Human Studies/ Cohort Studies
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| Article | Study objective/ findings |
| Effectiveness of tocotrienol-rich fraction combined with tamoxifen in the management of women with early breast cancer: a pilot clinical trial.
Nesaretnam, K., et.al (2010). Breast Cancer Res.
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A double-blinded, placebo-controlled pilot trial to test the effectiveness of adjuvant tocotrienol therapy in combination with tamoxifen for five years in women with early breast cancer. Two-hundred-forty women, aged between 40-60 years, with either tumor node metastases (TNM) Stage I or II breast cancer and estrogen receptor (ER) positive tumors were non-randomly assigned to two groups. The intervention group received tocotrienol rich fraction (TRF) plus tamoxifen whilst the control group received placebo plus tamoxifen, for five years. From the current study, there seems to be no association between adjuvant tocotrienol therapy and breast cancer specific survival in women with early breast cancer.
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| Tocotrienol level in adipose tissue of beningn and malignant breast lumps in patients in Malaysia.
Nesaretnam, K, et.al (2007). Asia Pac J Clin Nutr. |
To investigate whether there was a difference in tocopherol and tocotrienol concentrations in malignant and benign adipose tissue, in a Malaysian population consuming predominantly a palm oil diet. The study was undertaken using fatty acid levels in breast adipose tissue as a biomarker of qualitative dietary intake of fatty acids. The major fatty acids in breast adipose tissue of patients (benign and malignant) were oleic acid (45-46%), palmitic (28-29%) and linoleic (11-12%). No differences were evident in the fatty acid composition of the two groups. There was a significant difference (p=0.006) in the total tocotrienol levels between malignant (13.7 +/- 6.0 microg/g) and benign (20+/-6.0 microg/g) adipose tissue samples. However, no significant differences were seen in the total tocopherol levels (p=0.42) in the two groups. The study reveals that dietary intake influences adipose tissue fatty acid levels and that adipose tissue is a dynamic reservoir of fat soluble nutrients. The higher adipose tissue concentrations of tocotrienols in benign patients provide support for the idea that tocotrienols may provide protection against breast cancer. |
In- vivo / Animal Studies |
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| Article | Study objectives/ findings |
| Tocotrienol-adjuvanted dendritic cells inhibit tumor growth and metastasis: a murine model of breast cancer.
Abdul Hafid, S.R., et.al (2013). PLoS One.
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Tocotrienol-rich fraction (TRF) from palm oil is reported to possess anti-cancer and immune-enhancing effects. In this study, TRF supplementation was used as an adjuvant to enhance the anti-cancer effects of dendritic cells (DC)-based cancer vaccine in a syngeneic mouse model of breast cancer. Female BALB/c mice were inoculated with 4T1 cells in mammary pad to induce tumor. When the tumor was palpable, the mice in the experimental groups were injected subcutaneously with DC-pulsed with tumor lysate (TL) from 4T1 cells (DC+TL) once a week for three weeks and fed daily with 1 mg TRF or vehicle. Control mice received unpulsed DC and were fed with vehicle. The combined therapy of using DC+TL injections and TRF supplementation (DC+TL+TRF) inhibited (p<0.05) tumor growth and metastasis. Splenocytes from the DC+TL+TRF group cultured with mitomycin-C (MMC)-treated 4T1 cells produced higher (p<0.05) levels of IFN-γ and IL-12. The cytotoxic T-lymphocyte (CTL) assay also showed enhanced tumor-specific killing (p<0.05) by CD8(+) T-lymphocytes isolated from mice in the DC+TL+TRF group. This study shows that TRF has the potential to be used as an adjuvant to enhance effectiveness of DC-based vaccines.
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| Effect of annatto-tocotrienols supplementation on the development of mammary tumors in HER-2/neu transgenic mice.
Pierpaoli, E., et.al (2013). Carcinogenesis.
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The effects of dietary supplementation with annatto-T3 (90% δ-T3 and 10% γ-T3) on the spontaneous development of mammary tumors in HER-2/neu transgenic mice. Underlying mechanisms of the antitumor effect were evaluated by studying apoptosis, senescent-like growth arrest, immune modulation, oxidative effect and the expression of HER-2/neu in tumoral mammary glands of transgenic mice and in vitro in human and mice tumor cell lines. The results show that the antitumor effect of annatto-T3 supplementation in HER-2/neu transgenic mice is mainly related to the direct induction of oxidative stress, senescent-like growth arrest and apoptosis of tumor cells rather than to an immune modulation.
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| Tocotrienol-rich fraction from palm oil affects gene expression in tumors resulting from MCF-7 cell inoculation in athymic mice.
Nesaretnam, K., et.al (2004). Lipids.
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To investigate the molecular basis of the effect of tocotrienols, we injected MCF-7 breast cancer cells into athymic nude mice. Mice were fed orally with 1 mg/d of tocotrienol-rich fraction (TRF) for 20 wk. The observations indicate that TRF supplementation significantly and specifically affects MCF-7 cell response after tumor formation in vivo and therefore the host immune function. The observed effect on gene expression is possibly exerted independently from the antioxidant activity typical of this family of molecules. |
In-vitro Studies |
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| Article | Study objective/ findings |
| γ-Tocotrienol-induced disruption of lipid rafts in human breast cancer cells is associated with a reduction in exosome heregulin content
Alawin, O.A., et.al (2017). J Nutr Biochem. |
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 that act to stimulate 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.
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| Tocotrienol Nanoemulsion Platform of Curcumin Elicit Elevated Apoptosis and Augmentation of Anticancer Efficacy against Breast and Ovarian Carcinomas.
Steuber, N, et.al (2016). Int J Mol Sci.
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Vitamin E (VE) tocotrienols (T3), recognized for their cancer-specific anti-proliferative and pro-apoptotic activities, have been previously fabricated into bio-active nanoemulsion (NE) formulations. Here, our viscosity-adapted δ-T3 NE platform was developed to additionally incorporate curcumin (CUR), which is known for its potent suppression of signaling pathways involved in malignant cell growth, survival and metastasis. Thanks to efficient 70:30 wt % surfactant mix of Lutrol F-127:VE-TPGS, in conjunction with optimal CUR loading, a prototype CUR in δ-T3 NE was successfully prepared. Model CUR/δ-T3 NE demonstrated excellent nano-scale aspects (mean particle size = 261 nm, PDI = 0.27, and ζ-potential = -35 mV), pharmaceutical stability, and controlled release properties. Suitability for systemic administration was also verified via standardized in vitro biocompatibility and hemocompatibility assays. In two human cancer cells (MCF-7 and OVCAR-8), our CUR/δ-T3 NE prominently suppressed constitutive NF-κB activation, and significantly induced apoptosis. Finally, the combined CUR/δ-T3 NE produced superior cytotoxicity profiles, in concentration- and time-dependent manners (p ≤ 0.05), at least three to four folds lower IC50 than in closest CUR control. The strong synergism, estimated in both cultured carcinomas, revealed the augmented therapeutic efficacy of our CUR/δ-T3 NE combined platform, supporting its strong potential towards pharmaceutical development for cancer therapy.
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| Gamma-tocotrienol reverses multidrug resistance of breast cancer cells with a mechanism distinct from that of atorvastatin.
Ding, Y., et.al (2017). J Steroid Biochem Mol Biol.
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The role of the mevalonate pathway in γ-tocotrienol-mediated reversal of multidrug resistance in cancer cells was determined. It was found that both γ-tocotrienol and atorvastatin effectively reversed multidrug resistance of MCF-7/Adr and markedly inhibited the intracellular levels of FPP and GGPP. Exogenous addition of mevalonate or FPP and GGPP almost completely prevented the reversal ability of atorvastatin but only partly attenuated the reversal effect of γ-tocotrienol on doxorubicin resistance. In addition, γ-tocotrienol actively inhibited the expression of P-gp and increased the accumulation of doxorubicin in cells, which led to the enhanced G2/M arrest and cell apoptosis. Taken together, γ-tocotrienol reversed the multidrug resistance of MCF-7/Adr with a mechanism distinct from that of atorvastatin. Instead of the mevalonate pathway, the inhibition of P-gp expression is a potential mechanism by which γ-tocotrienol reverses multidrug resistance in MCF-7/Adr.
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| Role of Rac1/WAVE2 Signaling in Mediating the Inhibitory Effects of γ-Tocotrienol on Mammary Cancer Cell Migration and Invasion.
Algayadh, I.G., et.al (2016). Biol Pharm Bull.
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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.
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| γ-Tocotrienol reversal of epithelial-to-mesenchymal transition in human breast cancer cells is associated with inhibition of canonical Wnt signalling.
Ahmed, R.A., et.al (2016). Cell Prolif.
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To investigate the role of the canonical Wnt pathway in mediating inhibitory effects of γ-tocotrienol on EMT in human breast cancer cells. γ-Tocotrienol suppression of metastatic breast cancer cell proliferation and EMT was associated with suppression of the canonical Wnt/β-catenin signaling pathway. |
| Elimination of ALDH+ breast tumor initiating cells by docosahexanoic acid and/or gamma tocotrienol through SHP-1 inhibition of Stat3 signaling.
Xiong, A., et.al (2016). Mol Carcinog.
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Study investigated the ability of docosahexaenoic acid (DHA) alone and in combination with gamma-tocotrienol (γT3) to eliminate aldehyde dehydrogenase positive (ALDH+) cells and to inhibit mammosphere formation, biomarker and functional assay for tumor initiating cells (TICs), respectively, in human triple negative breast cancer cells (TNBCs), and investigated possible mechanisms of action. DHA upregulated Src homology region 2 domain-containing protein tyrosine phosphatase-1 (SHP-1) protein levels and suppressed levels of phosphorylated signal transducer and activator of transcription-3 (pStat3) and its downstream mediators c-Myc, and cyclin D1. siRNA to SHP-1 enhanced the percentage of ALDH+ cells and Stat-3 signaling, as well as inhibited, in part, the ability of DHA to reduce the percentage of ALDH+ cells and Stat-3 signaling. γT3 alone and in combination with DHA reduced ALDH+ TNBCs, up-regulated SHP-1 protein levels, and suppressed Stat-3 signaling. Taken together, data demonstrate the anti-TIC potential of achievable concentrations of DHA alone as well as in combination with γT3.
