General Health

Published Human Studies / Cohort Studies
Article	Study objective/ findings
Tocotrienol supplementation suppressed bone resorption and oxidative stress in postmenopausal osteopenic women: a 12-week randomized double-blinded placebo-controlled trial. Shen et al., (2018). Osteoporosis International.	To evaluate the effect of 12 weeks supplementation of tocotrienol on bone health, 89 postmenopausal women with osteopenia were randomly assigned into 3 groups: (1) placebo, (2) low TT (430 mg TT/day, 70% purity), and (3) high TT (860 mg TT/day, 70% purity). There were a decreases in serum BALP level and significant decreases in urine NTX levels, serum sRANKL, sRANKL/OPG ratio, and urine 8-OHdG concentrations and a significant increase in BALP/NTX ratio due to tocotrienol supplementation.

Published Human Studies / Cohort Studies

Article

Study objective/ findings

Tocotrienol supplementation suppressed bone resorption and oxidative stress in postmenopausal osteopenic women: a 12-week randomized double-blinded placebo-controlled trial.

Shen et al., (2018). Osteoporosis International.

To evaluate the effect of 12 weeks supplementation of tocotrienol on bone health, 89 postmenopausal women with osteopenia were randomly assigned into 3 groups: (1) placebo, (2) low TT (430 mg TT/day, 70% purity), and (3) high TT (860 mg TT/day, 70% purity). There were a decreases in serum BALP level and significant decreases in urine NTX levels, serum sRANKL, sRANKL/OPG ratio, and urine 8-OHdG concentrations and a significant increase in BALP/NTX ratio due to tocotrienol supplementation.

In- vivo / Animal Studies
Article	Study objective/ findings
Tocotrienol-rich fraction supplementation reduces hyperglycemia-induced skeletal muscle damage through regulation of insulin signaling and oxidative stress in type 2 diabetic mice Lee, H., Lim, Y., (2018). J Nutr Biochem.	In this study, the effect of palm tocotrienol-rich fraction (TRF) supplementation on skeletal muscle damage in diabetic mice is investigated. Normal control (CON) and diabetic control (DMC) groups were administrated with olive oil, while TRF treatment groups were administrated with TRF (dissolved in olive oil) at low dose (100 mg/kg BW, LT) or high dose (300 mg/kg BW, HT) by oral gavage for 12 weeks. TRF supplementation ameliorated muscle atrophy, plasma insulin concentration and homeostatic model assessment estimated insulin resistance in diabetic mice.
Comparison between tocotrienol and omeprazole on gastric growth factors in stress-exposed rats Azlina, M. F. N., et.al (2017). World J Gastroenterol.	The gastroprotective effect of tocotrienol was investigated and compared with omeprazole (a common drug used for to decrease excess stomach acid). Tocotrienol shown to have the similar protective effect as omeprazole, but through multiple protection mechanism which are by increasing expression of bFGF, TGF-α and VEGF in stress-induced gastric ulcer rat model.
γ-Tocotrienol suppresses growth and sensitises human colorectal tumours to capecitabine in a nude mouse xenograft model by down-regulating multiple molecules. Prasad, S., et.al (2016). Br J Cancer.	Both in vitro and in vivo effects of γ-tocotrienol (γ-T3) alone and in combination with capecitabine are investigated. Apoptosis and cytotoxicity assays were performed by MTT and FACS analysis, whereas expression of proteins was investigated using western blotting and immunohistochemistry. The findings suggest that γ-T3 inhibited the growth of human CRC and sensitised CRC to capecitabine by regulating proteins linked to tumourigenesis.
Tocotrienol-Rich Fraction (TRF) Suppresses the Growth of Human Colon Cancer Xenografts in Balb/C Nude Mice by the Wnt Pathway Zhang, J-S., et.al (2015). PLos One.	In this study, after feeding with tocotrienol-rich fraction from palm oil (TRF) for 2 weeks, Balb/c nude mice were inoculated human colon SW620 cancer cell and then continued to feed TRF for 4 weeks. At termination of experiments, xenografts were removed and determined the expression of Wnt-pathways related protein by immunohistochemistry or western blotting. Liver tissues were homogenated for determining the levels of antioxidative enzymes activity or malondialdehyde (MDA). The results showed that TRF significantly inhibited the growth of xenografts in nude mice. TRF also affected the activity of antioxidative enzymes in the liver tissue of mice. These changes were partly contributed to activation of wnt pathways or affecting their related protein. Thus, these finding suggested that the potent anticancer effect of TRF is associated with the regulation of Wnt signal pathways.
Vitamin E δ-tocotrienol prolongs survival in the LSL KrasG12D/+; LSLTrp 53R172H/+;Pdx-1-Cre (KPC) transgenic mouse model of pancreatic cancer. Husain, K, et.al (2013). Cancer Prev Res (Phila).	The effects of VEDT (vitamin E delta-tocotrienol) and the combination of VEDT and gemcitabine in the KPC mice. KPC mice were randomized into four groups: (i) vehicle [olive oil, 1.0 mL/kg per os twice a day and PBS 1.0 mL/kg intrapertoneally (i.p.) twice a week], (ii) gemcitabine (100 mg/kg i.p. twice a week), (iii) VEDT (200 mg/kg per os twice a day), and (iv) gemcitabine + VEDT were investigated. These results strongly support further investigation of VEDT alone and in combination with gemcitabine for pancreatic cancer prevention and treatment.
First evidence that gamma-tocotrienol inhibits the growth of human gastric cancer and chemosensitizes it to capecitabine in a xenograft mouse model through the modulation of NF-κβ pathway. Manu, K. A., et.al (2012). Clin Cancer Res.	The effect of γ-tocotrienol on proliferation of gastric cancer cell lines was examined by mitochondrial dye uptake assay, apoptosis by esterase staining, NF-κB activation by DNA-binding assay, and gene expression by Western blotting. The effect of γ-tocotrienol on the growth and chemosensitization was also examined in subcutaneously implanted tumors in nude mice. The results show that γ-tocotrienol can potentiate the effects of capecitabine through suppression of NF-κB-regulated markers of proliferation, invasion, angiogenesis, and metastasis.
