancer potentials of Inula viscosa methanol and water extracts (IVM and IVW) on MM cells, A2058 and MeWo, and normal fibroblasts. After the chromatographic and antioxidant activity analysis, their antiproliferative effects were determined with the increasing doses for 24-72 h. IVM induced more cell death in a dose and time-dependent manner in MM cells compared to IVW. This effect was probably due to the higher amount of phenolics in it. IVM significantly induced more apoptotic death in MM cells than fibroblasts (p < 0.01), which was also supported morphologically. IVM also caused cell cycle arrest at G0/G1 and G2/M phases in A2058 and MeWo, respectively, and suppressed the migration ability of MM cells (p < 0.01). Additionally, IVM was found to have significant potential in regulating MM-related miRNAs, upregulating miR-579 and miR-524, and downregulating miR-191 and miR-193, in MM cells (p < 0.05, p < 0.01). As a result, the anticancer effect of IVM via regulating miRNAs' expression has been demonstrated for the first time. Thus, IVM, with these potentials, may be a promising candidate for MM treatment.

Inhibitory Effects of Olea europaea Leaf Extract on Mesenchymal Transition Mechanism in Glioblastoma Cells.

BACKGROUND: Glioblastoma (GB) is the most aggressive form of brain tumor. Despite the current treatment methods, the survival rate of patients is very low. Therefore, there is a need to develop new therapeutic agents. The migration and invasion capacity of GB cells is related to mesenchymal transition (MT) mechanism. MATERIALS AND METHODS: The effect of OLE on MT was determined by analysis of the Twist, Snail, Zeb1, N-cadherin and E-cadherin genes in the EMT mechanism. The effect of OLE on cell migration was determined by wound healing test. RESULTS: 2 mg/ml OLE reduced Twist, Snail, Zeb1 and N-cadherin expression and the combination of OLE + TMZ (2 mg/ml OLE + 350 mM TMZ) increased E-cadherin and reduced Twist, Zeb1 and N-cadherin. In addition, co-treatment with OLE increased TMZ-induced anti-invasion properties thought suppressing transcription factors of MT mechanism. CONCLUSION: OLE can enhance the anti-MT activities of TMZ against GB and provide strong evidence that combined treatment with OLE and TMZ has the potential to be an effective alternative approach in GB therapy.

Investigation of new treatment option for hepatocellular carcinoma: a combination of sorafenib with usnic acid.

OBJECTIVES: Sorafenib (SOR) is an orally administered molecular targeted agent in the systemic chemotherapy of hepatocellular carcinoma (HCC). However, the partial response of SOR is limited due to its adverse side effect and high heterogeneity and resistant phenotype of HCC. In the current study, we investigated synergistic effects of SOR and usnic acid (UA) on HCC cell lines including HepG2 and SNU-449, and a normal cell line, HUVEC. METHODS: The antiproliferative and apoptotic effects of combination therapy and SOR alone were analysed by WST-1 and Annexin V analysis, respectively. Furthermore, cell cycle, gene expression analysis of SOR-targeted kinases and acridine orange-ethidium bromide staining were also performed in combined treatments. KEY FINDINGS: Our results demonstrated that SOR and UA combination indicated a strong synergism in HCC cell lines and reduced SOR toxicity in HUVEC cells. Additionally, the combination treatment SOR and UA significantly induced much more apoptotic cell death and G0/G1 arrest through downregulation of SOR-targeted kinases. CONCLUSIONS: Consequently, SOR and UA combination could be a new therapeutic strategy for HCC treatment.

Olea europaea Leaf Extract Improves the Efficacy of Temozolomide Therapy by Inducing MGMT Methylation and Reducing P53 Expression in Glioblastoma.

Unmethylated O6-methylguanine-DNA-methyltransferase (MGMT) promoter leads to Temozolomide (TMZ) resistance in most of the glioblastoma multiforme (GBM) patients. We previously investigated the synergistic effect of Olea europaea leaf extract (OLE) on TMZ cytotoxicity through modulating microRNA expression. To date, knowledge about the effect of OLE on MGMT methylation is insufficient. The aim of the current study was to evaluate the potential modulating effect of OLE on the TMZ response of GBM tumors through MGMT methylation. Exposure to 1 mg/mL OLE caused a significant induction of CpG island methylation in the MGMT gene using Methyl quantitative PCR assay (P < 0.001). In WST-1 analysis, the use of 350 µM TMZ plus 1 mg/mL OLE significantly increased the TMZ response of MGMT unmethylated cells (P = 0.003). Using the comet assay, the impact of 1 mg/mL OLE plus 350 µM TMZ on the formation of DNA strand breaks was significantly higher than that of 450 µM TMZ alone (P < 0.001) and Western blot analysis revealed that, when cells are treated with 1-mg/mL OLE, the total p53 protein levels tended to decrease. The results presented in this study uniquely demonstrated that OLE synergistically increased the TMZ response of GBM tumors by regulating MGMT gene methylation and p53 expression. However, further studies to validate our findings are required.