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| Antiproliferative effects of γ-tocotrienol are associated with lipid raft disruption in HER2-positive human breast cancer cells.
Alawin, O.A., et.al (2016). J Nutr Biochem.
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The effects of γ-tocotrienol on HER2 activation within the lipid raft microdomain in HER2-positive SKBR3 and BT474 human breast cancer cells were examined. In summary, these findings demonstrate that the anticancer effects of γ-tocotrienol are associated with its accumulation in the lipid raft microdomain and subsequent interference with HER2 dimerization and activation in SKBR3 and BT474 human breast cancer cells. |
| miR-429 mediates delta-tocotrienol-induced apoptosis in triple-negative breast cancer cells by targeting XIAP.
Wang C., et.al (2015). Int J Clin Exp Med. |
Vitamin E δ-tocotrienol has been reported to possess anticancer activity both in vitro and in vivo. However, the underlying molecular mechanisms of δ-tocotrienol induced apoptosis in triple-negative breast cancer are not fully understood. Here, we reported that microRNA-429 (miR-429) is up-regulated in two TNBC cell lines (MDA-MB-231 and MDA-MB-468), treated with δ-tocotrienol. Inhibition of miR-429 may partially rescue the apoptosis induced by δ-tocotrienol in MDA-MB-231 cells. We also showed that the forced expression of miR-429 was sufficient to lead to apoptosis in MDA-MB-231 cells. Furthermore, we identified X-linked inhibitor of apoptosis protein (XIAP) as one of miR-429’s target genes. These results suggest that the activation of miR-429 by δ-tocotrienol may be an effective approach for the prevention and treatment of triple-negative breast cancer.
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| HPLC Separation of Vitamin E and Its Oxidation Products and Effects of Oxidized Tocotrienols on the Viability of MCF-7 Breast Cancer Cells in Vitro.
Drotleff, A.M., et.al (2015). J Agric Food Chem.
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Tocotrienols, a vitamin E subgroup, exert potent anticancer effects, but easily degrade due to oxidation. Eight vitamin E reference compounds, α-, β-, γ-, or δ-tocopherols or -tocotrienols, were thermally oxidized in n-hexane. The corresponding predominantly dimeric oxidation products were separated from the parent compounds by diol-modified normal-phase HPLC-UV and characterized by mass spectroscopy. The composition of test compounds, that is, α-tocotrienol, γ-tocotrienol, or palm tocotrienol-rich fraction (TRF), before and after thermal oxidation was determined by HPLC-DAD, and MCF-7 cells were treated with both nonoxidized and oxidized test compounds for 72 h. Whereas all nonoxidized test compounds (0-100 μM) led to dose-dependent decreases in cell viability, equimolar oxidized α-tocotrienol had a weaker effect, and oxidized TRF had no such effect. However, the IC50 value of oxidized γ-tocotrienol was lower (85 μM) than that of nonoxidized γ-tocotrienol (134 μM), thereby suggesting that γ-tocotrienol oxidation products are able to reduce tumor cell viability in vitro.
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| Anticancer Effects of γ-Tocotrienol Are Associated with a Suppression in Aerobic Glycolysis.
Parajuli, P., et.al (2015). Biol Pharm Bull.
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To determine the effects of γ-tocotrienol on aerobic glycolysis in mouse +SA and human MCF-7 breast cancer cells. Treatment with γ-tocotrienol resulted in a dose-responsive inhibition of both +SA and MCF-7 mammary tumor cell growth, and induced a relatively large reduction in glucose utilization, intracellular ATP production and extracellular lactate excretion. These effects were also associated with a large decrease in enzyme expression levels involved in regulating aerobic glycolysis, including hexokinase-II, phosphofructokinase, pyruvate kinase M2, and lactate dehydrogenase A. γ-Tocotrienol treatment was also associated with a corresponding reduction in the levels of phosphorylated (active) Akt, phosphorylated (active) mTOR, and c-Myc, but not HIF-1α or glucose transporter 1 (GLUT-1). In summary, these findings demonstrate that the antiproliferative effects of γ-tocotrienol are mediated, at least in the part, by the concurrent inhibition of Akt/mTOR signaling, c-Myc expression and aerobic glycolysis.
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| PEGylated γ-tocotrienol isomer of vitamin E: Synthesis, characterization, in vitro cytotoxicity, and oral bioavailability.
Abu-Fayyad., A., et.al (2015). Eur J Pharm Biopharm.
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In this study two PEGylated γ-T3 variants with mPEG molecular weights of 350 (γ-T3PGS 350) and 1000 (γ-T3PGS 1000) were synthesized by a two-step reaction procedure and characterized by (1)H NMR, HPLC, and mass spectroscopy. The physical properties of their self-assemblies in water were characterized by zeta, CMC, and size analysis. Similar physical properties were found between the PEGylated T3 and vitamin E TPGS. PEGylated T3 were also found to retain the in vitro cytotoxic activity of the free T3 against the MCF-7 and the triple-negative MDA-MB-231 breast cancer cells. PEGylated γ-T3 also increased the oral bioavailability of γ-T3 by threefolds when compared to the bioavailability of γ-T3 formulated into a self-emulsified drug delivery system. No significant differences in biological activity were found between the PEG 350 and 100 conjugates. Results from this study suggest that PEGylation of γ-T3 represents a viable platform for the oral and parenteral delivery of γ-T3 for potential use in the prevention of breast cancer.
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| Synergistic anticancer effects of combined γ-tocotrienol and oridonin treatment is associated with the induction of autophagy.
Tiwari, R., et.al (2015). Mol Cell Biochem.
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γ-Tocotrienol and oridonin are natural phytochemicals that display potent anticancer activity. Studies showed that combined treatment with subeffective doses of γ-tocotrienol with oridonin resulted in synergistic autophagic and apoptotic effects in malignant +SA, but not normal CL-S1 mouse mammary epithelial cells in vitro. Specifically, combined treatment with low doses of γ-tocotrienol (8 µM) and oridonin (2 µM) for 24 h resulted in synergistic inhibition of +SA mammary cancer cells viability. This combination significantly enhanced the expression of autophagy cellular markers including the conversion of LC3B-I to LC3B-II, beclin-1, Atg3, Atg7, Atg5-Atg12, LAMP-1 and cathepsin-D, and pretreatment with the autophagy inhibitors 3-methyladenine (3-MA) or bafilomycin A1 (Baf1) blocked these effects. Furthermore, blockade of γ-tocotrienol and oridonin-induced autophagy with 3-MA or Baf1 induced a modest, but significant reduction in cytotoxicity resulting from the combined treatment of these phytochemicals. The anticancer effects of combination treatment was also associated with a large suppression in Akt/mTOR mitogenic signaling and corresponding increase in the levels of apoptotic cellular marker including cleaved caspase-3 and PARP, and Bax/Bcl-2 ratio in these tumor cells. These effects were also found to be selective against cancer cells, since similar combined treatment with γ-tocotrienol and oridonin did not induce autophagy or reduce viability of normal mouse CL-S1 mammary epithelial cells. These findings indicate that combined γ-tocotrienol and oridonin-induced autophagy plays a role in mediating the synergistic anticancer effects of these phytochemicals.
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| γ-Tocotrienol-induced endoplasmic reticulum stress and autophagy act concurrently to promote breast cancer cell death.
Tiwari, R.V., et.al (2015). Biochem Cell Biol.
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The anticancer effects of γ-tocotrienol are associated with the induction of autophagy and endoplasmic reticulum (ER) stress-mediated apoptosis, but a direct relationship between these events has not been established. Treatment with 40 μmol/L of γ-tocotrienol caused a time-dependent decrease in cancer cell viability that corresponds to a concurrent increase in autophagic and endoplasmic reticulum (ER) stress markers in MCF-7 and MDA-MB-231 human breast cancer cells. γ-Tocotrienol treatment was found to cause a time-dependent increase in early phase (Beclin-1, LC3B-II) and late phase (LAMP-1 and cathepsin-D) autophagy markers, and pretreatment with autophagy inhibitors Beclin-1 siRNA, 3-MA or Baf1 blocked these effects. Furthermore, blockage of γ-tocotrienol-induced autophagy with Beclin-1 siRNA, 3-MA, or Baf1 induced a modest, but significant, reduction in γ-tocotrienol-induced cytotoxicity. γ-Tocotrienol treatment was also found to cause a decrease in mitogenic Erk1/2 signaling, an increase in stress-dependent p38 and JNK1/2 signaling, as well as an increase in ER stress apoptotic markers, including phospho-PERK, phospho-eIF2α, Bip, IRE1α, ATF-4, CHOP, and TRB3. In summary, these finding demonstrate that γ-tocotrienol-induced ER stress and autophagy occur concurrently, and together act to promote human breast cancer cell death.
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| Synergistic inhibition of cancer cell proliferation with a combination of delta-tocotrienol and ferulic acid.
Eitsuka, T., et.al (2014). Biochem Biophys Res Commun. |
The synergistic inhibitory effects of its components, particularly δ-tocotrienol (δ-T3) and ferulic acid (FA), against the proliferation of an array of cancer cells, including DU-145 (prostate cancer), MCF-7 (breast cancer), and PANC-1 (pancreatic cancer) cells are investigated. The combination of δ-T3 and FA markedly reduced cell proliferation relative to δ-T3 alone, and FA had no effect when used alone. Although δ-T3 induced G1 arrest by up-regulating p21 in PANC-1 cells, more cells accumulated in G1 phase with the combination of δ-T3 and FA. This synergistic effect was attributed to an increase in the cellular concentration of δ-T3 by FA. The results suggest that the combination of δ-T3 and FA may present a new strategy for cancer prevention and therapy.
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| δ-Tocotrienol oxazine derivative antagonizes mammary tumor cell compensatory response to CoCl2-induced hypoxia
Ananthula, S., et.al (2014). Biomed Res Int. |
Studies were conducted to determine the role of δ-tocotrienol and a semisynthetic δ-tocotrienol oxazine derivative, compound 44, on +SA mammary tumor cell hypoxic response. These findings demonstrate that treatment with the δ-tocotrienol oxazine derivative, compound 44, significantly attenuates +SA mammary tumor cell compensatory responses to hypoxia and suggests that this compound may provide benefit in the treatment of rapidly growing solid breast tumors.