Novel tocotrienol-entrapping vesicles can eradicate solid tumors after intravenous administration. Fu, J.Y., et.al. (2011). J Control Release.	The therapeutic potential of tocotrienol, a vitamin E extract with anti-cancer properties, is hampered by its failure to specifically reach tumors after intravenous administration. In this work, we demonstrated that novel transferrin-bearing, tocopheryl-based multilamellar vesicles entrapping tocotrienol significantly improved tocotrienol uptake by cancer cells overexpressing transferrin receptors. This led to a dramatically improved therapeutic efficacy in vitro, ranging from 17-fold to 72-fold improvement depending on the cell lines, compared to the free drug. In vivo, the intravenous administration of this novel tocotrienol formulation led to complete tumor eradication for 40% of B16-F10 murine melanoma tumors and 20% of A431 human epidermoid carcinoma tumors. Animal survival was improved by more than 20 days compared to controls, for the two tumor models tested. These therapeutic effects, together with the lack of toxicity, potentially make transferrin-bearing vesicles entrapping tocotrienol a highly promising therapeutic system as part as an anti-cancer therapeutic strategy.
Simultaneous induction of non-neoplastic and neoplastic lesions with highly proliferative hepatocytes following dietary exposure of rats to tocotrienols for 2 years. Tasaki, M., et.al (2009). Arch Toxicol.	It was recently shown that 1-year chronic exposure of rats to tocotrienol (TT) induced highly proliferative liver lesions, nodular hepatocellular hyperplasia (NHH), and independently increased the number of glutathione S-transferase placental form (GST-P)-positive hepatocytes. Focusing attention on the pathological intrinsic property of NHH, a 104-week carcinogenicity study was performed in male and female Wistar Hannover rats given TT at concentrations of 0, 0.4 or 2% in the diet. The high-dose level was adjusted to 1% in both sexes from week 51 because the survival rate of the high-dose males dropped to 42% by week 50. At necropsy, multiple cyst-like nodules were observed, as in the chronic study, but were further enlarged in size, which consequently formed a protuberant surface with a partly pedunculated shape in the liver at the high dose in both sexes. Unlike the chronic study, NHH was not always accompanied by spongiosis, and instead angiectasis was prominent in some nodules. However, several findings in the affected hepatocytes such as minimal atypia, no GST-P immunoreactivity and heterogeneous proliferation, implied that NHH did not harbor neoplastic characteristics from increased exposure despite sustained high cell proliferation. On the other hand, in the high-dose females, the incidence of hepatocellular adenomas was significantly higher than in the control. There was no TT treatment-related tumor induction in any other organs besides the liver. Thus, the overall data clearly suggested that NHH is successively enlarged by further long-term exposure to TT, but does not become neoplastic. In contrast, TT induces low levels of hepatocellular adenomas in female rats.
Induction of characteristic hepatocyte proliferative lesion with dietary exposure of Wistar Hannover rats to tocotrienol for 1 year. Tasaki, M., et.al (2008). Toxicology.	Tocotrienol is an antioxidant which has found commercial application as a food additive and health supplement all over the world. Since there have been no reports regarding toxicological effects of long-term exposure, we performed a 52-week chronic study using Wistar Hannover rats of both sexes given the compound at doses of 0, 0.08, 0.4 or 2% in powdered basal diet. Since 6 animals in the 2% male group died of hemorrhage of several organs by week 50, the maximum dose level was changed to 1% in both sexes for the last 2 weeks. Decrease of body weight gain was observed in the 2% males from week 5 and females from week 10, this persisting to the end of the study. With the high dose, prolongation of prothrombin time and increase of serum ALT in males, and increase of serum ALP in both sexes were observed with statistical significance. In male and female rats receiving 0.4% or less, there were no toxicological changes in any of the parameters examined. At necropsy, multiple cyst-like nodules on the liver surface were macroscopically pronounced in both sexes receiving 2%. On histopathological examination, hepatocellular nodules were evident with distortion of hepatic cords and compression of the surrounding tissue, almost all including areas of spongiosis hepatis. The constituent hepatocytes were immunohistochemically stained with proliferation cell nuclear antigen at high rates. Nevertheless, they did not exhibit overt atypia and the basic lobular architecture remained intact. Additionally, they were consistently negative for glutathione S-transferase placental form (GST-P). Accordingly, we propose the newly categorized but previously used name ‘nodular hepatocellular hyperplasia’, which may not necessarily have a neoplastic or regenerative nature. However, quantitative GST-P analysis of the liver sections overall showed numbers of GST-P foci in the high dose females to be significantly elevated as compared to the control value. Based on the present data demonstrating nodular liver lesions only at the high dose of both sexes, we conclude that the no-observed-adverse-effect level (NOAEL) is 0.4% (303 mg/kg/day for males, and 472 mg/kg/day for females).
In Vivo Angiogenesis Is Suppressed by Unsaturated Vitamin E, Tocotrienol Nakagawa, K., et.al (2007). American Society for Nutrition.	Antiangiogenic therapy using drugs and food components is a recognized strategy for the prevention of various angiogenesis-mediated disorders such as tumor growth, diabetic retinopathy, and rheumatoid arthritis. Our preliminary cell culture studies, using both bovine aortic endothelial cells and human umbilical vein endothelial cells (HUVEC) on screening for food-derived antiangiogenic compounds, showed tocotrienol (T3), an unsaturated version of vitamin E, to be a potential angiogenic inhibitor. We therefore investigaged the in vivo antiangiogenic properties of T3 using 2 well-characterized angiogenic models [mouse dorsal air sac (DAS) assay and the chick embryo chorioallantoic membrane (CAM) assay]. In the DAS assay, the increased neovascularization (angiogenesis index, 4.8 ± 0.6) in tumor cell–implanted mice was suppressed (angiogenesis index, 2.7 ± 0.6) by dietary supplementation of 10 mg T3-rich oil/d (equivalent to 4.4 mg T3/d). In the CAM assay, T3 (500–1000 μg/egg) inhibited new blood vessel formation on the growing CAM and increased the frequency of avascular zone (36–50%). To evaluate the antiangiogenic mechanism, we conducted cell-culture studies and found that T3 significantly reduced fibroblast growth factor -induced proliferation, migration, and tube formation in HUVEC (P < 0.05), with δ-T3 having the highest activity. Western blot analysis revealed that δ-T3 suppressed the phosphorylation of phosophoinositide-dependent protein kinase (PDK) and Akt, and increased the phosphorylation of apoptosis signal-regulating kinase and p38 in fibroblast growth factor-treated HUVEC, indicating that the antiangiogenic effects of T3 are associated with changes in growth factor-dependent phosphatidylinositol-3 kinase /PDK/Akt signaling as well as induction of apoptosis in endothelial cells. Our findings suggest that T3 has potential as a therapeutic dietary supplement for preventing angiogenic disorders, and therefore future clinical study will be required to evaluate the efficacy and safety of T3.
