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Luciferase assay was performed on (a) MCF-7 and (b) MDA-MB-231 human breast cancer cells. The cells were initially plated at a density of 2×104 cells/well in 96-well plates. Cells were then transfected by adding 32 ng of PPRE X3-TK-luc and 3.2 ng of renilla luciferase plasmid in 0.8 μL of lipofectamine 2000 transfection reagent. Following a 6-h incubation period, MCF-7 cells were treated with control or treatment media containing 0–2 μM γ-tocotrienol, 0–3.2 μM rosiglitazone, 0–3.2 μM troglitazone, 0–3.2 μM GW9662, or 0–6.4 μM T0070907 alone or in combination. MDA-MB-231 cells were initially plated in a similar manner and treated with control or treatment media containing 0–3 μM γ-tocotrienol, 0–6.4 μM rosiglitazone, 0–6.4 μM troglitazone, 0–6.4 μM GW9662, or 0–6.4 μM T0070907 alone or in combination. All cells were fed fresh treatment media every other day for 4-day incubation period. Afterwards, cells were lysed with 75 μL of passive lysis buffer and treated according to manufacturer’s instructions using the dual-glo luciferase assay system. Results were calculated as raw luciferase units divided by raw renilla units. Vertical bars indicate PPRE mediated reporter activity ± SEM (arbitrary units) in each treatment group. P*<0.05 as compared with vehicle-treated controls.
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γ-Tocotrienol is a natural vitamin E that displays potent anticancer activity, and previous studies suggest that these effects involve alterations in PPARγ activity. Treatment with 0.5-6 μM γ-tocotrienol, 0.4-50 μM PPARγ agonists (rosiglitazone or troglitazone), or 0.4-25 μM PPARγ antagonists (GW9662 or T0070907) alone resulted in a dose-responsive...
Citations
... Moreover, due to the crucial role of PARP-1 in the DNA repair system, the inhibition of PARP-1 leads to cell apoptosis as a result of damage to DNA, which creates a new strategy in the development of cancer drugs [70]. It has been described that both rosiglitazone and GW9662 do not significantly change PARP expression [71,72], which is similar to our observation for the A549 cells. On the other hand, Girnun et al. (2007) showed that rosiglitazone and carboplatin (cytostatic drug) cleaved PARP-1, while 0.5 µM rosiglitazone alone did not change PARP-1 expression and activation in A549 cells [73]. ...
Cancer is one of the most important problems of modern societies. Recently, studies have reported the anticancer properties of rosiglitazone related to its ability to bind peroxisome proliferator receptor γ (PPARγ), which has various effects on cancer and can inhibit cell proliferation. In this study, we investigated the effect of new 4-thiazolidinone (4-TZD) hybrids Les-4369 and Les-3467 and their effect on reactive oxygen species (ROS) production, metabolic activity, lactate dehydrogenase (LDH) release, caspase-3 activity, and gene and protein expression in human foreskin fibroblast (BJ) cells and lung adenocarcinoma (A549) cells. The ROS production and caspase-3 activity were mainly increased in the micromolar concentrations of the studied compounds in both cell lines. Les-3467 and Les-4369 increased the mRNA expression of PPARG, P53 (tumor protein P53), and ATM (ATM serine/threonine kinase) in the BJ cells, while the mRNA expression of these genes (except PPARG) was mainly decreased in the A549 cells treated with both of the tested compounds. Our results indicate a decrease in the protein expression of AhR, PPARγ, and PARP-1 in the BJ cells exposed to 1 µM Les-3467 and Les-4369. In the A549 cells, the protein expression of AhR, PPARγ, and PARP-1 increased in the treatment with 1 µM Les-3467 and Les-4369. We have also shown the PPARγ modulatory properties of Les-3467 and Les-4369. However, both compounds prove weak anticancer properties evidenced by their action at high concentrations and non-selective effects against BJ and A549 cells.
... For instance, its coupling to tyrosine kinase inhibitors such as gefitinib or erlotinib, resulted in the inhibition of epidermal growth factordependent mitogenic signaling in murine mammary tumor cells [15]. Similarly, Malaviya and Sylvester, showed, that a combined treatment involving γ-tocotrienol with PPARγγ antagonists such as GW9662 or T0070907, synergistically inhibited the cell growth of breast cancer cell lines, MCF-7 and MDA-MB-231 [16]. Those studies indicate that γ-tocotrienol alone, or in combination to other chemotherapeutic drugs, exert anti-cancerous effects on various cancer types. ...