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| Autophagy inhibitor 3-methyladenine potentiates apoptosis induced by dietary tocotrienols in breast cancer cells.
Tran, A.T., et.al (2015). Eur J Nutr.
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Tocomin® represents commercially available mixture of naturally occurring tocotrienols (T3s) and tocopherols extracted from palm oil/palm fruits that possess powerful antioxidant, anticancer, neuro/cardioprotective and cholesterol-lowering properties. Cellular autophagy represents a defense mechanism against oxidative stress and several anticancer compounds. Recently, we reported that T3s induce apoptosis and endoplasmic reticulum stress in breast cancer cells. Together, the data indicate anticancer effects of Tocomin® in breast cancer cells, which is potentiated by the autophagy inhibitor 3-MA.
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| Entrapment into nanoemulsions potentiates the anticancer activity of tocotrienols against the highly malignant (+SA) mouse mammary epithelial cells.
Alayoubi, A., et.al (2014). J Nanosci Nanotechnol.
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The objective of this work was to test whether the activity of tocotrienols in lipid nanoemulsions against the +SA cells was retained. A secondary objective was to test whether stabilizing the nanoemulsions with poloxamer or sodium oleate would affect their activity. Nanoemulsions were found to be significantly more potent than tocotrienol/albumin conjugate. The IC50 values of the poloxamer and sodium oleate nanoemulsions were 3 and 6 microM, respectively, whereas the IC50 value of the conjugate was 10 microM. The antiproliferative activity of the nanoemulsions was also found to inversely correlate with particle size. No activity was observed with nanoemulsions loaded with alpha-tocopherol or vehicle, which confirmed the cytotoxic activity of tocotrienols and the potential use of nanoemulsions in cancer therapy.
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| Synergistic Antiproliferative Effects of Combined γ -Tocotrienol and PPAR γ Antagonist Treatment Are Mediated through PPAR γ -Independent Mechanisms in Breast Cancer Cells.
Malaviya, A., et.al (2014). PPAR Res.
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Studies were conducted to characterize the role of PPAR γ in mediating the effects of combined treatment of γ -tocotrienol with PPAR γ agonists or antagonists on the growth of PPAR γ negative +SA mammary cells and PPAR γ -positive and PPAR γ -silenced MCF-7 and MDA-MB-231 breast cancer cells. Combined treatment of γ -tocotrienol with PPAR γ antagonist decreased, while combined treatment of γ -tocotrienol with PPAR γ agonist increased, growth of all cancer cells. However, treatment with high doses of 15d-PGJ2, an endogenous natural ligand for PPAR γ , had no effect on cancer cell growth. Western blot and qRT-PCR studies showed that the growth inhibitory effects of combined γ -tocotrienol and PPAR γ antagonist treatment decreased cyclooxygenase (COX-2), prostaglandin synthase (PGDS), and prostaglandin D2 (PGD2) synthesis. In conclusion, the anticancer effects of combined γ -tocotrienol and PPAR γ antagonists treatment in PPAR γ negative/silenced breast cancer cells are mediated through PPAR γ -independent mechanisms that are associated with a downregulation in COX-2, PGDS, and PGD2 synthesis.
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| γ-Tocotrienol-induced autophagy in malignant mammary cancer cells.
Tiwari, R.V., et.al (2014). Exp Biol Med (Maywood).
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γ-Tocotrienol, a member of the vitamin E family of compounds, displays potent antiproliferative and cytotoxic effects in a variety of cancer cell types at treatment doses that have little or no effect on normal cell viability or growth. Autophagy is a tightly regulated lysosomal self-digested process that can either promote cell survival or programmed cell death, but the role of autophagy in mediating γ-tocotrienol-induced cytotoxicity in breast cancer is not presently completely understood. Mouse (+SA) and human (MCF-7 and MDA-MD-231) mammary tumor cells lines were exposed to 0-40 µmol/L γ-tocotrienol for a 24 h treatment period. γ-Tocotrienol treatment caused a relatively large increase in the accumulation of monodansylcadaverine (MDC)-labeled vacuoles, a marker of autophagosome formation, in all tumor cell lines. Results also showed that γ-tocotrienol treatment induced an increased conversion of microtubule-associated protein, 1A/1B-light chain 3, from its cytosolic form (LC3B-I) to its lipidated form (LC3B-II), increased Beclin-1 levels, and increased acridine orange staining as determined by flow cytometry analysis, providing further evidence of γ-tocotrienol-induced autophagy in these mammary cancer cell lines. In contrast, similar treatment with γ-tocotrienol was not found to increase autophagy marker expression in immortalized mouse (CL-S1) and human (MCF-10 A) normal mammary epithelial cell lines. Treatment with γ-tocotrienol also caused a reduction in PI3K/Akt/mTOR signaling and a corresponding increase in the Bax/Bcl-2 ratio, cleaved caspase-3, and cleaved poly (ADP-ribose) polymerase (PARP) levels in these cancer cell lines, suggesting that γ-tocotrienol-induced autophagy may be involved in the initiation of apoptosis. In summary, these findings demonstrate that the cytotoxic effects of γ-tocotrienol are associated with the induction of autophagy in a mouse and human mammary cancer cells.
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| Combined gamma-tocotrienol and Met inhibitor treatment suppresses mammary cancer cell proliferation, epithelial-to-mesenchymal transition and migration.
Ayoub, N.M., et.al (2013). Cell Prolif. |
Dysregulation of Met signalling is associated with malignant transformation. Combined treatment has been shown to reduce Met activation and mammary tumour cell proliferation. Experiments here, were conducted to determine mechanisms involved in mediating anti-cancer effects of combined γ-tocotrienol and SU11274 (Met inhibitor) treatment in various mammary cancer cell lines. It is suggest that combined γ-tocotrienol and Met inhibitor treatment may provide benefit in treatment of breast cancers characterized by aberrant Met activity.
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| Tocotrienols promote apoptosis in human breast cancer cells by inducing poly(ADP-ribose) polymerase cleavage and inhibiting nuclear factor kappa-B activity.
Loganathan, R., et.al (2013). Cell Prolif.
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The study described here has evaluated anti-cancer activity of vitamin E to elucidate mechanisms of cell death, using human breast cancer cells. Tocotrienols induced anti-proliferative and apoptotic effects in association with DNA fragmentation, poly(ADP-ribose) polymerase cleavage and NF-κB inhibition in the two human breast cancer cell lines. |
| Eliminating drug resistant breast cancer stem-like cells with combination of simvastatin and gamma-tocotrienol.
Gopalan, A., et.al (2013). Cancer Lett.
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Present study shows that drug resistant human breast cancer cells are enriched in cancer stem-like cells (CSCs) and express elevated levels of Stat-3 signaling mediators, which contribute to CSC enrichment. Simvastatin (SVA) and gamma-tocotrienol (γT3) eliminate enriched CSCs and suppress expression of Stat-3 signaling mediators via inhibition of the mevalonate pathway and activation of de novo ceramide synthesis pathway, respectively. Combination of SVA+γT3 at low doses enhanced these actions via inhibition of the mevalonate pathway. Data demonstrate that SVA and γT3 alone or in combination possess the ability to eliminate CSCs in drug resistant human breast cancer cells. |
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Viola, V., et.al (2013). Biofactors. |
Anticancer activity and mitochondrial mechanism of the vitamin E form δ-tocotrienol (δ-T3) was investigated in HER-2/neu-overexpressing human SKBR3 and murine TUBO breast cancer cells. In conclusion, synthetic derivatives of δ-T3 with enhanced apoptotic activity in breast carcinoma cells are investigated for the first time in this study also describing mechanistic aspects of mitochondrial effects of δ-T3. Further investigation in preclinical models of HER2/neu-high breast adenocarcinoma is underway to identify other and more effective forms of VE in this type of cancer.
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| Sesamin synergistically potentiates the anticancer effects of γ-tocotrienol in mammary cancer cell lines
Mohamed, R., et.al (2013). Fitorapia.
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Since sesamin inhibits the metabolic degradation of tocotrienols, studies were conducted to determine if combined treatment with sesamin potentiates the antiproliferative effects of γ-tocotrienol on neoplastic mouse (+ SA) and human (MCF-7 and MDA-MB-231) mammary cancer cells. These findings indicate that the synergistic antiproliferative action of combined γ-tocotrienol and sesamin treatment in mouse and human mammary cancer cells is cytostatic, not cytotoxic, and results from G1 cell cycle arrest.
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| Mechanisms mediating the synergistic anticancer effects of combined gamma-tocotrienol and sesamin treatment.
Akl, M.R., et.al (2012). Planta Med.
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In this study, the combined treatment of subeffective doses of γ-tocotrienol and sesamin caused a synergistic inhibition of murine +SA mammary epithelial cell growth, as determined by the MTT assay and immunofluorescent Ki-67 staining. The findings demonstrate that the synergistic growth inhibitory effects of γ-tocotrienol and sesamin treatment are associated with suppression of EGF-dependent mitogenic signaling in mammary tumor cells and suggest that dietary supplementation with these phytochemicals may provide some benefits in the prevention and/or treatment of breast cancer.
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| Involvement of de novo ceramide synthesis in gamma-tocopherol and gamma-tocotrienol-induced apoptosis in human breast cancer cells.
Gopalan, A., et.al (2012). Mol Nutr Food Res.
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This study further examines mechanisms involved in the pro-apoptotic action of gamma-tocopherol (γT) and gamma-tocotrienol (γT3) in human breast cancer cell lines. Data show that both γT and γT3 induce apoptosis via de novo ceramide synthesis dependent activation of JNK/CHOP/DR5 pro-apoptotic signaling. |
| Gamma-tocotrienol induced apoptosis is associated with unfolded protein response in human breast cancer cells.
Patacsil, D., et.al (2012). J Nutr Biochem.