Tocotrienols potentiate lovastatin-mediated growth suppression in vitro and in vivo. McAnally, J.A., et.al (2007). Exp Biol Med.	3-Hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase is the rate-limiting enzyme in the mevalonate pathway that provides essential intermediates for the membrane anchorage and biologic functions of growth-related proteins. Contrary to preclinical studies showing the growth-suppressive activity of statins, competitive inhibitors of HMG CoA reductase, clinical application of statins in cancer is precluded by their lack of activity at levels prescribed for the prevention of cardiovascular disease and by their dose-limiting toxicities at high doses. The dysregulated and elevated HMG CoA reductase activity in tumors retains sensitivity to the isoprenoid-mediated posttranscriptional down-regulation, an action that complements the statin-mediated inhibition and may lead to synergistic impact of blends of isoprenoids and lovastatin on tumor HMG CoA reductase activity and consequently tumor growth. d-gamma- and d-delta-tocotrienols, vitamin E isomers containing an isoprenoid moiety, and lovastatin-induced concentration-dependent inhibition of the 48-hr proliferation of murine B16 melanoma cells with IC50 values of 20 +/- 3, 14 +/- 3, and 1.5 +/- 0.4 microM respectively. A blend of lovastatin (1 microM) and d-gamma-tocotrienol (5 microM) totally blocked cell growth, an impact far exceeding the sum of inhibitions induced by lovastatin (12%) and d-gamma-tocotrienol (8%) individually. Synergistic impact of these two agents was also shown in human DU145 prostate carcinoma and human A549 lung carcinoma cells. C57BL6 mice were fed diets supplemented with 12.5 mg lovastatin/kg body weight, 62.5 mg d-delta-tocotrienol/kg body weight, or a blend of both agents for 22 days following B16 cell implantation; only the latter had significantly lower tumor weight than those with no supplementation. Co-administration of isoprenoids that posttranscriptionally down-regulate tumor reductase may lower the effective dose of statins and offer a novel approach to cancer chemo-prevention and/or therapy.
Suppression of diethynitrosamine and 2-acetylaminofluorene-induced hepatocarcinogenesis in rats by tocotrienol-rich fraction isolated from rice bran oil. Iqbal, J.et.al (2004). Eur J Cancer Prev.	The anticancer efficacy of tocotrienol-rich fraction (TRF) was evaluated during diethylnitrosamine (DEN)/2-acetylaminofluorene (AAF)-induced hepatocarcinogenesis in male Sprague-Dawley rats. Long-term intake of TRF could reduce cancer risk by preventing hepatic lipid peroxidation and protein oxidation damage due to its antioxidant actions.
Suppresssion of 7,12-dimethylben[alpha]anthracene-induced carcinogenesis and hypercholetsrolaemia in rats by tocotrienol-rich fraction isolated from rice bran oil. Iqbal, J., et.al (2003). Eur J Cancer Prev.	The anti-tumour and anti-cholesterol impacts of tocotrienol-rich fraction (TRF) were investigated in rats treated with the chemical carcinogen 7,12-dimethylbenz [alpha]anthracene (DMBA), which is known to induce mammary carcinogenesis and hypercholesterolaemia. DMBA administration to rats was associated with the appearance of multiple tumours on mammary glands after 6 months. Results indicate that TRF has potent anti-cancer and anti-cholesterol effects in rats.
Long-term administration of tocotrienols and tumor-maker enzyme activities during hepatocarcinogenesis in rats. Rahmat, A., et.al (1993). Nutrition.	The effects of long-term administration of tocotrienol on hepatocarcinogenesis in rats induced by diethylnitrosamine (DEN) and 2-acetylaminofluorene (AAF) were investigated by determining the activities of gamma-glutamyl transpeptidase (GGT), alkaline phosphatase (ALP), glutathione S-transferases (GSTs), and glutathione (GSH) levels in blood and liver. Twenty-eight male 7- to 8-wk-old Rattus norwegicus rats, weighing 120-160 g, were used in this study. The rats were divided into four treatment groups: a control group on a basal diet, a group fed a basal diet supplemented with tocotrienol (30 mg/kg food), a group treated with DEN/AAF, and a group treated with DEN/AAF and fed a diet supplemented with tocotrienol (30 mg/kg food). Blood was collected monthly, and GGT, ALP, and GSH levels were determined. The rats were killed after 9 mo, and the livers were examined morphologically. Grayish white nodules (2/liver) were found in all the DEN/AAF-treated rats (n = 10), but only one of the rats treated with DEN/AAF and supplemented with tocotrienol (n = 6) had liver nodules. A significant increase in the level of blood and liver GSH, ALP, and GGT activities was observed in the DEN/AAF-treated rats. Liver GSTs were similarly increased with DEN/AAF treatment. Tocotrienol supplementation attenuated the impact of the carcinogens in the rats.
Effect of tocotrienols on hepatocarcinogenesis induced by 2-acetylaminofluorene in rats. Ngah, W.Z., et.al (1991). Am J Clin Nutr.	The effects of tocotrienols on hepatocarcinogenesis in rats fed with 2-acetylaminofluorene (AAF) were followed morphologically and histologically for a period of 20 wk. No differences between treated and control rats in the morphology and histology of their livers was observed. Cell damage was extensive in the livers of AAF-treated rats but less extensive in the AAF-tocotrienols-treated rats when compared with normal and tocotrienols-treated rats. 2-Acetylaminofluorene significantly increases the activities of both plasma and liver microsomal gamma-glutamyltranspeptidase (GGT) and liver microsomal UDP-glucuronyltransferase (UDP-GT). Tocotrienols administered together with AAF significantly decrease the activities of plasma GGT after 12 and 20 wk (P less than 0.01, P less than 0.002, respectively) and liver microsomal UDP-GT after 20 wk (P less than 0.02) when compared with the controls and with rats treated only with tocotrienols. Liver microsomal GGT also showed a similar pattern to liver microsomal UDP-GT but the decrease was not significant. These results suggest that tocotrienols administered to AAF-treated rats reduce the severity of hepatocarcinogenesis.