Acute myeloid leukemia (AML) is a blood cancer characterized by the formation of faulty defective myelogenous cells with morphological heterogeneity and cytogenic aberrations leading to a loss of their function. In an attempt to find an effective and safe AML treatment, vitamin E derivatives, including tocopherols were considered as potential anti-tumor compounds. Recently, other isoforms of vitamin E, namely tocotrienols have been proposed as potential potent anti-cancerous agents, displaying promising therapeutic effects in different cancer types. In this study we evaluated the anti-cancerous effects of γ-tocotrienol, on AML cell lines in vitro. For this purpose, AML cell lines incubated with γ-tocotrienol were examined for their viability, cell cycle status, apoptotic cell death, DNA fragmentation, production of reactive oxygen species and expression of proapoptotic proteins. Our results showed that γ-tocotrienol exhibits time and dose-dependent anti-proliferative, pro-apoptotic and antioxidant effects on U937 and KG-1 cell lines, through the upregulation of proteins involved in the intrinsic apoptotic pathway.
... For instance, its coupling to tyrosine kinase inhibitors such as gefitinib or erlotinib, resulted in the inhibition of epidermal growth factordependent mitogenic signaling in murine mammary tumor cells [15]. Similarly, Malaviya and Sylvester, showed, that a combined treatment involving γ-tocotrienol with PPARγγ antagonists such as GW9662 or T0070907, synergistically inhibited the cell growth of breast cancer cell lines, MCF-7 and MDA-MB-231 [16]. Those studies indicate that γ-tocotrienol alone, or in combination to other chemotherapeutic drugs, exert anti-cancerous effects on various cancer types. ...
Acute myeloid leukemia (AML) is a blood cancer characterized by the formation of faulty defective myelogenous cells with morphological heterogeneity and cytogenic aberrations leading to a loss of their function. In an attempt to find an effective and safe AML treatment, vitamin E derivatives, including tocopherols were considered as potential anti-tumor compounds. Recently, other isoforms of vitamin E, namely tocotrienols have been proposed as potential potent anti-cancerous agents, displaying promising therapeutic effects in different cancer types. In this study we evaluated the anti-cancerous effects of γ-tocotrienol, on AML cell lines in vitro. For this purpose, AML cell lines incubated with γ-tocotrienol were examined for their viability, cell cycle status, apoptotic cell death, DNA fragmentation, production of reactive oxygen species and expression of proapoptotic proteins. Our results showed that γ-tocotrienol exhibits time and dose-dependent anti-proliferative, pro-apoptotic and antioxidant effects on U937 and KG-1 cell lines, through the upregulation of proteins involved in the intrinsic apoptotic pathway.
... For instance, its coupling to tyrosine kinase inhibitors such as gefitinib or erlotinib, resulted in the inhibition of epidermal growth factordependent mitogenic signaling in murine mammary tumor cells [15]. Similarly, Malaviya and Sylvester, showed, that a combined treatment involving γ-tocotrienol with PPARγγ antagonists such as GW9662 or T0070907, synergistically inhibited the cell growth of breast cancer cell lines, MCF-7 and MDA-MB-231 [16]. Those studies indicate that γ-tocotrienol alone, or in combination to other chemotherapeutic drugs, exert anti-cancerous effects on various cancer types. ...
Acute myeloid leukemia (AML) is a blood cancer characterized by the formation of faulty defective myelogenous cells with morphological heterogeneity and cytogenic aberrations leading to a loss of their function. In an attempt to find an effective and safe AML treatment, vitamin E derivatives, including tocopherols were considered as potential anti-tumor compounds. Recently, other isoforms of vitamin E, namely tocotrienols have been proposed as potential potent anti-cancerous agents, displaying promising therapeutic effects in different cancer types. In this study we evaluated the anti-cancerous effects of γ-tocotrienol, on AML cell lines in vitro. For this purpose, AML cell lines incubated with γ-tocotrienol were examined for their viability, cell cycle status, apoptotic cell death, DNA fragmentation, production of reactive oxygen species and expression of proapoptotic proteins. Our results showed that γ-tocotrienol exhibits time and dose-dependent anti-proliferative, pro-apoptotic and antioxidant effects on U937 and KG-1 cell lines, through the upregulation of proteins involved in the intrinsic apoptotic pathway.