Olea europaea leaf extract and bevacizumab synergistically exhibit beneficial efficacy upon human glioblastoma cancer stem cells through reducing angiogenesis and invasion in vitro.

Patients with glioblastoma multiforme (GBM) that are cancer stem-cell-positive (GSC [+]) essentially cannot benefit from anti-angiogenic or anti-invasive therapy. In the present study, the potential anti-angiogenic and anti-invasive effects of Olea europaea (olive) leaf extract (OLE) were tested using GSC (+) tumours. OLE (2mg/mL) caused a significant reduction in tumour weight, vascularisation, invasiveness and migration (p=0.0001, p<0.001, p=0.004; respectively) that was associated with reducing the expression of VEGFA, MMP-2 and MMP-9. This effect was synergistically increased in combination with bevacizumab. Therefore, our current findings may contribute to research on drugs that inhibit the invasiveness of GBM.

Ficus carica latex prevents invasion through induction of let-7d expression in GBM cell lines.

Glioblastoma multiforme (GBM) is one of the deadliest human malignancies. A cure for GBM remains elusive, and the overall survival time is less than 1 year. Thus, the development of more efficient therapeutic approaches for the treatment of these patients is required. Induction of tumor cell death by certain phytochemicals derived from medicinal herbs and dietary plants has become a new frontier for cancer therapy research. Although the cancer suppressive effect of Ficus carica (fig) latex (FCL) has been determined in a few cancer types, the effect of this latex on GBM tumors has not been investigated. Therefore, in the current study, the anti-proliferative activity of FCL and the effect of the FCL-temozolomide (TMZ) combination were tested in the T98G, U-138 MG, and U-87 MG GBM cell lines using the WST-1 assay. The mechanism of cell death was analyzed using Annexin-V/FITC and TUNEL assays, and the effect of FCL on invasion was tested using the chick chorioallantoic membrane assay. To determine the effect of FCL on GBM progression, the expression levels of 40 GBM associated miRNAs were analyzed in T98G cells using RT-qPCR. According to the obtained data, FCL causes cell death in GBM cells with different responses to TMZ, and this effect is synergistically increased in combination with TMZ. In addition, the current study is the first to demonstrate the effect of FCL on modulation of let-7d expression, which may be an important underlying mechanism of the anti-invasive effect of this extract.

Olea europaea leaf extract alters microRNA expression in human glioblastoma cells.

PURPOSE: Glioblastoma multiforme (GBM) is the most common and the most lethal form of primary malignant tumors in the central nervous system. There is an increasing need for the development of more efficient therapeutic approaches for the treatment of these patients. One of the most attractive cancer therapy methods to date is the induction of tumor cell death by certain phytochemicals. Interestingly, bioactive compounds have been shown to alter micro RNA (miRNA) expression involved in several biological processes at the posttranscriptional level. The present study aimed to evaluate whether Olea europaea leaf extract (OLE) has an anticancer effect and modulates miRNA expression in GBM. MATERIALS AND METHODS: Firstly, the anti-proliferative activity of OLE and the nature of the interaction with temozolomide (TMZ) of OLE were tested in human glioblastoma cell line T98G cells by trypan blue and WST-1 assays and than realized miRNA PCR array analysis. Potential mRNA targets were analyzed bioinformatically. RESULTS: OLE exhibited anti-proliferative effects on T98G cell lines. Cells were treated with temozolomide (TMZ) in the presence OLE, and changes to miRNA expression levels were identified by PCR array analysis. miRNA target genes are involved in cell cycle and apoptotic pathways. Specifically, miR-181b, miR-153, miR-145, miR-137, and let-7d were significantly upregulated after treatment with both TMZ and OLE. CONCLUSION: Our results suggest that OLE modulates the expression of some miRNAs related to anticancer activity in GBM and the response to TMZ. Further studies and validations are needed, but we suggest that OLE might be used for in vivo studies and future medical drug studies.