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PI3K/AKT/mTOR signaling promotes cell survival, proliferation and progression in cancer cells. Targeting this pathway may lead to the development of novel therapeutic approaches for human cancers. Here, we examined the effects of (-)-epigallocatechin-3-gallate (EGCG) on the PI3K/AKT/mTOR pathway in pancreatic cancer cells, and assessed its therapeutic potential. In this study, the proliferation and apoptosis of PANC-1 cells were examined by MTT assay and flow cytometry, respectively. The expression of genes and proteins involved in the PI3K/AKT/mTOR pathway were measured by RT-PCR and western blotting, respectively. Our results revealed that EGCG dramatically inhibited the proliferation of PANC-1 cells and induced apoptosis simultaneously. Furthermore, it upregulated PTEN mRNA and protein expression levels, as well as downregulating the expression of phospho-AKT and phospho-mTOR. In conclusion, these results suggest that EGCG can suppress proliferation and induce apoptosis of PANC-1 cells in a time- and dose-dependent manner; moreover, EGCG also can upregulate PTEN expression and downregulate the expression of pAKT and p-mTOR to modulate the PI3K/AKT/mTOR signaling pathway.

α-Phellandrene (α-PA) is a cyclic monoterpene. To investigate the induction of autophagy by α-PA and its mechanism, human liver tumor cells (J5) were incubated with α-PA and analyzed for cell viability and the molecular regulation of pre-autophagosome origination and autophagosome formation. According to the results, PI3K-I, mTOR, and Akt protein levels were decreased after α-PA treatment compared to those of the control group (p < 0.05). The phosphorylation of Bcl-2, and PI3K-III, LC3-II and Beclin-1 protein levels in J5 cells were increased after α-PA treatment (p < 0.05). In addition, α-PA up-regulated nuclear p53 and down-regulated cytoplasmic p53 expression in J5 cells. The NF-κB pathway was activated, as indicated by increase in cytosolic phosphorylated IκB, nuclear NF-κB levels, and the DNA-binding activity of NF-κB after α-PA treatment in J5 cells (p < 0.05). These results suggest that α-PA can induce J5 cell autophagy by regulating mTOR and LC-3II expression, p53 signaling, and NF-κB activation in J5 cells.

Aloe-emodin (AE) is derived from Aloe vera and rhubarb (Rheum palmatum) and exhibits anticancer activities via multiple regulatory mechanisms in various cancers. AE can also enhance the anticancer efficacy of cisplatin, doxorubicin, docetaxel, and 5-fluorouracil; however, its effects remain poorly characterized. MCF-7, MDA-MB-231, MDA-MB-468, BT-474, and HCC-1954 breast cancer cell lines were treated with the indicated conditions of AE, and cell viability assays were performed. The expression levels of signaling proteins were determined by western blot analysis, intracellular reactive oxygen species (ROS), cell cycle distributions, and rates of apoptosis as estimated by flow cytometry. In comparison with other cells, MCF-7 cells were more sensitive to AE treatment; AE enhanced the cytotoxicity of 9$\mu$g/ml tamoxifen by reducing EGFR, ER$\alpha$, Ras, ERK, c-Myc, and mTOR protein expression and blocking PI3K and mTOR activation. Finally, although co-treatment of AE with tamoxifen increased intracellular ROS, there were no effects on cell cycle progression. Besides facilitating tamoxifen-induced cell death, AE also enhanced the antiproliferative activity of tamoxifen by blocking Ras/ERK and PI3K/mTOR pathways in breast cancer cells, thus demonstrating the chemosensitizing potential of AE.

