Miyake N, Chikumi H, Takata M, Nakamoto M, Igishi T, Shimizu ERapamycin induces p53-independent apoptosis through the mitochondrial pathway in non-small cell lung cancer cells. Oncol Rep 28: 848-854
Division of Medical Oncology and Molecular Respirology, Department of Multidisciplinary Internal Medicine, Tottori University, 36-1 Nishi-cho, Yonago-shi, Tottori-ken 683-8504, Japan. Oncology Reports
(Impact Factor: 2.3).
06/2012; 28(3):848-54. DOI: 10.3892/or.2012.1855
The mammalian target of rapamycin (mTOR) is a key kinase acting downstream of growth factor receptor PI3K and AKT signaling, leading to processes resulting in increased cell size and proliferation through translation control. Rapamycin, a specific inhibitor of mTOR, results predominately in G1 cell cycle arrest through translation control and occasionally, cell type-dependent apoptosis by an unknown mechanism. In this study, we investigated the effect and mechanism of action of rapamycin on non-small cell lung cancer (NSCLC) cell lines with p53 mutations. Cell proliferation was evaluated by modified MTT assay. The apoptotic effect of rapamycin was measured by caspase-3 activation and flow cytometric analysis of Annexin V binding. The expression of Bcl-2 and the release of cytochrome c from mitochondria were evaluated by western blotting. We found that rapamycin induced apoptosis in NSCLC cell lines with p53 mutations. Western blot analysis demonstrated that rapamycin downregulates the expression levels of Bcl-2, which leads to increased cytochrome c release from mitochondria and subsequent activation of caspase cascades. These findings suggest that rapamycin induces p53-independent apoptosis through downregulation of Bcl-2 and the mitochondrial pathway in NSCLC cell lines as a novel antitumor mechanism.
Available from: Cinzia Antognelli
- "It is broadly accepted that p53 is a master tumour suppressor protein promoting apoptosis in many cellular systems. However, it has also been reported that caspase-mediated apoptosis does not always require p53 (Miyake et al, 2012; Yeh et al, 2012; Amir et al, 2013). Here, we showed that IR-induced apoptosis, via GI inhibition and AP-modified Hsp27 protein accumulation, occurred "
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Glyoxalase I (GI) is a cellular defence enzyme involved in the detoxification of methylglyoxal (MG), a cytotoxic byproduct of glycolysis, and MG-derived advanced glycation end products (AGEs). Argpyrimidine (AP), one of the major AGEs coming from MG modifications of proteins arginines, is a pro-apoptotic agent. Radiotherapy is an important modality widely used in cancer treatment. Exposure of cells to ionising radiation (IR) results in a number of complex biological responses, including apoptosis. The present study was aimed at investigating whether, and through which mechanism, GI was involved in IR-induced apoptosis.
Apoptosis, by TUNEL assay, transcript and protein levels or enzymatic activity, by RT–PCR, western blot and spectrophotometric methods, respectively, were evaluated in irradiated MCF-7 breast cancer cells, also in experiments with appropriate inhibitors or using small interfering RNA.
Ionising radiation induced a dramatic reactive oxygen species (ROS)-mediated inhibition of GI, leading to AP-modified Hsp27 protein accumulation that, in a mechanism involving p53 and NF-κB, triggered an apoptotic mitochondrial pathway. Inhibition of GI occurred at both functional and transcriptional levels, the latter occurring via ERK1/2 MAPK and ERα modulation.
Glyoxalase I is involved in the IR-induced MCF-7 cell mitochondrial apoptotic pathway via a novel mechanism involving Hsp27, p53 and NF-κB.
Available from: sciencedirect.com
- "Rapamycin, a mTOR inhibitor, inhibits cell proliferation and induces apoptosis by decreasing the phosphorylation of mTOR and its downstream targets, including 70 kDa ribosomal S6 kinase (p70S6K) and transcription initiation factor 4E-binding protein 1 (4E-BP1) (Karlsson et al., 2013). Rapamycin and its derivates have been identified as the therapeutic drugs for non-small cell lung cancer (Miyake et al., 2012; Huang et al., 2013), but the immunosuppression effect limited its application as well. "
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Cardamonin has previously demonstrated that it had the antiproliferation of vascular smooth muscle cells by inhibiting the activity of mammalian target of rapamycin (mTOR). The antiproliferative effect and the possible mechanism of combining with mTOR of cardamonin were investigated on A549 cells.
The cell proliferation, cell cycle and apoptosis were measured by methyl thiazolyl tetrazolium (MTT) and flow cytometry, respectively. mTOR and 12 kDa FK506 bind protein (FKBP12) were transfected into A549 cells by Lipofectamine. Western blots were used to examine the mTOR expressions and its activities, and the expressions of 70 kDa ribosomal S6 kinase (p70S6K), FKBP12 and Interleukin-2 (IL-2), respectively.
Treated with cardamonin, the proliferation of A549 cells was inhibited. Meanwhile, the cell cycle was blocked and DNA synthesis was decreased whereas the cell apoptosis was promoted, and the activation of mTOR and p70S6K was decreased by cardamonin. Transfected with mTOR or FKBP12, proliferation of A549 cells was increased. Rapamycin had the degree of similarity effect of antiproliferation on both transfected cells. However, the antiproliferative effect of cardamonin on mTOR transfected cells was stronger than that on FKBP12 transfected cells. Both rapamycin and cardamonin decreased the phosphorylation of mTOR and p70S6K in two kinds of transfected cells. Cardamonin had no effect on the expression of FKBP12 and IL-2, whereas the expressions were decreased by rapamycin.
Cardamonin inhibited proliferation and induced apoptosis of A549 cells via mTOR. It might directly interact with mTOR independent of binding with FKBP12.
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We investigated the effects of bevacizumab and rapamycin on central corneal opacity and apoptotic keratocyte number after photorefractive keratectomy (PRK) followed by ultraviolet B (UV-B) irradiation.
A total of 60 right eyes of Sprague-Dawley rats in four groups (n = 15 each) underwent PRK ablation to 80 μm with a 3-mm zone. Sponges soaked with 0.02% mitomycin C (MMC), 2.5% bevacizumab, 0.01% rapamycin, and balanced saline solution were applied for 2 minutes to these eyes in the MMC, bevacizumab, rapamycin, and control groups, respectively. At 3 weeks after PRK, all right eyes were exposed to 100 mJ/cm(2) UV-B irradiation. Biomicroscopy was used to determine the amount of haze, and TUNEL staining for apoptosis and histology were performed at 3, 6, and 12 weeks.
Contrary to the results at 3 weeks, central corneal haze, and apoptotic keratocyte and keratocyte number decreased significantly in the MMC, bevacizumab, and rapamycin groups compared to the control group, and the keratocyte number was lower in the MMC group than the bevacizumab and rapamycin groups at 6 weeks (all P < 0.05). At 12 weeks, the apoptotic keratocyte number was lower in the MMC, bevacizumab, and rapamycin groups than the control group, and the keratocyte number was significantly lower in the MMC than the rapamycin and control groups (all P < 0.05).
Intraoperative bevacizumab and rapamycin administration decreases central corneal haze and apoptotic keratocyte number after PRK. Bevacizumab and rapamycin may be safe alternatives to MMC during refractive surgery to prevent postoperative corneal opacity less affecting the keratocyte number.
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