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In the present study, γ-T3 induces apoptosis in MDA-MB 231 and MCF-7 breast cancer cells as evident by PARP cleavage and caspase-7 activation has been demonstrated. In summary, γ-T3 modulates ER stress signaling and have identified ATF3 as a molecular target for γ-T3 in breast cancer cells is demonstrated. |
| Palm tocotrienols decrease levels of pro-angiogenic markers in human umbilical vein endothelial cells (HUVEC) and murine mammary cancer cells
Kangarani, S., et.al (2012). Genes Nutr.
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In this study, the anti-angiogenic activity of palm tocotrienol-rich fraction (TRF) and its individual components (γ- and δ-tocotrienol) were first investigated in vitro in human umbilical vein endothelial cells (HUVEC) and 4T1 mouse mammary cancer cells. This study confirms previous observations that palm tocotrienols exhibit anti-angiogenic properties that may inhibit tumour progression. |
| Gamma-tocotrienol inhibits HGF-dependent mitogenesis and Met activation in highly malignant mammary tumor cells.
Ayoub, N.M., et.al (2011). Cell Prolif. |
Aberrant Met signalling is associated with aggressive cancer cell phenotypes. γ-tocotrienol displays potent anti-cancer activity that is associated with suppression of HER/ErbB receptor signalling. Experiments were conducted to investigate the effects of γ-tocotrienol treatment on HGF-dependent +SA mammary tumour cell proliferation, upon Met activation. These findings show, for the first time, the inhibitory effects of γ-tocotrienol on Met expression and activation, and strongly suggest that γ-tocotrienol treatment may provide significant health benefits in prevention and/or treatment of breast cancer, in women with deregulated HGF/Met signaling.
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| Tocotrienol-treated MCF-7 human breast cancer cells show down-regulation of AP15 and up-regulation of MIG6 genes.
Ramdas, P., et.al (2011). Cancer Genomics Proteomics.
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This study aimed to identify the genes that are regulated in human breast cancer cells following exposure to various isomers of vitamin E as these may be potential targets for the treatment of breast cancer. Treatment of human MCF-7 cells with tocotrienol isomers causes the down-regulation of the API5 gene and up-regulation of the MIG6 gene and the differential expression of other genes reported to play a key role in breast cancer biology. |
| Palm tocotrienols inhibit proliferation of murine mammary cancer cells and induce expression of interleukin-24 mRNA.
Selvaduray, K.R., et.al (2010). J Interferon Cytokine Res.
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In this study, for the first time, the anticancer effect of tocotrienols is linked to increased expression of interleukin-24 (IL-24) mRNA, a cytokine reported to have antitumor effects in many cancer models. It is reported that tocotrienols have potent antiangiogenic and antitumor effects that is associated with increased levels of IL-24 mRNA. |
| Tocotrienols induce apoptosis in breast cancer cells lines via an endoplasmic reticulum stress-dependent increase in extrinsic death receptor signaling.
Park, S.K., et.al (201). Breast Cancer Res Treat. |
This study focused on investigating anticancer effects of tocotrienols and the mechanisms of apoptosis induction by tocotrienols in vivo and in vitro. Dietary delivery of γ-tocotrienol (γ-T3) suppressed tumor growth in a syngeneic implantation mouse mammary cancer model by inhibiting cell proliferation and inducing apoptosis. In cell culture studies, γ-T3 inhibited colony formation of a mouse mammary cancer cell line and human breast cancer cell lines. The anti-proliferative effects of tocotrienols were highly correlated with an increase in apoptosis based on Annexin V assessment. Treatment of human MDA-MB-231 and MCF-7 cells with γ-T3 induced cleavages of PARP as well as caspase-8, -9, and -3. Additional analyses showed that γ-T3 activated c-Jun NH(2)-terminal kinase (JNK) and p38 MAPK, and upregulated death receptor 5 (DR5) and C/EBP homologous protein (CHOP), an endoplasmic reticulum (ER) stress marker. Silencing either JNK or p38 MAPK reduced the increase in DR5 and CHOP and partially blocked γ-T3-induced apoptosis. Both DR5 and CHOP upregulation were required for γ-T3-induced apoptosis, and DR5 was transcriptionally regulated by CHOP after γ-T3 treatment. Moreover, γ-T3 increased the level of other ER-stress markers. Taken together, these results suggest that upregulation of DR5 by γ-T3 treatment is dependent on JNK and p38 MAPK activation which is mediated by ER-stress.
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| Redox-silent tocotrienol esters as breast cancer proliferation and migration inhibitors.
Behrey, F.A., et.al (2010). Bioorg Med Chem. |
Tocotrienols’ chemical instability, poor water solubility, NPC1L1-mediated transport, and rapid metabolism are examples of such obstacles which hinder the therapeutic use of these valuable natural products. Vitamin E esters like α-tocopheryl succinate were prepared to significantly improve chemical and metabolic stability, water solubility, and potency. Thus, 12 semisynthetic tocotrienol ester analogues 4-15 were prepared by direct esterification of natural tocotrienol isomers with various acid anhydrides or chlorides. Esters 4-15 were evaluated for their ability to inhibit the proliferation and migration of the mammary tumor cells +SA and MDA-MB-231, respectively. Esters 5, 9, and 11 effectively inhibited the proliferation of the highly metastatic +SA rodent mammary epithelial cells with IC(50) values of 0.62, 0.51, and 0.86μM, respectively, at doses that had no effect on immortalized normal mouse CL-S1 mammary epithelial cells. Esters 4, 6, 8-10, and 13 inhibited 50% of the migration of the human metastatic MDA-MB-231 breast cancer cells at a single 5μM dose in wound-healing assay. The most active ester 9 was 1000-fold more water-soluble and chemically stable versus its parent α-tocotrienol (1). These findings strongly suggest that redox-silent tocotrienol esters may provide superior therapeutic forms of tocotrienols for the control of metastatic breast cancer.
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| Differential suppression of proliferation in MCF-7 and MDA-MB-231 breast cancer cells exposed to alpha-, gamma-, and delta-tocotrienols is accompanied by altered expression of oxidative stress modulatory enzymes.
Hsieh, T.C., et.al (2010). Anticancer Res. |
The present studies were performed to test whether the differential growth inhibition resulting from exposure to α-, γ- and δ-tocotrienols in estrogen receptor-positive human MCF-7 and estrogen receptor-negative MDA-MB-231 breast cancer cells might be accompanied by changes in phase II antioxidant enzymes. These studies demonstrate that different tocotrienols show distinct and selective activity in regulating the NRF2-KEAP1, in coordination with the induced expression of cytoprotective oxidative stress modulatory genes and regulation of proliferation in breast cancer cells. |
| Gamma-tocotrienol controls proliferation, modulates expression of cell cycle regulatory proteins and up-regulates quinone reducatase NQO2 in MCF-7 breast cancer cells.
Hsieh, T.C., et.al (2010). Anticancer Res.
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In this study, the effects of gamma-tocotrienol were examined with regard to its ability to suppress cell proliferation via modulation of cell cycle regulatory protein expression, and also from the perspective of control of cellular oxidoreductive status through regulation of detoxification enzymes, e.g., quinone reductase NQO2, using estrogen receptor-positive MCF-7 human breast cancer cells. By exerting control on expression of specific cell cycle regulatory proteins in concomitance with suppression of cell proliferation, as well as the induction of NQO2, gamma-tocotrienol offers promise as an added chemopreventive and/or chemotherapeutic agent against breast cancer carcinogenesis.
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| Tocotrienols activity in MCF-7 breast cancer cells: Involvement of ERbeta signal transduction.
Cormitato, R., et.al (2010). Mol Nutr Foo Res. |
In this study, it is reported that, in MCF-7 breast cancer cell, expressing both ERalpha and ERbeta, PTRF treatment increases ERbeta nuclear translocation, as demonstrated by immunofluorescence experiments and significantly inhibits ERalpha expression (-458.91-fold of change) and complete disappearing of the protein from the nucleus. Moreover, PTRF treatment induces ER-dependent genes expression (macrophage inhibitory cytokine-1, early growth response-1 and Cathepsin D) which is inhibited by the ER inhibitor, ICI 182.780, and induces DNA fragmentation. Finally, cDNA-array experiments suggest that the activation of specific pathways in cells treated with gamma-TT with respect to alpha-TT. The data suggest a novel potential molecular mechanism for TTs activity.
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| Id1, inhibitor of differentiation, is a key protein mediating anti-tumor responses of gamma-tocotrienol in breast cancer cells.
Yap, W.N., et.al (2010). Cancer Lett. |
This study aimed at deciphering the molecular pathways responsible for its activity. Our results showed that treatment of BCa cells with gamma-tocotrienol resulted in induction of apoptosis as evidenced by activation of pro-caspases, accumulation of sub-G1 cells and DNA fragmentations. The results suggested that the anti-proliferative and chemosensitization effect of gamma-tocotrienol on BCa cells may be mediated through downregulation of Id1 protein.
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| Enhanced antiproliferative and apoptotic response to combined treatment of gamma-tocotrienol with erlotinib or gefitinib in mammary tumor cells.
Bachawal, S.V., et.al (2010). BMC Cancer. |
Aberrant ErbB receptor signaling is associated with various types of malignancies. gamma-Tocotrienol is a member of the vitamin E family of compounds that displays potent anticancer activity that is associated with suppression in ErbB receptor phosphorylation and mitogenic signaling. Erlotinib and gefitinib are tyrosine kinase inhibitors that block ErbB1 receptor activation, whereas trastuzumab is a monoclonal antibody that has been designed to specifically inhibit ErbB2 receptor activation. However, the clinical effectiveness of these agents have been disappointing because of cooperation between different ErbB family members that can rescue cancer cells from agents directed against a single ErbB receptor subtype. It was hypothesized that targeting multiple ErbB receptor subtypes with combined treatment of gamma-tocotrienol and ErbB receptor inhibitors would provide greater anticancer effects than monotherapy targeting only a single ErbB receptor subtype. Combination treatment of gamma-tocotrienol with specific ErbB receptor inhibitors is more effective in reducing mammary tumor cell growth and viability than high dose monotherapy, suggesting that targeting multiple ErbB receptors with combination therapy may significantly improve the therapeutic response in breast cancer patients.