... Subsequent studies showed that tocotrienols are able to inhibit a wide range of receptors such as VEGFR [67], HER2-4 [24,68,69], MET [70,71], and Frizzled-7 (FZD7) [72]. Furthermore, combined treatment of tocotrienols with other phytochemicals and chemotherapeutic agents shows synergistic anticancer activity [24,25,70,[73][74][75]. In addition to suppressing mitogen-dependent mitogenesis, treatment with very high doses of γ-tocotrienol is cytotoxic and initiates apoptosis, necrosis, and autophagy in cancer cells [15,76,77]. ...
γ-Tocotrienol, a natural isoform within the vitamin E family of compounds, displays potent antiproliferative and apoptotic activity against a wide range of cancers, particularly breast cancer. A large percentage of human breast cancers characteristically display aberrant receptor tyrosine kinase activity, including receptors within the HER family, c-Met, and VEGF among others. Lipid rafts are specialized microdomains within the plasma membrane that are required for receptor tyrosine kinase dimerization, activation, and signal transduction. Recent studies demonstrate that the anticancer effects of γ-tocotrienol are associated with its accumulation in the lipid raft microdomain where it appears to interfere with the receptor tyrosine kinase dimerization and activation in human breast cancer cells. Heregulin is a potent ligand that activates HER3 and HER4 receptors, and overexpression of this ligand is associated with the development of chemotherapy resistance. Exosomes released from cancer cells are small vesicles originating from the outward budding of lipid rafts that carry various mitogenic proteins that then act locally in an autocrine/paracrine manner to stimulate cancer cell growth. However, exosomes isolated from the culture media of breast cancer cells treated with γ-tocotrienol contain significantly less heregulin content and are significantly less potent in stimulating HER3/HER4 heterodimerization, activation, and mitogenic signaling, as compared to exosomes isolated from the culture media of breast cancer cells grown in control media. These findings provide strong evidence that the anticancer effects of γ-tocotrienol result, at least in part, by directly disrupting lipid raft integrity by directly interfering with HER receptor dimerization and signaling within the lipid rafts and indirectly by reducing exosome heregulin content and subsequent autocrine/paracrine mitogenic stimulation.
... This cell line also expresses wild-type p53, PPARγ and NR4A1 [12]. Of the breast cancer lines, MCF-7 expresses estrogen receptors and its growth is estrogen-dependent [15,18], whereas MDA-MB-231 does not express estrogen receptors and its growth is estrogen-independent [18]. Wild-type tumor suppressor p53 and mutant nonfunctional p53 are expressed in MCF-7 and MDA-MB-231 cells, respectively [26]. ...
... This cell line also expresses wild-type p53, PPARγ and NR4A1 [12]. Of the breast cancer lines, MCF-7 expresses estrogen receptors and its growth is estrogen-dependent [15,18], whereas MDA-MB-231 does not express estrogen receptors and its growth is estrogen-independent [18]. Wild-type tumor suppressor p53 and mutant nonfunctional p53 are expressed in MCF-7 and MDA-MB-231 cells, respectively [26]. ...
... Wild-type tumor suppressor p53 and mutant nonfunctional p53 are expressed in MCF-7 and MDA-MB-231 cells, respectively [26]. NR4A1 is expressed in MCF-7 cells [1], but not in MDA-MB-231 cells [1], and PPARγ is expressed in both lines [17,18]. Of the prostate cancer cell lines, LNCaP cells are androgen-dependent and express the mutant androgen receptor (AR) (Thr877Ala), which is activated by both the androgen dihydrotestosterone (DHT) and the anti-androgen hydroxyflutamide [25]. ...