In-vitro Studies
Article	Study objectives/ findings
Tocotrienol-Rich Fraction (TRF) Treatment Promotes Proliferation Capacity of Stress-Induced Premature Senescence Myoblasts and Modulates the Renewal of Satellite Cells: Microarray Analysis Lim, J.J., et al., (2019). Oxid Med Cell Longev	Little is known on the mechanism of tocotrienol towards the oxidative stress-associated senescence in myoblasts, especially human myoblasts. This study aimed to reveal the novel insight into the differential gene expression by stress-induced human myoblasts with the treatment of TRF. The study findings proposed that TRF treatment not only promotes the proliferation capacity of SIPS myoblasts through regulation of the ErbB signalling pathway (upregulation of expression of EREG, SHC1, and SHC3) and FoxO signalling pathway (downregulation of expression of MSTN and SMAD3). At the same time, TRF treatment is proposed to modulate the renewal of satellite cells through regulation of p53 signalling (upregulation of RRM2B and SESN1), cell cycle, Wnt signalling pathway, and expression of MRF.
Gamma-tocotrienol profoundly alters sphingolipids in cancer cells by inhibition of dihydroceramide desaturase and possibly activation of sphingolipid hydrolysis during prolonged treatment. Jang, Y., et.al (2017). J Nutr Biochem.	Vitamin E gamma-tocotrienol (γ-T3) is known to have anticancer effects, but mechanisms underlying these actions are not clear. Gamma-tocotrienol (γ-T3) induced marked changes of sphingolipids including rapid elevation of dihydrosphingosine and dihydroceramides (dhCers) in various types of cancer cells. The elevation of dihydrosphingolipids coincided with increased cellular stress, as indicated by JNK phosphorylation, and was prior to any sign of induction of apoptosis. Chemically blocking de novo synthesis of sphingolipids partially counteracted γ-T3-induced apoptosis and autophagy. This study demonstrates that γ-T3 altered sphingolipid metabolism by inhibiting DEGS activity and possibly by activating SM hydrolysis during prolonged treatment in cancer cells.
Synthesis, characterization and in-vitro anti-tumor activity of polyethylene glycol (350 and 1000) succinate derivatives of the tocopherol and tocotrienol isomers of vitamin E. Abu-Fayyad & Nazzal. (2017). Int J Pharm.	The current work was therefore undertaken to synthesize and characterize the water soluble polyethylene glycol (PEG 350 and 1000) derivatives of T and T₃. The results from the current work have demonstrated the feasibility of synthesizing PEGylated conjugates of vitamin E isomers and highlighted the potential use of these conjugates in drug delivery as functional and safer excipients especially for γ-T₃PGS 1000 and δ-T₃PGS 1000 conjugate.
A Combination of δ-Tocotrienol and Ferulic Acid Synergistically Inhibits Telomerase Activity in DLD-1 Human Colorectal Adenocarcinoma Cells. Eitsuka, T, et.al (2016). J Nutr Sci Vitaminol (Tokyo)	The synergistic suppressive effects of the combination of delta-tocotrienol (δ-T3) and fatty acid (FA) on telomerase activity in DLD-1 human colorectal adenocarcinoma cells. FA significantly increased the intracellular concentration of δ-T3, suggesting that FA improved the bioavailability of δ-T3, thereby increasing the inhibitory potency of δ-T3 on telomerase. FA may be a promising candidate for augmenting the anti-cancer activity of δ-T3.
Vitamin E δ-tocotrienol triggers endoplasmic reticulum stress-mediated apoptosis in human melanoma cells. Montagnani, M. M., et.al (2016). Sci Rep.	The effects of delta-tocotrienol (δ-T3) on melanoma cell growth and the involvement of the endoplasmic reticulum (ER) stress in this activity. In conclusion, δ-T3exerts a proapoptotic activity on melanoma cells, through activation of the ER stress-related pathways. δ-T3might represent an effective option for novel chemopreventive/therapeutic strategies for melanoma.
Enhanced effectiveness of tocotrienol-based nano-emulsified system for topical delivery against skin carcinomas. Pham, J., et.al (2016). Drug Deliv.	The emphasis of the present work is to develop a pharmaceutically scalable, low-energy nano-emulsification approach for optimized incorporation of T3-rich palm oil (Tocomin®), possessing anticancer activity as a potential cutaneous delivery platform for adjunctive therapy of skin carcinomas, either alone or in combination with other chemotherapeutic agents. Adopted hybrid nano-emulsification of Tocomin®, correlated with highest preservation of DPPH-radical scavenging capacity of active T3 in prototype formulation, Tocomin®-NE, which effectively permeated diffusion cell membranes 4-folds higher than propyleneglycol (PG)-admixed Tocomin® control. Against two different cell models of human cutaneous carcinoma, Tocomin®-hybrid NE demonstrated significantly stronger cytotoxic profiles (p ≤ 0.01), visible in both concentration- and time- dependent manners, with at least 5-folds lower IC50 values, compared to those estimated for the closest Tocomin®-control. The proposed hybrid nano-emulsified formulation of Tocomin® provides simple and stable delivery platform, for effective topical application against keratinocyte tumors.
Synergistic cytotoxic effects of combined δ-tocotrienol and jerantinine B on human brain and colon cancers Abubakar, I.B., et.al (2016). J Ethnopharmacol.	The present study was undertaken to study the mechanism of apoptosis induction in human glioblastoma (U87MG) and colorectal adenocarcinoma (HT-29) cancer cells by a novel indole alkaloid, jerantinine B isolated from T. corymbosa, δ-tocotrienol and the combined low-dose treatments of δ-tocotrienol with IC20 dose of jerantinine B. The present study demonstrated the mechanism for cytotoxic potency of δ-tocotrienol and jerantinine B against U87MG and HT-29 cells. Furthermore, combined low-dose treatments induced concurrent synergistic inhibition of cancer cell growth with concomitant dose reduction thus minimizing toxicity to normal cells and improving potency of δ-tocotrienol and jerantinine B.