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| Combined gamma-tocotrienol and erlontibib.gefitinib treatment suppresses Stat and Akt signaling in murine mammary tumor cells.
Bachawal, S.V., et.al (2010). Anticancer Res. |
Heterodimer cooperation between ErbB receptors has limited clinical usefulness of receptor tyrosine kinase inhibitors (TKIs), erlotinib and gefitinib in the treatment of cancer. However, combination treatment of TKIs with gamma-tocotrienol targets multiple ErbB receptors and significantly inhibit +SA murine mammary tumor cell growth. Combined treatment of gamma-tocotrienol with erlotinib or gefitinib prevents ErbB receptor heterodimer cooperation and inhibits EGF-dependent mitogenic signaling in +SA murine mammary tumor cells. These findings strongly suggest that combination treatment may significantly improve therapeutic responsiveness in breast cancer patients.
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| Anti-proliferative effects of gamma-tocotrienol on mammary tumor cells are associated with suppression of cell cycle progression. Samant, G.V., et.al (2010). Cell Prolif. |
Previous studies have shown that gamma-tocotrienol induces potent anti-proliferative effects on +SA mammary tumour cells in culture; here, investigations have been conducted to determine its effects on intracellular signalling proteins involved in regulating cell cycle progression. These findings strongly suggest that anti-proliferative effects of gamma-tocotrienol are associated with reduction in cell cycle progression from G(1) to S, as evidenced by increased p27 levels, and a corresponding decrease in cyclin D1, CDK2, CDK4, CDK6 and phosphorylated Rb levels.
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| Endoplasmic reticulum stress mediates gamma-tocotrienol-induced apoptosis in mammary tumor cells.
Wali, V.B., et.al (2009). Apoptosis. |
Studies were conducted to determine the role of endoplasmic reticulum (ER) stress in mediating gamma-tocotrienol-induced apoptosis in +SA mammary tumor cells. Treatment with 15-40 microM gamma-tocotrienol induced +SA cell death in a dose-responsive manner, and these effects were associated with a corresponding increase in poly (ADP-ribose) polymerase (PARP)-cleavage and activation of protein kinase-like endoplasmic reticulum kinase/eukaryotic translational initiation factor/activating transcription factor 4 (PERK/eIF2alpha/ATF-4) pathway, a marker of ER stress response. These results demonstrate that ER stress apoptotic signaling is associated with gamma-tocotrienol-induced apoptosis in +SA mammary tumor cells.
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| A novel mechanism of natural vitamin E tocotrienol activity: involvement of ERβ signal transduction
Camitato, R., et.al (2009). Am J Physiol Endocrinol Metab.
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Vitamin E is a generic term used to indicate all tocopherol (TOC) and tocotrienol (TT) derivates. In the last few years, several papers have shown that a TT-rich fraction (TTRF) extracted from palm oil inhibits proliferation and induces apoptosis in a large number of cancer cells. However, the molecular mechanism(s) involved in TT action is still unclear. In the present study, we proposed for the first time a novel mechanism for TT activity that involves estrogen receptor (ER) signaling. In silico simulations and in vitro binding analyses indicated a high affinity of TTs for ERβ but not for ERα. In addition, in ERβ-containing MDA-MB-231 breast cancer cells, we demonstrated that TTs increase the ERβ translocation into the nucleus, which in turn activates estrogen-responsive genes (MIC-1, EGR-1 and cathepsin D), as demonstrated by cell preincubation with the ER inhibitor ICI-182,780. Finally, we observed that TT treatment is associated with alteration of cell morphology, DNA fragmentation, and caspase-3 activation. Altogether, these experiments elucidated the molecular mechanism underling γ- and δ-TT effects.
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| Combined treatment of gamma-tocotrienol with statins induce mammary tumor cell cycle arrest in G1.
Wali, V.B., et.al (2009). Exp Biol Med (Maywood). |
To investigate the mechanism mediating this growth inhibition, studies were conducted to determine the effect of combination low dose gamma-tocotrienol and statin treatment on +SA mammary tumor cell cycle progression. These findings demonstrate that combination low dose statin and gamma-tocotrienol treatment induced mammary tumor cell cycle arrest at G1, resulting from an increase in p27 expression, and a corresponding decrease in cyclin D1, CDK2, and hypophosphorylation of Rb protein. These findings suggest that combined treatment of statins with gamma-tocotrienol may provide significant health benefits in the treatment of breast cancer in women, while avoiding myotoxicity associated with high dose statin monotherapy.
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| Design and QSAR study of analogs of gamma-tocotrienol with enhanced antiproliferative activity against human breast cancer cells.
Nikolic, K., et.al (2008). J Mol Graph Model. |
Quantitative structure-activity relationships (QSAR) study has been performed for two sets of the antitumor drugs against human breast cancer MCF-7 cell lines, alpha-tocopherol and cholesterol derivatives. Constitutional, geometrical, physico-chemical and electronic descriptors (using the density functional theory, B3LYP/6-31G (d,p) basis set) were computed and analyzed. The most relevant of these descriptors were grouped and multiple linear regressions have been carried out. Optimal QSAR models with three and four variables, R(2)>0.95 and cross-validation parameter q(pre)(2)>0.88, were selected. Based on the QSAR study, novel vitamin-E derivatives (compounds D-1 and D-2) were designed and their antiproliferative activities were evaluated using the proposed regression models. Calculated antiproliferative activities of the designed compounds, IC(50) (D-1): 3.09 microM and IC(50) (D-2): 3.54 microM, were significantly stronger than anticancer effect of the other analyzed compounds IC(50): 4-1461 microM. |
| Suppression of cell proliferation and gene expression by combinatorial synergy of EGCG, resveratrol and gamma-tocotrienol in estrogen receptor-positive MCF-7 breast cancer cells.
Hsieh & Wu, (2008). Int J Oncol.
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The estrogen receptor-positive MCF-7 breast cancer cells as a model, whether the combination of epigallocatechin gallate (EGCG), resveratrol and gamma-tocotrienol at suboptimal doses elicits synergism in suppressing cell proliferation, modulating gene expression, and increasing antioxidant activity, as compared to each of the three phytochemicals added alone were investigated. The results suggest that diet-based protection against breast cancer may partly derive from synergy amongst dietary phytochemicals directed against specific molecular targets in responsive breast cancer cells, and provide support for the feasibility of the development of a diet-based combinatorial approach in the prevention and treatment of breast cancer.
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| Growth Inhibition of Human MDA-MB-231 Breast Cancer Cells by δ-Tocotrienol Is Associated with Loss of Cyclin D1/CDK4 Expression and Accompanying Changes in the State of Phosphorylation of the Retinoblastoma Tumor Suppressor Gene Product
Elangovan, S., et.al (2008). Anticancer Res.
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In this study, the effect of δ-tocotrienol on cell cycle arrest was assessed by studying the retinoblastoma protein (Rb) levels and phosphorylation status, levels of E2F (a transcription factor critically involved in the G1/S-phase transition of the mammalian cell cycle; originally identified as a DNA-binding protein essential for early region 1A-dependent activation of the adenovirus promoter designated E2), and other cell cycle controlling proteins in estrogen receptor-negative MDA-MB-231 breast cancer cells. To the best of our knowledge, this study was the first to reveal that the target of cell proliferative inhibitory action of δ-tocotrienol in a model estrogen receptor-negative human breast cancer cell line MDAMB-231 is mediated by the loss of cyclin D1 and associated suppression of site-specific Rb phosphorylation, suggesting its future development and use as an anticancer agent.
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| Synergistic antiproliferative effects of gamma-tocotrienol and statin treatment on mammary tumor cells.
Wali, .B., et.al (2007). Lipids.
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Studies were conducted to determine if combined low dose treatment of gamma-tocotrienol with individual statins resulted in a synergistic antiproliferative effect on neoplastic mouse +SA mammary epithelial cells. Treatment with 3-4 microM gamma-tocotrienol or 2-8 microM simvastatin, lovastatin or mevastatin alone resulted in a significant decrease, whereas treatment with 10-100 microM pravastatin had no effect on +SA cell growth. However, combined treatment of subeffective doses (0.25 or 10 microM) of individual statins with 0.25-2.0 microM gamma-tocotrienol resulted in a dose-responsive synergistic inhibition in +SA cell proliferation. Additional studies showed that treatment with subeffective doses of individual statins or gamma-tocotrienol alone had no effect, whereas combined treatment of these compounds resulted in a relatively large decrease in intracellular levels of phosphorylated (activated) MAPK, JNK, p38, and Akt. These findings strongly suggest that combined low dose treatment of gamma-tocotrienol with individual statins may have potential value in the treatment of breast cancer without causing myotoxicity that is associated with high dose statin treatment.
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| Gamma-tocotrienol inhibits ErbB3-dependent PI3K/Akt mitogenic signaling in neoplastic mammary epithelial cells.
Samant, G.V., et.al (2006).Cell Prolif. |
Studies were conducted to investigate the direct effects of gamma-tocotrienol treatment on specific components within the PI3K/PDK-1/Akt mitogenic pathway. +SA cells were grown in culture and maintained in serum-free media containing 10 ng/ml EGF as a mitogen. These findings strongly suggest that the antiproliferative effects of gamma-tocotrienol in neoplastic +SA mouse mammary epithelial cells are mediated by a suppression in ErbB3-receptor tyrosine phosphorylation and subsequent reduction in PI3K/PDK-1/Akt mitogenic signaling.
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| Intracellular signaling mechanisms mediating the antiproliferative and apoptotic effects of gamma-tocotrienol in neoplastic mammary epithelial cells.
Sylvester, P.W., et.al (2005). J Plant Physiol. |
Tocotrienols, a subgroup within the vitamin E family of compounds, display potent antiproliferative and apoptotic activity against neoplastic mammary epithelial cells at treatment doses that have little or no effect on normal cell growth and function. Recent studies have shown that treatment with a growth inhibitory, but non-cytotoxic dose (4 microM) of gamma-tocotrienol inhibits phosphatidylinositol-3-kinase-dependent kinase (Pl3K)/Pl3K-dependent kinase 1 (PDK-1)/mitogenic signaling over a 2-3 day period following treatment exposure, and these effects were not found to be associated with an increased in either phosphatase and tensin homologue deleted from chromosome 10 (PTEN) or protein phosphatase type 2A (PP2A) phosphatase activity. In summary, both the antiproliferative and apoptotic effects of gamma-tocotrienol appear to be mediated by a reduction in the Pl3K/PDK-1 /Akt signaling, an important pathway associated with cell proliferation and survival in neoplastic mammary epithelial cells.