Puerarin is a traditional Chinese medicine with beneficial effects of reduced depression-like behaviors in mice with stress. Previous studies also show that puerarin can produce neuroprotective effect via activating the Akt or increased brain-derived neurotrophic factor (BDNF) expression. Interestingly, BDNF and Akt downstream target, mammalian target of rapamycin (mTOR) mediate the fast-acting antidepressant properties of ketamine. Until now, the involvement of the mTOR signaling pathway or BDNF on puerarin-induced antidepressant effect remains unknown. We aimed to investigate whether the antidepressant-like effect induced by puerarin would associate mTOR signaling pathway and BDNF release. The antidepressant-like effects of puerarin were evaluated using the forced swim test. The activation of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionaic acid receptor (AMPAR)-mTOR signaling pathway and release of BDNF in the prefrontal cortex were determined. We also investigated the effect of puerarin on AMPAR trafficking through measuring the PKA phosphorylation of AMPAR subunit GluR1. Our present results show that puerarin exerted antidepressant-like responses that was mediated by AMPAR-induced mTOR signaling pathway and associated with increased BDNF release. Moreover, a significant increase in the GluR1 phosphorylation at its PKA site was noted following puerarin treatment. Our findings are the first to demonstrate that the antidepressant-like actions of puerarin require AMPAR–mTOR signaling pathway activation, are associated with an increased BDNF level and facilitate AMPAR membrane insertion. These findings provide preclinical evidence that puerarin may possess antidepressant property which is mediated by the glutamatergic system.

Autophagy is a process of active programmed cell death, where a dying cell induces autophagosomes and subsequently regulated by degradative machinery. The aim of this study was to investigate the mechanism behind induction of autophagic cell death by Naringin flavonoid in AGS cancer cells. Growth inhibition of AGS cells showed downregulation of PI3K/Akt/mTOR signaling by Naringin treatment. Transmission electron microscopy observation showed swollen mitochondria and lysosome near peri-nuclear zone fused with autophagic vacuoles. Rapamycin pre-treatment with Naringin showed significant decrease in mTOR phosphorylation and increase in LC3B activation in AGS cells. Decrease in mTOR phosphorylation is associated with lysosomal function activation was observed by time-dependent treatment of Naringin. Induction of lysosomal membrane permeabilization (LMP) was observed by LAMP1 activation leading lysosomal cell death by releasing Cathepsin D from lysosomal lumen to cytosol. Naringin treated AGS cells showed up-regulating BH3 domain Bad, down-regulating Bcl-xL, and Bad phosphorylation and significant mitochondrial fluorescence intensity expression. Significant localization of mitochondria and LC3B activation was examined by person coefficient correlation. Activation of ERK1/2-p38 MAPKs and production of intracellular ROS has been observed over Naringin treatment. It has also been elucidated that pre-treatment with NAC inhibited mitochondria-LC3B colocalization, where ROS acted as upstream of ERK1/2-p38 MAPKs activation. Lysosomal cell death involvement has been evaluated by BAF A1 pre-treatment, inhibiting LAMP1, Cathepsin D, ROS, and blocking autophagolysosome in AGS cell death. Taken together, these findings show that, Naringin induced autophagy cell death involves LMP mediated lysosomal damage and BH3 protein Bad activation in AGS cancer cells.

Glioblastoma multiforme (GBM) is a deadly malignant brain tumor that is resistant to most clinical treatments. Novel therapeutic agents that are effective against GBM are required. Antrodia cinnamomea has shown antiproliferative effects in GBM cells. However, the exact mechanisms and bioactive components remain unclear. Thus, the present study aimed to investigate the effect and mechanism of 4-acetylantrocamol LT3 (4AALT3), a new ubiquinone from Antrodia cinnamomeamycelium, in vitro. U87 and U251 cell lines were treated with the indicated concentration of 4AALT3. Cell viability, cell colony-forming ability, migration, and the expression of proteins in well-known signaling pathways involved in the malignant properties of glioblastoma were then analyzed by CCK-8, colony formation, wound healing, and western blotting assays, respectively. We found that 4AALT3 significantly decreased cell viability, colony formation, and cell migration in both in vitro models. The epidermal growth factor receptor (EGFR), phosphatidylinositol-3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR), Hippo/yes-associated protein (YAP), and cAMP-response element binding protein (CREB) pathways were suppressed by 4AALT3. Moreover, 4AALT3 decreased the level of DNA repair enzyme O⁶-methylguanine-DNA methyltransferase and showed a synergistic effect with temozolomide. Our findings provide the basis for exploring the beneficial effect of 4AALT3 on GBM in vivo.