γ-Tocotrienol upregulates aryl hydrocarbon receptor expression and enhances the anticancer effect of baicalein. Yamashita, S., et.al (2016). Biochem Biophys Res Commun.	The effects of naturally abundant γ-T3 on FFS-related gene expressions in melanoma using a DNA chip were investigated. The results showed that γ-T3 increased the expression level of aryl hydrocarbon receptor (AhR), a sensing molecule to plant polyphenol baicalein. The co-treatment with γ-T3 and baicalein enhanced the anti-proliferative activity of baicalein, accompanied by the downstream events of AhR-activation induced by baicalein. These data suggest that γ-T3 upregulates AhR expression and enhances its sensitivity to baicalein.
Cytotoxicity Induced by a Redox-silent Analog of Tocotrienol in Human Mesothelioma H2452 Cell Line via Suppression of Cap-dependent Protein Translation. Sato, A., et.al (2016). Anticancer Res.	Tocotrienol-induced potent cytotoxicity aganist MM (malignant mesothelioma) cells were investigated. Overall, T3E had a cytotoxic effect on MM cells via disruption of the activated cap-dependent translation complex through inactivation of RAS.
Gamma tocotrienol targets tyrosine phosphatase SHP2 in mammospheres resulting in cell death through RAS/ERK pathway Gu, W., et.al (2015). BMC Cancer.	There is increasing evidence supporting the concept of cancer stem cells (CSCs), which are responsible for the initiation, growth and metastasis of tumors. CSCs are thus considered the target for future cancer therapies. To achieve this goal, identifying potential therapeutic targets for CSCs is essential. SHP1 (Src homology 2 domain-containing phosphatase 1) and SHP2 are potential therapeutic targets for breast CSCs and gamma-T3 is a promising natural drug for future breast cancer therapy.
Differentiation and apoptosis induction by lovastatin and γ-tocotrienol in HL-60 cells via Ras/ERK/NF-κB and Ras/Akt/NF-κB signaling dependent down-regulation of glyoxalase 1 and HMG-CoA reductase. Chen, C.C., et.al (2015). Cell Signal.	In this study, treatment of HL-60 cells with lovastatin induced characteristic apoptosis in a dose-dependent manner. We demonstrated that lovastatin treatment inhibited Ras and Raf protein translocation to cell membrane and eliminated the phosphorylation of the downstream effectors Akt and ERK, and the subsequent NF-κB translocation into nucleus. These results suggest that both lovastatin and gamma-tocotrienol could induce differentiation and followed by apoptosis.
Gamma-tocotrienol and 6-gingerol in combination synergistically induce cytotoxicity and apoptosis in HT-29 and SW837 human colorectal cancer cells. Yusof, K.M., et.al (2015). Molecules.	The objective of this study was to determine the anti-proliferative effect of γ-tocotrienol (γ-T3) and 6-gingerol (6G) in combination by evaluating apoptosis and active caspase-3 in HT-29 and SW837 colorectal cancer cells. In conclusion, γ-T3 and 6G when used in combination act synergistically increasing cytotoxicity and apoptosis in cancer cells.
δ-Tocotrienol treatment is more effective against hypoxic tumor cells than normoxic cells: potential implications for cancer therapy. Shibata, A., et.al (2015). J Nutr Biochem.	In this study, we demonstrated that δ-tocotrienol (δ-T3) could be used as a novel anticancer agent against human colorectal adenocarcinoma (DLD-1) cells under both normoxic and hypoxic conditions. These data suggest that tocotrienols are potent antitumor agents capable of inducing apoptosis and inhibiting angiogenesis under both hypoxic and normoxic conditions. Tocotrienols could have significant therapeutic potential in the clinical treatment of tumors.
δ-Tocotrienol Induces Human Bladder Cancer Cell Growth Arrest, Apoptosis and Chemosensitization through Inhibition of STAT3 Pathway Ye, C.X., et.al (2015). PLoS One.	Vitamin E intake has been implicated in reduction of bladder cancer risk. However, the mechanisms remain elusive. Here we reported that δ-tocotrienol (δ-T3), one of vitamin E isomers, possessed the most potent cytotoxic capacity against human bladder cancer cells, compared with other Vitamin E isomers. δ-T3 inhibited cancer cell proliferation and colonogenicity through induction of G1 phase arrest and apoptosis. Western blotting assay revealed that δ-T3 increased the expression levels of cell cycle inhibitors (p21, p27), pro-apoptotic protein (Bax) and suppressed expression levels of cell cycle protein (Cyclin D1), anti-apoptotic proteins (Bcl-2, Bcl-x_L and Mcl-1), resulting in the Caspase-3 activation and cleavage of PARP. Moreover, the δ-T3 treatment inhibited ETK phosphorylation level and induced SHP-1 expression, which was correlated with downregulation of STAT3 activation. In line with this, δ-T3 reduced the STAT3 protein level in nuclear fraction, as well as its transcription activity. Knockdown of SHP-1 partially reversed δ-T3-induced cell growth arrest. Importantly, low dose of δ-T3 sensitized Gemcitabine-induced cytotoxic effects on human bladder cancer cells. Overall, our findings demonstrated, for the first time, the cytotoxic effects of δ-T3 on bladder cancer cells and suggest that δ-T3 might be a promising chemosensitization reagent for Gemcitabine in bladder cancer treatment.
Alkaloid extracts of Ficus species and palm oil-derived tocotrienols synergistically inhibit proliferation of human cancer cells Abubakar, I.B., et.al (2014). Nat Prod Res.	This study explored the potential synergism of antiproliferative effects induced by individual alkaloid extracts of Ficus fistulosa, Ficus hispida and Ficus schwarzii combined with δ- and γ-tocotrienols against human brain glioblastoma (U87MG), lung adenocarcinoma (A549) and colorectal adenocarcinoma (HT-29) cells. Cell viability and morphological results demonstrated that extracts containing a mixture of alkaloids from the leaves and bark of F. schwarzii inhibited the proliferation of HT-29 cells, whereas the alkaloid extracts of F. fistulosa inhibited the proliferation of both U87MG and HT-29 cells and showed synergism in combined treatments with either δ- or γ-tocotrienol resulting in 2.2–34.7 fold of reduction in IC₅₀ values of tocotrienols. The observed apoptotic cell characteristics in conjunction with the synergistic antiproliferative effects of Ficus species-derived alkaloids and tocotrienols assuredly warrant future investigations towards the development of a value-added chemotherapeutic regimen against cancers.