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| Gamma-tocotrienol inhibits neoplastic mammary epithelial cell proliferation by decreasing Akt and nuclear factor kappaB activity.
Shah, S.J., et.al (2005). Exp Biol Med (Maywood). |
Tocotrienols, a subgroup within the vitamin E family of compounds, have been shown to display potent anticancer activity and inhibit preneoplastic and neoplastic mammary epithelial cell proliferation at treatment doses that have little or no effect on normal cell growth and function. However, the specific intracellular mechanisms mediating the antiproliferative effects of tocotrienols are presently unknown. Because Akt and nuclear factor kappaB (NFkappaB) are intimately involved in mammary tumor cell proliferation and survival, studies were conducted to determine the effects of gamma-tocotrienol on Akt and NFkappaB activity in neoplastic +SA mammary epithelial cells in vitro. Treatment with 0-8 microM gamma-tocotrienol for 0-3 days caused a dose-responsive inhibition in +SA cell growth and mitotic activity, as determined by MTT colorimetric assay and proliferating cell nuclear antigen immunocytochemical staining, respectively. Studies also showed that treatment with 4 microM gamma-tocotrienol, a dose that inhibited +SA cell growth by more than 50% compared with that of untreated control cells, decreased intracellular levels of activated phosphotidylinositol 3-kinase-dependent kinase (PI3K)-dependent kinase 1 (phospho-PDK-1) and Akt, and reduced phospho-Akt kinase activity. Furthermore, these effects were not found to be associated with an increase in either phosphatase and tensin homologue deleted from chromosome 10 (PTEN) or protein phosphatase type 2A phosphatase activity. In addition, gamma-tocotrienol treatment was shown to decrease NFkappaB transcriptional activity, apparently by suppressing the activation of IkappaB-kinase-alpha/beta, an enzyme associated with inducing NFkappaB activation. In summary, these findings demonstrate that the antiproliferative effects of gamma-tocotrienol result, at least in part, from a reduction in Akt and NFkappaB activity in neoplastic +SA mammary epithelial cells.
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| Tocotrienol-induced cytotoxicity is unrelated to mitochondrial stress apoptotic signaling in neoplastic mammary epithelial cells.
Shah, S.J. (2005). Biochem Cell Biol. |
Tocotrienols and tocopherols represent the 2 subgroups within the vitamin E family of compounds, but tocotrienols display significantly greater apoptotic activity against a variety of cancer cell types. However, the exact mechanism mediating tocotrienol-induced apoptosis is not understood. Studies were conducted to determine the effects of tocotrienols on mitochondrial-stress-mediated apoptotic signaling in neoplastic +SA mammary epithelial cells grown in vitro. Exposure for 24 h to 0-20 micromol/L gamma-tocotrienol resulted in a dose-responsive increase in +SA cells undergoing apoptosis, as determined by flow cytometric analysis of Annexin V staining. However, tocotrienol-induced apoptosis was not associated with a disruption or loss of mitochondrial membrane potential, or the release of mitochondrial cytochrome c into the cytoplasm, as determined by JC-1 flow cytometric staining and ELISA assay, respectively. Interestingly, apoptotic +SA cells showed a paradoxical decrease in mitochondrial levels of pro-apoptotic proteins Bid, Bax, and Bad, and a corresponding increase in mitochondrial levels of anti-apoptotic proteins, Bcl-2 and Bcl-xL, suggesting that mitochondrial membrane stability and integrity might actually be enhanced for a limited period of time following acute tocotrienol exposure. In summary, these findings clearly demonstrate that tocotrienol-induced apoptosis occurs independently of mitochondrial stress apoptotic signaling in neoplastic +SA mammary epithelial cells.
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| Tocotrienol-rich fraction from palm oil and gene expression in human breast cancer cells.
Nesaretnam, K., et.al (2004). Ann N Y Acad Sci. |
Vitamin E is important not only for its cellular antioxidant and lipid-lowering properties, but also as an antiproliferating agent. It has also been shown to contribute to immunoregulation, antibody production, and resistance to implanted tumors. It has recently been shown that tocotrienols are the components of vitamin E responsible for growth inhibition in human breast cancer cells in vitro as well as in vivo through estrogen-independent mechanisms. Although tocotrienols act on cell proliferation in a dose-dependent manner and can induce programmed cell death, no specific gene regulation has yet been identified. In order to investigate the molecular basis of the effect of a tocotrienol-rich fraction (TRF) from palm oil, we performed a cDNA array analysis of cancer-related gene expression in estrogen-dependent (MCF-7) and estrogen-independent (MDA-MB-231) human breast cancer cells. The human breast cancer cells were incubated with or without 8 mug/mL of tocotrienols for 72 h. RNA was subsequently extracted and subjected to reverse transcription before being hybridized onto cancer arrays. Tocotrienol supplementation modulated significantly 46 out of 1200 genes in MDA-MB-231 cells. In MCF-7 cells, tocotrienol administration was associated with a lower number of affected genes. Interestingly, only three were affected in a similar fashion in both cell lines: c-myc binding protein MM-1, 23-kDa highly basic protein, and interferon-inducible protein 9-27 (IFITM-1). These proteins are most likely involved in the cell cycle and can exert inhibitory effects on cell growth and differentiation of the tumor cell lines. These data suggest that tocotrienols are able to affect cell homeostasis, possibly independent of their antioxidant activity.
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| Pro-apoptotic mechanisms of action of a novel vitamin E analog (alpha-TEA) and a naturally occurring form of vitamin E (delta-tocotrienol) in MDA-MB-435 human breast cancer cells.
Shun, M.C., et.al (2004). Nutr Cancer. |
Objectives of these studies were to characterize signaling events involved in the pro-apoptotic actions of a naturally occurring form of vitamin E, delta-tocotrienol, and a novel vitamin E analog, alpha-tocopherol ether acetic acid analog [alpha-TEA; 2,5,7,8-tetramethyl-2R-(4R,8R,12-trimethyltridecyl)chroman-6-yloxyacetic acid]. Like VES, alpha-TEA and delta-tocotrienol induced estrogen-nonresponsive MDA-MB-435 and estrogen-responsive MCF-7 human breast cancer cells to undergo high levels of apoptosis in a concentration- and time-dependent fashion. Like VES, the two compounds induced either no or lower levels of apoptosis in normal human mammary epithelial cells and immortalized but nontumorigenic human MCF-10A cells. The pro-apoptotic mechanisms triggered by the structurally distinct alpha-TEA and delta-tocotrienol were identical to those previously reported for VES, that is, alpha-TEA- and delta-tocotrienol-induced apoptosis involved up-regulation of TGF-beta receptor II expression and TGF-beta-, Fas- and JNK-signaling pathways. These data provide a better understanding of the anticancer actions of a dietary form of vitamin E (delta-tocotrienol) and a novel nonhydrolyzable vitamin E analog (alpha-TEA).
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| Tocotrienol-induced caspase-8 activation is unrelated to death receptor apoptotic signaling in neoplastic mammary epithelial cells.
Shah, S., et.al (2004). Exp Biol Med (Maywood). |
Tocotrienols, a subclass in the vitamin E family of compounds, have been shown to induce apoptosis by activating caspase-8 and caspase-3 in neoplastic mammary epithelial cells. Since caspase-8 activation is associated with death receptor apoptotic signaling, studies were conducted to determine the exact death receptor/ligand involved in tocotrienol-induced apoptosis. Highly malignant +SA mouse mammary epithelial cells were grown in culture and maintained in serum-free media. Treatment with 20 microM gamma-tocotrienol decreased+SA cell viability by inducing apoptosis, as determined by positive terminal dUTP nick end labeling (TUNEL) immunocytochemical staining. Western blot analysis showed that gamma-tocotrienol treatment increased the levels of cleaved (active) caspase-8 and caspase-3. Combined treatment with caspase inhibitors completely blocked tocotrienol-induced apoptosis. Additional studies showed that treatment with 100 ng/ml tumor necrosis factor-alpha (TNF-alpha), 100 ng/ml FasL, 100 ng/ml TNF-related apoptosis-inducing ligand (TRAIL), or 1 microg/ml apoptosis-inducing Fas antibody failed to induce death in +SA cells, indicating that this mammary tumor cell line is resistant to death receptor-induced apoptosis. Furthermore, treatment with 20 microM gamma-tocotrienol had no effect on total, membrane, or cytosolic levels of Fas, Fas ligand (FasL), or Fas-associated via death domain (FADD) and did not induce translocation of Fas, FasL, or FADD from the cytosolic to the membrane fraction, providing additional evidence that tocotrienol-induced caspase-8 activation is not associated with death receptor apoptotic signaling. Other studies showed that treatment with 20 microM gamma-tocotrienol induced a large decrease in the relative intracellular levels of phospho-phosphatidylinositol 3-kinase (PI3K)-dependent kinase 1 (phospho-PDK-1 active), phospho-Akt (active), and phospho-glycogen synthase kinase3, as well as decreasing intracellular levels of FLICE-inhibitory protein (FLIP), an antiapoptotic protein that inhibits caspase-8 activation, in these cells. Since stimulation of the PI3K/PDK/Akt mitogenic pathway is associated with increased FLIP expression, enhanced cellular proliferation, and survival, these results indicate that tocotrienol-induced caspase-8 activation and apoptosis in malignant +SA mammary epithelial cells is associated with a suppression in PI3K/PDK-1/Akt mitogenic signaling and subsequent reduction in intracellular FLIP levels.
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| Role of GTP-binding proteins in reversing the antiproliferative effects of tocotrienols in preneoplastic mammary epithelial cells.