... In growth studies, cells were exposed to their respective experimental treatments containing various doses of γ-tocotrienol for a 3-day culture period. Afterwards, viable cell number was determined using the 3-(4, 5dimethylthiazol-2yl)-2,5-diphenyl tetrazolium bromide (MTT) colorimetric assay as described previously [27]. Briefly, the media in all treatment groups was replaced with fresh control media containing 0.5 mg/ml MTT. ...
... T47D cells were isolated with trypsin, and then whole cell lysates were prepared as previously described [27]. Proteins concentrations from the cells and the exosomes were determined using the Bio-Rad protein assay kit (Bio-Rad, Hercules, CA, USA). ...
Overexpression of heregulin, a potent ligand that activates HER3 and HER4 receptors, plays a significant role in the development of chemotherapy resistance in breast cancer patients. Exosomes released from cancer cells are small vesicles originating from the outward budding of lipid rafts that carry various mitogenic proteins that then act locally in an autocrine/paracrine manner to stimulate cancer cell growth. Since the anticancer activity of γ-tocotrienol has been shown to be mediated in part through the disruption of lipid rafts, studies were conducted to determine the effect of γ-tocotrienol on exosomes mitogenic biopotency. Exosomes isolated from the media of cultured T47D breast cancer cells were found to stimulate T47D cell growth in a dose-dependent manner. These growth stimulating effects were due to the high levels of heregulin contained in the exosomes which that acts to stimulate of HER3 and HER4 activation, heterodimerization and mitogenic signaling. Exposure to 5 μM γ-tocotrienol resulted in the selective accumulation and disruption in the integrity of the lipid raft microdomain and a corresponding decrease in exosome heregulin content and mitogenic biopotency. These findings provide strong evidence indicating that the anticancer effects of γ-tocotrienol are mediated, at least in part, by directly disrupting HER dimerization and signaling within the lipid rafts and indirectly by reducing exosome heregulin content and subsequent autocrine/paracrine mitogenic stimulation.
... 25) Specifically, studies have shown that breast cancer cells exposed to growth inhibiting doses of γ-tocotrienol display significant inhibition in HER2/ErbB2, HER3/ErbB3, HER4/ErbB4, and c-Met activation, [25][26][27]37,[40][41][42] and their downstream effectors including mitogen-activated protein kinase (MAPK), phosphatidylinositol 3-kinase (PI3K)/Akt, Stat, and nuclear factorkappa B (NFκB). 35,40,[43][44][45] Furthermore, γ-tocotrienol suppres-sion of c-Met receptor signaling was directly associated with a suppression in EMT, as evidence by an increased expression of epithelial markers such as E-cadherin, β-catenin, and cytokeratins 8/18 and corresponding decrease in the expression of vimentin, a mesenchymal marker. 26,37) It is now well established that Rac1/WAVE2 signaling plays an important role in promoting EMT and metastasis in a variety of cancer cell types. ...
The majority of breast cancer deaths result from the progression of this disease to a metastatic phenotype. Rac1 and Cdc42 are Rho family members that together with their downstream effectors, Wiskott-Aldrich Syndrome protein-family verprolin-homologous protein 2 (WAVE2) and Arp2/3, play an important role in cytoskeletal reorganization and the formation of membrane protrusions that promote cancer cell migration and invasion. γ-Tocotrienol, is a natural isoform within the vitamin E family of compounds that inhibits breast cancer cell growth and progression by suppressing various signaling pathways involved in mitogenic signaling and metastatic progression. Studies were conducted to examine the effects of γ-tocotrienol on Rac1/WAVE2 signaling dependent migration and invasion in highly metastatic mouse +SA and human MDA-MB-231 mammary cancer cells. Exposure to γ-tocotrienol resulted in a dose-responsive decrease in Rac1/WAVE2 signaling as characterized by a suppression in the levels of Rac1/Cdc42, phospho-Rac1/Cdc42, WAVE2, Arp2, and Arp3 expression. Additional studies also demonstrated that similar treatment with γ-tocotrienol resulted in a significant reduction in tumor cell migration and invasion. Taken together, these findings indicate that γ-tocotrienol treatment effectively inhibits Rac1/WAVE2 signaling and reduces metastatic phenotypic expression in mammary cancer cells, suggesting that γ-tocotrienol may provide some benefit as a novel therapeutic approach in the treatment of metastatic breast cancer.