Gamma-tocotrienol and hydroxy-chavicol synergistically inhibits growth and induces apoptosis of human glioma cells Abdul Rahman, A., et.al (2014). BMC Complement Altern Med.	The interaction of GTT (gamma-tocotrienol) and HC (hydroxy-chavicol) and its mode of cell death on glioma cell lines was investigated. GTT or HC alone and in combination were tested for cytotoxicity on glioma cell lines 1321N1 (Grade II), SW1783 (Grade III) and LN18 (Grade IV) by [3-(4,5-dimethylthiazol-2- yl)-5-(3-carboxymethoxy-phenyl)-2-(4-sulfophenyl)- 2H- tetrazolium, inner salt] MTS assay. The interactions of each combination were evaluated by using the combination index (CI) obtained from an isobologram. In conclusion, combined treatments with sub-effective doses of GTT and HC resulted in synergistic inhibition of cell proliferation through the induction of apoptosis of human glioma cells in vitro.
A redox-silent analogue of tocotrienol inhibits cobalt(II) chloride-induced VEGF expression via Yes signaling in mesothelioma cells. Sato, A., et.al (2014). Biol Pharm Bull.	In this study we examine whether tocotrienol (T3) can suppress the secretion of vascular endothelial growth factor (VEGF) in human malignant mesothelioma (MM) cells exposed to mimic hypoxia by cobalt chloride (CoCl2). We found that CoCl2-induced hypoxia treatment leads to increased up-regulated hypoxia-inducible factor-2α (HIF-2α) and subsequently induced the secretion of VEGF in MM cells. This up-regulation activation mainly depended on the activation of Yes, a member of the Src family of kinases. Treatment of hypoxic MM cells with T3E effectively inhibited the secretion of VEGF, On the other hand, T3E inhibited CoCl2-induced gene expression of VEGF due to the inactivation of Yes/HIF-2α signaling. These data suggest that Yes/HIF2-α/VEGF could be a promising therapeutic target of T3E in MM cells.
Synergistic effect of combined treatment with gamma-tocotrienol and statin on human malignant mesothelioma Tuerdi, G., et.al (2013). Cancer Lett.	The present study is the first to demonstrate the synergetic effect of statins (atorvastatin and simvastatin) and gamma-tocotrienol (γ-T3) on human malignant mesothelioma (MM). Statin + γ-T3 combinations induced greater cell growth inhibition more than each single treatment via inhibition of mevalonate pathway, a well-known target of both γ-T3 and statins. γ-T3 was necessary for endoplasmic reticulum stress markers CHOP and GRP78, whereas an intrinsic apoptotic marker, caspase 3 activation was induced only in the presence of statins. Overall, the combination of γ-T3 and statins could be useful for MM therapy and functions in a complementary style.
Optimization of tocotrienols as antiproliferative and antimigratory leads. Behery, F.A., et.al (2013). Eur J Med Chem.	The vitamin E family members γ- and δ-tocotrienols (2 and 3, respectively) are known natural products with documented anticancer activities. Redox-silent structural modifications, such as esterification, etherification and carbamoylation, of 2 and 3 significantly enhanced their anticancer activities. However, hit-to-lead optimization of tocotrienols and their analogs was yet to be reported at the outset of the project described herein. Subjecting the chroman ring of 2 and 3 to the electrophilic substitution reactions, namely, Mannich and Lederer-Manasse procedures, afforded 42 new products. These included the 3,4-dihydro-1,3-oxazines 3-29 and 35-44, Mannich bases 30-31, and the hydroxymethyl analogs 32-34. Of these, the δ-tocotrienol analogs 8, 11, 18, 24, 25, 27, and 40 inhibited the proliferation of the highly metastatic +SA mammary epithelial cancer cell line, with IC(50) values in the nanomolar (nM) range. In NCI’s 60 human tumor cell line panel, 8, 17, 38, and 40 showed antiproliferative activity, with nM GI(50) values. The δ-tocotrienol analogs 10 and 38 inhibited the migration of the highly metastatic human breast cancer cell line MDA-MB-231 with IC(50) values of 1.3 and 1.5 μM, respectively, in the wound-healing assay. A dose of 0.5 mg/day for 14 days of one of the active analogs, 30, significantly slowed the growth of +SA mammary tumors in the syngeneic BALB/c mouse model, compared to the vehicle- and the parent γ-tocotrienol-treated control groups. Electrophilic substitution reactions promoted tocotrienols to lead level and can enable their future use to control metastatic breast malignancies.
Vitamin E δ-tocotrienol induces p27(Kip1)-dependent cell-cycle arrest in pancreatic cancer cells via an E2F-1-dependent mechanism. Hodul, P.J., et.al (2013). PLoS One.	δ-tocotrienol exerted significant cell growth inhibition pancreatic ductal cancer (PDCA) cells without affecting normal human pancreatic ductal epithelial cell growth. We also showed that δ-tocotrienol-induced growth inhibition occurred concomitantly with G(1) cell-cycle arrest and increased p27(Kip1) nuclear accumulation. This finding is significant considering that loss of nuclear p27(Kip1) expression is a well-established adverse prognostic factor in PDCA. Furthermore, δ-tocotrienol inactivated RAF-MEK-ERK signaling, a pathway known to suppress p27(Kip1) expression. To determine whether p27(Kip1) induction is required for δ-tocotrienol inhibition of PDCA cell proliferation, we stably silenced the CDKN1B gene, encoding p27(Kip1), in MIAPaCa-2 PDCA cells and demonstrated that p27(Kip1) silencing suppressed cell-cycle arrest induced by δ-tocotrienol. Furthermore, δ-tocotrienol induced p27(Kip1) mRNA expression but not its protein degradation. p27(Kip1) gene promoter activity was induced by δ-tocotrienol through the promoter’s E2F-1 binding site, and this activity was attenuated by E2F-1 depletion using E2F-1 small interfering RNA. Finally, decreased proliferation, mediated by Ki67 and p27(Kip1) expression by δ-tocotrienol, was confirmed in vivo in a nude mouse xenograft pancreatic cancer model. Our findings reveal a new mechanism, dependent on p27(Kip1) induction, by which δ-tocotrienol can inhibit proliferation in PDCA cells, providing a new rationale for p27(Kip1) as a biomarker for δ-tocotrienol efficacy in pancreatic cancer prevention and therapy.