Sylvester, P.W., et.al (2002). Asia Pac J Clin Nutr. |
Tocotrienols are a subclass of vitamin E compounds that display potent anticancer activity. Determining the anticancer mechanism of action of tocotrienols will provide essential information necessary for understanding the potential health benefits of these compounds in reducing the risk of breast cancer in women. Epidermal growth factor (EGF) is a potent mitogen for normal and neoplastic mammary epithelial cells. Initial events in EGF-receptor (EGF-R) mitogenic-signalling are G-protein activation, stimulation of adenylyl cyclase and cyclic AMP (cAMP) production. Studies were conducted to determine if the antiproliferative effects of tocotrienols are associated with reduced EGF-induced G-protein and cAMP-dependent mitogenic signalling. Preneoplastic CL-S1 mouse mammary epithelial cells were grown in culture and maintained on serum-free media containing 0-25 micro mol/L tocotrienol-rich fraction of palm oil and/or different doses of pharmacological agents that alter intracellular cAMP levels. Tocotrienol-induced effects on EGF-receptor levels of tyrosine kinase activity, as well as EGF-dependent mitogen-activated pathway kinase (MAPK) and Akt activation, were determined by western blot analysis. Results demonstrate that the antiproliferative effects of tocotrienols in preneoplastic mammary epithelial cells do not reflect a reduction in EGF-receptor mitogenic responsiveness, but rather, result from an inhibition in early post-receptor events involved in cAMP production upstream from EGF-dependent MAPK and phosphoinositide 3-kinase/Akt mitogenic signalling. In summary, these data further characterise the mechanism of action of tocotrienols in suppressing preneoplastic mammary epithelial cell proliferation, and advance the current understanding of the potential health benefits of these compounds in reducing the risk of breast cancer in women.
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| Tocotrienols inhibit growth of ZR-75-1 breast cancer cells.
Nesaretnam, K., et.al (2000). Int J Food Sci Nutr. |
The vitamin E component of palm oil provides a rich source of tocotrienols which have been shown previously to be growth inhibitory to two human breast cancer cell lines: responsive MCF7 cells and unresponsive MDA-MB-231 cells. Data presented here shows that the tocotrienol-rich fraction (TRF) of palm oil and individual fractions (alpha, gamma and delta) can also inhibit the growth of another responsive human breast cancer cell line, ZR-75-1. At low concentrations in the absence of oestrogen tocotrienols stimulated growth of the ZR-75-1 cells, but at higher concentrations in the presence as well as in the absence of oestradiol, tocotrienols inhibited cell growth strongly. As for MCF7 cells, alpha-tocopherol had no effect on growth of the ZR-75-1 cells in either the absence or presence of oestradiol. In studying the effects of tocotrienols in combination with antioestrogens, it was found that TRF could further inhibit growth of ZR-75-1 cells in the presence of tamoxifen (10(-7) M and 10(-8) M). Individual tocotrienol fractions (alpha, gamma, delta) could inhibit growth of ZR-75-1 cells in the presence of 10(-8) M oestradiol and 10(-8) M pure antioestrogen ICI 164,384. The immature mouse uterine weight bioassay confirmed that TRF could not exert oestrogen antagonist action in vivo. These results provide evidence of wider growth-inhibitory effects of tocotrienols beyond MCF7 and MDA-MB-231 cells, and with an oestrogen-independent mechanism of action, suggest a possible clinical advantage in combining administration of tocotrienols with antioestrogen therapy.
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| Antiproliferative and apoptotic effects of tocopherols and tocotrienols on preneoplastic and neoplastic mouse mammary epithelial cells.
Mclntyre, B.S., et.al (2000). Proc Soc Exp Biol Med.
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Studies were conducted to determine the comparative effects of tocopherols and tocotrienols on preneoplastic (CL-S1), neoplastic (-SA), and highly malignant (+SA) mouse mammary epithelial cell growth and viability in vitro. These data also showed that highly malignant +SA cells were the most sensitive, whereas the preneoplastic CL-S1 cells were the least sensitive to the antiproliferative and apoptotic effects of tocotrienols, and suggest that tocotrienols may have potential health benefits in preventing and/or reducing the risk of breast cancer in women. |
| Antiproliferative and apoptotic effects of tocopherols and tocotrienols on normal mouse mammary epithelial cells.
Mclntyre, B.S., et.al (2000). Lipids.
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Studies were conducted to determine the comparative effects of tocopherols and tocotrienols on normal mammary epithelial cell growth and viability. Cells isolated from midpregnant BALB/c mice were grown within collagen gels and maintained on serum-free media. Treatment with 0-120 microM alpha- and gamma-tocopherol had no effect, whereas 12.5-100m microM tocotrienol-rich fraction of palm oil (TRF), 100-120 microM delta-tocopherol, 50-60 microM alpha-tocotrienol, and 8-14 microM gamma- or delta-tocotrienol significantly inhibited cell growth in a dose-responsive manner. In acute studies, 24-h exposure to 0-250 microM alpha-, gamma-, and delta-tocopherol had no effect, whereas similar treatment with 100-250 microM TRF, 140-250 microM alpha-, 25-100 microM gamma- or delta-tocotrienol significantly reduced cell viability. Growth-inhibitory doses of TRF, delta-tocopherol, and alpha-, gamma-, and delta-tocotrienol were shown to induce apoptosis in these cells, as indicated by DNA fragmentation. Results also showed that mammary epithelial cells more easily or preferentially took up tocotrienols as compared to tocopherols, suggesting that at least part of the reason tocotrienols display greater biopotency than tocopherols is because of greater cellular accumulation. In summary, these findings suggest that the highly biopotent gamma- and delta-tocotrienol isoforms may play a physiological role in modulating normal mammary gland growth, function, and remodeling.
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| Induction of apoptosis in human breast cancer cells by tocopherols and tocotrienols.
Yu, W., et.al (1999). Nutr Cancer. |
The apoptosis-inducing properties of RRR-alpha-, beta-, gamma-, and delta-tocopherols, alpha-, gamma-, and delta-tocotrienols, RRR-alpha-tocopheryl acetate (vitamin E acetate), and RRR-alpha-tocopheryl succinate (vitamin E succinate) were investigated in estrogen-responsive MCF7 and estrogen-nonresponsive MDA-MB-435 human breast cancer cell lines in culture. Apoptosis was characterized by two criteria: 1) morphology of 4,6-diamidino-2-phenylindole-stained cells and oligonucleosomal DNA laddering. Vitamin E succinate, a known inducer of apoptosis in several cell lines, including human breast cancer cells, served as a positive control. The estrogen-responsive MCF7 cells were more susceptible than the estrogen-nonresponsive MDA-MB-435 cells, with concentrations for half-maximal response for tocotrienols (alpha, gamma, and delta) and RRR-delta-tocopherol of 14, 15, 7, and 97 micrograms/ml, respectively. The tocotrienols (alpha, gamma, and delta) and RRR-delta-tocopherol induced MDA-MB-435 cells to undergo apoptosis, with concentrations for half-maximal response of 176, 28, 13, and 145 micrograms/ml, respectively. With the exception of RRR-delta-tocopherol, the tocopherols (alpha, beta, and gamma) and the acetate derivative of RRR-alpha-tocopherol (RRR-alpha-tocopheryl acetate) were ineffective in induction of apoptosis in both cell lines when tested within the range of their solubility, i.e., 10-200 micrograms/ml. In summary, these studies demonstrate that naturally occurring tocotrienols and RRR-delta-tocopherol are effective apoptotic inducers for human breast cancer cells.
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| Tocotrienols inhibit the growth of human breast cancer cells irrespective of estrogen receptor status. Nesaretnam, K., et.al (1998). Lipids. |
Potential antiproliferative effects of tocotrienols, the major vitamin E component in palm oil, were investigated on the growth of both estrogen-responsive (ER+) MCF7 human breast cancer cells and estrogen-unresponsive (ER-) MDA-MD-231 human breast cancer cells, and effects were compared with those of α-tocopherol (αT). The tocotrienol-rich fraction (TRF) of palm oil inhibited growth of MCF7 cells in both the presence and absence of estradiol with a nonlinear dose-response but such that complete suppression of growth was achieved at 8 μg/mL. MDA-MB-231 cells were also inhibited by TRF but with a linear dose-response such that 20 μg/mL TRF was needed for complete growth suppression. Separation of the TRF into individual tocotrienols revealed that all fractions could inhibit growth of both ER+ and ER- cells and of ER+ cells in both the presence and absence of estradiol. However, the γ- and δ-fractions were the most inhibitory. Complete inhibition of MCF7 cell growth was achieved at 6 μg/mL of γ-tocotrienol/δ-tocotrienol (γT3/δT3) in the absence of estradiol and 10μm/mL of δT3 in the presence of estradiol, whereas complete suppression of MDA-MB-231 cell growth was not achieved even at concentrations of 10μg/mL of δT3. By contrast to these inhibitory effects of tocotrienols, αT had no inhibitory effect on MCF7 cell growth in either the presence or the absence of estradiol, nor on MDA-MB-231 cell growth. These results confirm studies using other sublines of human breast cancer cells and demonstrate that tocotrienols can exert direct inhibitory effects on the growth of breast cancer cells. In searching for the mechanism of inhibition, studies of the effects of TRF on estrogen-regulated pS2 gene expression in MCF7 cells showed that tocotrienols do not act via an estrogen receptor-mediated pathway and must therefore act differently from estrogen antagonists. Furthermore, tocotrienols did not increase levels of growth-inhibitory insulin-like growth factor binding proteins (IGFBP) in MCF7 cells, implying also a different mechanism from that proposed for retinoic acid inhibition of estrogen-responsive breast cancer cell growth. Inhibition of the growth of breast cancer cells by tocotrienols could have important clinical implications not only because tocotrienols are able to inhibit the growth of both ER+ and ER- phenotypes but also because ER+ cells could be growth-inhibited in the presence as well as in the absence of estradiol. Future clinical applications of TRF could come from potential growth suppression of ER+ breast cancer cells otherwise resistant to growth inhibition by antiestrogens and retinoic acid.
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| Inhibition of Proliferation of Estrogen Receptor–Negative MDA-MB-435 and –Positive MCF-7 Human Breast Cancer Cells by Palm Oil Tocotrienols and Tamoxifen, Alone and in Combination
Gutherie, N., et.al (1997). J Nutr.