... Considering this, several studies are in progress to screen for molecules that can synergize with T3 in order to augment its potency. Combinations of T3 and certain drugs, such as statins [22], erlotinib/gefitinib [23], celecoxib [24], SU11274 [25], GW966/T0070907 [26], oridonin [27], and baicalein [28], have synergistic actions on cancer cell growth and signaling pathways. In addition, co-treatment with T3 and dietary components, including epigallocatechin gallate (EGCG)/resveratrol [29], sesamin [30], and ferulic acid [31], also exhibit synergistic effects. ...
... These results suggested that T3 inhibited prostate cancer cell growth through, in part, PPARγ-dependent mechanisms. Malaviya and Sylvester examined the anti-proliferative effect of γ-T3 in combination with PPARγ agonists or antagonists on breast cancer cells (MCF-7 and MDA-MB-231) [26]. Single treatment with γ-T3, PPARγ agonists (rosiglitazone or troglitazone) or antagonists (GW9662 or T0070907) resulted in a dose-dependent growth inhibition of breast cancer cells. ...
... Unexpectedly, a combination with γ-T3 and the agonists promoted cancer cell proliferation, whereas co-treatment of γ-T3 and the antagonists synergistically suppressed the cell growth. The contradictory results might be derived from the cancer cell types used in this experiment; the authors [26] employed breast cancer cells while Campbell et al. [79] used prostate cancer cells. They concluded that the synergistic effect between γ-T3 and PPARγ antagonists was mediated through PPARγ-independent mechanisms [80]. ...
Tocotrienol (T3), unsaturated vitamin E, is gaining a lot of attention owing to its potent anticancer effect, since its efficacy is much greater than that of tocopherol (Toc). Various factors are known to be involved in such antitumor action, including cell cycle arrest, apoptosis induction, antiangiogenesis, anti-metastasis, nuclear factor-?B suppression, and telomerase inhibition. Owing to a difference in the affinity of T3 and Toc for the ?-tocopherol transfer protein, the bioavailability of orally ingested T3 is lower than that of Toc. Furthermore, cellular uptake of T3 is interrupted by coadministration of ?-Toc in vitro and in vivo. Based on this, several studies are in progress to screen for molecules that can synergize with T3 in order to augment its potency. Combinations of T3 with chemotherapeutic drugs (e.g., statins, celecoxib, and gefitinib) or dietary components (e.g., polyphenols, sesamin, and ferulic acid) exhibit synergistic actions on cancer cell growth and signaling pathways. In this review, we summarize the current status of synergistic effects of T3 and an array of agents on cancer cells, and discuss their molecular mechanisms of action. These combination strategies would encourage further investigation and application in cancer prevention and therapy.
... For instance, different studies have recently reported that tocotrienols at 40 mM have cytotoxic effects in human breast cancer cells MCF-7 and MDA-MB-231, which is associated with autophagy induction and ER stress-mediated apoptosis Tiwari et al., 2015). Furthermore, tocotrienols have been successfully used as anticancer agents in combination therapies (Abdul Rahman et al., 2014;Kani et al., 2013;Malaviya and Sylvester, 2013;Rezaei et al., 2014;Sylvester et al., 2011;Tiwari et al., 2015;Tuerdi et al., 2013;Yusof et al., 2015), as agents against cardiovascular diseases (Prasad, 2011) and as radioprotectors in patients undergoing radiotherapy (Kulkarni et al., 2010;Singh et al., 2013). ...
Dihydroceramide desaturase (Des1) is the last enzyme in the de novo synthesis of ceramides (Cer). It catalyzes the insertion of a double bond into dihydroceramides (dhCer) to convert them to Cer, both of which are further metabolized to more complex (dihydro)sphingolipids. For many years dhCer have received poor attention, mainly due to their supposed lack of biological activity. It was not until about ten years ago that the concept that dhCer might have regulatory roles in biology emerged for the first time. Since then, multiple publications have established that dhCer are implicated in a wide spectrum of biological processes. Physiological and pathophysiological functions of dhCer have been recently reviewed. In this review we will focus on the biochemical features of Des1 and on its inhibition by different compounds with presumably different modes of action.
Copyright © 2015. Published by Elsevier Ireland Ltd.