Delta-tocotrienol suppresses Notch-1 pathway by upregulating miR-34a in nonsmall cell lung cancer cells Ji, X., et.al (2012). Int J Cancer.	MicroRNAs (miRNAs) are small noncoding RNAs that play critical roles in regulating various cellular functions by transcriptional silencing. miRNAs can function as either oncogenes or tumor suppressors (oncomirs), depending on cancer types. In our study, using miRNA microarray, we observed that downregulation of the Notch-1 pathway, by delta-tocotrienol, correlated with upregulation of miR-34a, in nonsmall cell lung cancer cells (NSCLC). Moreover, re-expression of miR-34a by transfection in NSCLC cells resulted in inhibition of cell growth and invasiveness, induction of apoptosis and enhanced p53 activity. Furthermore, cellular mechanism studies revealed that induction of miR-34a decreased the expression of Notch-1 and its downstream targets including Hes-1, Cyclin D1, Survivin and Bcl-2. Our findings suggest that delta-tocotrienol is a nontoxic activator of mir-34a which can inhibit NSCLC cell proliferation, induce apoptosis and inhibit invasion, and thus offering a potential starting point for the design of novel anticancer agents.
Mechanisms mediating the synergistic anticancer effects of combined gamma-tocotrienol and sesamin treatment. Akl, M.R., et.al (2012). Planta Med.	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.
Inhibition of Mitochondrial Cytochrome c Release and Suppression of Caspases by Gamma-Tocotrienol Prevent Apoptosis and Delay Aging in Stress-Induced Premature Senescence of Skin Fibroblasts Makpol, S., et.al (2012). Oxid Med Cell Longev.	In this study, we determined the molecular mechanism of γ-tocotrienol (γ-T3) in preventing cellular aging by focusing on its anti-apoptotic effect in stress-induced premature senescence (SIPS) model of human diploid fibroblasts (HDFs). The findings suggested that γ-T3 inhibits apoptosis by modulating the upstream apoptosis cascade, causing the inhibition of cytochrome crelease from the mitochondria with concomitant suppression of caspase-9 and caspase-3 activation. In conclusion, GTT delays cellular senescence of human diploid fibroblasts through the inhibition of intrinsic mitochondria-mediated pathway which involved the regulation of pro- and anti-apoptotic genes and proteins.
Gamma-tocotrienol inhibits cell viability through suppression of β-catenin/Tcf signaling in human colon carcinoma HT-29 cells. Xu, H., et.al (2011). J Nutr Biochem.	In this study, the mechanisms underlying γ-tocotrienol-mediated growth inhibition of human carcinoma HT-29 cells were further investigated, especially in correlation with the involvement of β-catenin/T-cell factor (Tcf) signaling pathway. The results demonstrated that γ-tocotrienol-inhibited growth and -induced apoptosis in HT-29 cells were accompanied by significant inhibition of β-catenin/Tcf signaling. Blocking the expression of β-catenin with small interfering RNA significantly suppressed the ability of γ-tocotrienol to reduce viability and induce apoptosis in HT-29 cells. Thus, our data suggested that γ-tocotrienol exerts its anticancer activity through β-catenin/Tcf signaling, and β-catenin is a target for γ-tocotrienol in the Wnt/β-catenin signaling pathway.
Delta-tocotrienol augments cisplatin-induced suppression of non-small cell lung cancer cells via inhibition of the Notch-1 pathway. Ji, X., et.al (2012). Anticancer Res.	Non-small cell lung cancer (NSCLC), accounting for 80% of lung cancers, is the leading cause of all cancer deaths. Previously, we demonstrated that delta-tocotrienol inhibits NSCLC cell proliferation, invasion and induces apoptosis by down-regulation of the Notch-1 signaling pathway. The objective of this study was to investigate whether delta-tocotrienol, could enhance the anticancer effects of cisplatin. Treatment with a combination of delta-tocotrienol and cisplatin resulted in a dose-dependent, significant inhibition of cell growth, migration, invasiveness, and induction of apoptosis in NSCLC cells, as compared to the single agents. This was associated with a decrease in NF-κB DNA binding activity, decrease in Notch-1, Hes-1, Bcl-2 and increase in cleaved Caspase-3 and PARP expressions. These results suggest that down-regulation of Notch-1, via inhibition of NF-κB signaling pathways by delta-tocotrienol and cisplatin, in combination, could provide a potential novel approach for tumor arrest in NSCLC, while lowering the effective dose of cisplatin.
Delta- and gamma-tocotrienols induce classical ultrastructural apoptotic changes in human T lymphoblastic leukemic cells. Wong, R. S.Y., et.al (2012). Microsc Microanal.	The present study investigated the effects of three isomers of tocotrienols (alpha, delta, and gamma) on a human T lymphoblastic leukemic cell line (CEM-SS). These findings were confirmed biochemically by the demonstration of phosphatidylserine externalization via flow cytometry analysis. This is the first study showing classical ultrastructural apoptotic changes induced by delta- and gamma-tocotrienols in human T lymphoblastic leukemic cells.
Gamma-tocotrienol inhibits angiogenesis of human umbilical vein endothelial cell induced by cancer cell Li, Y., et.al (2011). J Nutr Biochem.	The purpose of this study was to evaluate the inhibitory effect and possible mechanism of gamma-tocotrienol on tumor angiogenesis. The results showed that gamma-tocotrienol (10-40 mumol/L) significantly suppressed proliferation, migration and tube formation of human umbilical vein endothelial cells (HUVECs) induced by SGC-7901 cell conditioned medium in a dose-dependent manner. gamma-Tocotrienol (800-1200 mug/egg) also inhibited new blood vessel formation on the growing chick embryo chorioallantoic membrane in a dose-dependent manner. Moreover, the inhibitory effects of gamma-tocotrienol on HUVECs were correlated with inducing the apoptosis and arresting cell cycle at the G(0)/G(1) phase at a dose of 40 mumol/L gamma-tocotrienol. In addition, gamma-tocotrienol inhibited angiogenesis in HUVECs by down-regulation of beta-catenin, cyclin D1, CD44, phospho-VEGFR-2 and MMP-9. The antiangiogenic effects of gamma-tocotrienol on HUVECs may be attributable to regulation of Wnt signaling by decreasing beta-catenin expression. Thus, our results suggest that gamma-tocotrienol has a potential chemopreventive agent via antiangiogenesis.