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Tocotrienols are a form of vitamin E, having an unsaturated isoprenoid side-chain rather than the saturated side-chain of tocopherols. The tocotrienol-rich fraction (TRF) from palm oil contains α-tocopherol and a mixture of α-, γ- and δ-tocotrienols. Earlier studies have shown that tocotrienols display anticancer activity. We previously reported that TRF, α-, γ- and δ-tocotrienols inhibited proliferation of estrogen receptor-negative MDA-MB-435 human breast cancer cells with 50% inhibitory concentrations (IC50) of 180, 90, 30 and 90 μg/mL, respectively, whereas α-tocopherol had no effect at concentrations up to 500 μg/mL. Further experiments with estrogen receptor-positive MCF-7 cells showed that tocotrienols also inhibited their proliferation, as measured by [3H] thymidine incorporation. The IC50s for TRF, α-tocopherol, α-, γ- and δ-tocotrienols were 4, 125, 6, 2 and 2 μg/mL, respectively. Tamoxifen, a widely used synthetic antiestrogen inhibits the growth of MCF-7 cells with an IC50 of 0.04 μg/mL. We tested 1:1 combinations of TRF, α-tocopherol and the individual tocotrienols with tamoxifen in both cell lines. In the MDA-MB-435 cells, all of the combinations were found to be synergistic. In the MCF-7 cells, only 1:1 combinations of γ- or δ-tocotrienol with tamoxifen showed a synergistic inhibitory effect on the proliferative rate and growth of the cells. The inhibition by tocotrienols was not overcome by addition of excess estradiol to the medium. These results suggest that tocotrienols are effective inhibitors of both estrogen receptor-negative and -positive cells and that combinations with tamoxifen should be considered as a possible improvement in breast cancer therapy.
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Study objectives/ findings |
| Tocotrienols and breast cancer: the evidence to date
Nesaretnam, K., et.al (2012). Genes Nutr. |
Breast cancer is the second most frequent cancer affecting women worldwide after lung cancer. The toxicity factor associated with synthetic drugs has turned the attention toward natural compounds as the primary focus of interest as anticancer agents. Vitamin E derivatives consisting of the well-established tocopherols and their analogs namely tocotrienols have been extensively studied due to their remarkable biological properties. While tocopherols have failed to offer protection, tocotrienols, in particular, α-, δ-, and γ-tocotrienols alone and in combination have demonstrated anticancer properties. The discovery of the antiangiogenic, antiproliferative, and apoptotic effects of tocotrienols, as well as their role as an inducer of immunological functions, not only reveals a new horizon as a potent antitumor agent but also reinforces the notion that tocotrienols are indeed more than antioxidants. On the basis of a transcriptomic platform, we have recently demonstrated a novel mechanism for tocotrienol activity that involves estrogen receptor (ER) signaling. In silico simulations and in vitro binding analyses indicate a high affinity of specific forms of tocotrienols for ERβ, but not for ERα. Moreover, we have demonstrated that specific tocotrienols increase ERβ translocation into the nucleus which, in turn, activates the expression of estrogen-responsive genes (MIC–1, EGR–1 and Cathepsin D) in breast cancer cells only expressing ERβ cells (MDA-MB-231) and in cells expressing both ER isoforms (MCF-7). The binding of specific tocotrienol forms to ERβ is associated with the alteration of cell morphology, caspase-3 activation, DNA fragmentation, and apoptosis. Furthermore, a recently concluded clinical trial seems to suggest that tocotrienols in combination with tamoxifen may have the potential to extend breast cancer-specific survival.
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| Disruption of mitochondria during tocotrienol-induced apoptosis in MDA-MB-231 human breast cancer cells.
Takahashi, K., et.al (2004). Biochem Pharmacol. |
The objective of this study was to clarify the apoptotic effects of tocotrienols, with especial emphasis in determining if the mitochondria-mediated death pathway is activated when human breast cancer cells are incubated with a specific tocotrienol isomer. During incubation with gamma-tocotrienol, MDA-MB-231 human breast cancer cells showed membrane blebbing, and apoptotic bodies were present. Upon 4′,6-diamidino-2-phenylindole staining of the cells, chromatin condensation and fragmentation were observed. Additionally, the annexin V-binding assay detected the translocation of membrane phospholipid during earlier analysis of the cells. Taken together, these results further establish that gamma-tocotrienol can induce apoptosis in human breast cancer cells. To help elucidate how gamma-tocotrienol induced the apoptosis, some important parameters related to the mitochondria-mediated death pathway were examined next. In gamma-tocotrienol-treated cells, the mitochondria were disrupted. Collapse of the mitochondrial membrane potential was detected, and cytochrome c was released later from mitochondria. However, expression of Bax and Bcl-2 (mRNA and protein) did not change. Furthermore, poly-(ADP-ribose)-polymerase cleavage was not detected, suggesting that caspases were not involved in the gamma-tocotrienol-induced apoptosis. These results imply that cytochrome c is not the critical protein released from mitochondria that triggers gamma-tocotrienol-induced apoptosis in MDA-MB-231 cells.
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| Antioxidants in dietary oils: Their potential role in breast cancer prevention
Sylvester, P.W., (2002). Malays J Nutr. |
Edible oils contain variable amounts of natural antioxidants such as vitamin E. Antioxidants act not only to prevent lipid peroxidation and free-radical production, but also display potent anticancer activity. The vitamin E family of compounds is divided into two subgroups called tocopherols and tocotrienols, but only tocotrienols display potent anticancer activity at treatment doses that have little or no effect on normal cell growth or viability. Palm oil contains the highest concentrations of natural tocotrienols. Tocotrienols induced apoptosis or programmed cell death in breast cancer cells. Morphological and biochemical characteristics of apoptosis, such as nuclear and cytoplasmic condensation and DNA fragmentation, are mediated by the activation of cysteine proteases called caspases. Apoptosis is triggered by the activation of initiator caspases (caspase-8 or 9) that subsequently activate effector caspases (caspase-3, 6, and 7). Studies were conducted using the highly malignant +SA mouse mammary epithelial cell line to determine if tocotrienol-induced programmed cell death is mediated through the caspase-8 or caspase-9 pathway. Treatment with cytotoxic doses of tocotrienol resulted in a large increase in caspase-8 and caspase-3, but not caspase-9 activity. Combined treatment of tocotrienol with selective caspase-8 or caspase-3 inhibitors completely blocked tocotrieno-linduced apoptosis and activation of caspase-8 and caspase-3, respectively. These findings demonstrate that tocotrienolinduced apoptosis in highly malignant mammary epithelial cells is mediated through caspase-8 activation, and may provide essential information necessary for understanding the potential health benefits of these compounds in preventing and/or reducing the risk of breast cancer in women.
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| Does lack of tocopherols and tocotrienols put women at increased risk of breast cancer?
Schwenke, D.C., et.al (2002). J Nutr Biochem. |
Breast cancer is the leading site of new cancers in women and the second leading cause (after lung cancer) of cancer mortality in women. Observational studies that have collected data for dietary exposure to alpha-tocopherol with or without the other related tocopherols and tocotrienols have suggested that vitamin E from dietary sources may provide women with modest protection from breast cancer. However, there is no evidence that vitamin E supplements confer any protection whatever against breast cancer. Observational studies that have assessed exposure to vitamin E by plasma or adipose tissue concentrations of alpha-tocopherol have failed to provide consistent support for the idea that alpha-tocopherol provides any protection against breast cancer. In addition, evidence from studies in experimental animals suggest that alpha-tocopherol supplementation alone has little effect on mammary tumors. In contrast, studies in breast cancer cells indicate that alpha- gamma-, and delta-tocotrienol, and to a lesser extent delta-tocopherol, have potent antiproliferative and proapoptotic effects that would be expected to reduce risk of breast cancer. Many vegetable sources of alpha-tocopherol also contain other tocopherols or tocotrienols. Thus, it seems plausible that the modest protection from breast cancer associated with dietary vitamin E may be due to the effects of the other tocopherols and the tocotrienols in the diet. Additional studies will be required to determine whether this may be the case, and to identify the most active tocopherol/tocotrienol.
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| Effect of tocotrienols on the growth of a human breast cancer cell line in culture.
Nesaretnam, K., et.al (1995). Lipids. |
The tocotrienol-rich fraction (TRF) of palm oil consists of tocotrienols and some α-tocopherol (α-T). Tocotrienols are a form of vitamin E having an unsaturated side-chain, rather than the saturated side-chain of the more common tocopherols. Because palm oil has been shown not to promote chemically-induced mammary carcinogenesis, we tested effects of TRF and α-T on the proliferation, growth, and plating efficiency (PE) of MDA-MB-435 estrogen-receptor-negative human breast cancer cells. TRF inhibited the proliferation of these cells with a concentration required to inhibit cell proliferation by 50% of 180 μg/mL, whereas α-T had no effect at concentrations up to 1000 μg/mL as measured by incorporation of [3H] thymidine. The effects of TRF and α-T also were tested in longer-term growth experiments, using concentrations of 180 and 500 μg/mL. We found that TRF inhibited the growth of these cells by 50%, whereas α-T did not. Their effect on the ability of these cells to form colonies also was studied, and it was found that TRF inhibited PE, whereas α-T had no effect. These results suggest that the inhibition is due to the presence of tocotrienols in TRF rather than α-T.
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| A comparison of tocopherol and tocotrienol for the chemoprevention of chemically induced rat mammary tumors.
Gould, M.N., et.al (1991). Am J Clin Nutr. |
Two forms of vitamin E, tocopherol and tocotrienol, were tested for chemopreventive activity in two chemically induced rat mammary-tumor models. When mammary tumors were induced by 7,12-dimethylbenz(a)anthracene (DMBA, 50 mg/kg), only the tocotrienol group had a statistically significant increase in tumor latency. There was no effect of either compound on tumor multiplicity. When tumors were induced by N-nitrosomethylurea (NMU, 30 mg/kg), neither analogue of vitamin E modified latency, whereas tocotrienol increased tumor multiplicity. In summary, neither vitamin analog had a major impact on mammary-tumor development after tumor induction with either DMBA or NMU.
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