Inhibition of cell growth and induction of apoptosis in non-small cell lung cancer cells by delta-tocotrienol is associated with notch-1 down-regulation. Ji, X., et.al (2011). J Cell Biochem.	The objective of this study was to investigate whether delta-tocotrienol, a naturally occurring isoform of Vitamin E, inhibits non-small cell lung cancer cell growth via Notch signaling. Treatment with delta-tocotrienol resulted in a dose and time dependent inhibition of cell growth, cell migration, tumor cell invasiveness, and induction of apoptosis. Real-time RT-PCR and western blot analysis showed that antitumor activity by delta-tocotrienol was associated with a decrease in Notch-1, Hes-1, Survivin, MMP-9, VEGF, and Bcl-XL expression. In addition, there was a decrease in NF-κB-DNA binding activity. These results suggest that down-regulation of Notch-1, via inhibition of NF-κB signaling pathways by delta-tocotrienol, could provide a potential novel approach for prevention of tumor progression in non-small cell lung cancer.
A paraptosis-like cell death induced by delta-tocotrienol in human colon carcinoma SW620 cells is associated with the suppression of the Wnt signaling pathway. Zhang J.S., et.al (2011). Toxicology.	In this study, we focused on the effects of δ-tocotrienol and its possible mechanism on induction of death in human colon cancer SW620 cells. δ-Tocotrienol inhibited proliferation of SW620 cell in a dose-dependent manner. Our findings showed that δ-tocotrienol effectively induced paraptosis-like death in SW620 cells, correlated with the vacuolation that may be from welling and fusion of mitochondria and/or the endoplasmic reticulum (ER) as well as caspase-3 nonactivated. However, there were no changes in apoptosis based on flow cytometry analysis. Of being noted, δ-tocotrienol reduced the expression of β-catenin and wnt-1 proteins by about 50% at the highest dose (20μmol/L). δ-Tocotrienol also decreased cyclin D1, c-jun and MMP-7 protein levels in SW620 cells. Altogether, these data indicate that δ-tocotrienol induces paraptosis-like cell death, which is associated with the suppression of the Wnt signaling pathway. Thus, our findings may provide a novel application in treatment of human colon carcinoma.
Role of immunoregulatory transcription factors in differential immunomodulatory effects of tocotrienols. Wilankar, C., et.al (2011). Free Radic Biol Med.	This report describes novel immunomodulatory effects of tocotrienols in murine lymphocytes. γ-Tocotrienol (GT) was more effective in suppressing concanavalin A (Con A)-induced T cell proliferation and cytokine production compared to α-tocotrienol (AT) when present continuously in the culture. GT inhibited T cell activation markers and costimulatory molecule. GT modulated intracellular glutathione in lymphocytes, and the suppressive effects of GT could not be abrogated by thiol or nonthiol antioxidants, indicating a poor link between anti-inflammatory properties of tocotrienols and cellular redox status. It was also observed that GT suppressed Con A-induced activation of NF-κB, AP-1, and NF-κB-dependent gene expression. Cellular uptake studies with tocotrienols showed higher accumulation of GT compared to AT. Similar immunosuppressive effects of GT were also observed when administered to mice. In contrast, transient exposure of lymphocytes to GT (4 h) resulted in higher survival and proliferation of lymphocytes in vitro and in vivo in syngeneic and allogeneic hosts. This was attributed to the ability of GT to induce NF-κB, AP-1, and mTOR activation in lymphocytes upon transient exposure. Our results demonstrated that antioxidants such as tocotrienols may exhibit pleiotropic effects by activating multiple mechanisms in cells.
Gamma-tocotrienol induces apoptosis in human T cell lymphoma through activation of both intrinsic and extrinsic pathways. Wilankar, C., et.al (2011). Curr Pharm Des.	The potential of α-tocotrienol (AT) and γ-tocotrienol (GT) in inhibiting the proliferation of human T cell lymphoma Jurkat cells and elucidate the pathways involved in anti tumor effects of GT. GT but not AT inhibited proliferation and induced apoptosis in Jurkat cells in a dose dependent manner was investigated. Our results show antitumor effects of GT in human lymphoma cells via increased mitochondrial ROS generation and activation of both intrinsic and extrinsic apoptotic pathways.
Induction of apoptosis by gamma-tocotrienol in human cancer cell lines and leukemic blasts from patients: Dependency on Bid, cytochrome c and caspase pathway. Inoue, A., et.al (2011). Nutr Cancer.	Tocotrienol-induced apoptosis in human hematological cancer cell lines. α-, δ-, and γ-T3 induced concentration-dependent apoptosis, and γ-T3 demonstrated more effective induction than the other Toc3 derivatives in HL-60 cells. γ-T3 may have induced apoptosis by activation of the caspase cascade, cytochrome c (Cyt.c) release, Bid cleavage, and mitochondorial membrane depolarization in HL-60, NB-4, Raji, and SY-5Y cells. Furthermore, 10–30 μM γ-T3 showed cytotoxicity for leukemic cells from various patients regardless of lymphoblastic, myeloblastic, or relapsed leukemia, but the cytotoxic effect was weak in normal mononuclear cells, interestingly. γ-T3 may have a role in cancer prevention and potential for treating hematological malignancies.
γ-Tocotrienol is a novel inhibitor of constitutive and inducible STAT3 signalling pathway in human hepatocellular carcinoma: potential role as an antiproliferative, pro-apoptotic and chemosensitizing agent Rajendran, P., et.al (2011). Br J Pharmacol.	The effect of γ-tocotrienol on STAT3 activation, associated protein kinases and phosphatase, STAT3-regulated gene products, cellular proliferation and apoptosis in HCC cells was investigated. Overall, these results suggest that γ-tocotrienol is a novel blocker of the STAT3 activation pathway, with a potential role in future therapies for HCC and other cancers.
Tocotrienols have potent antifibrogenic effects in human intestinal fibroblasts. Luna, L, et.al (2011). Inflamm Bowel Dis.	The effects of tocotrienols on human intestinal fibroblast (HIF) proliferation, apoptosis, autophagy, and synthesis of ECM (extracellular matrix) were investigated. Tocotrienols exert multiple effects on HIF, reducing cell proliferation, enhancing programmed cell death through apoptosis and autophagy, and decreasing ECM production. Considering their in vitro antifibrogenic properties, tocotrienols could be useful to treat or prevent bowel fibrosis in CD patients.
Tocotrienol-Rich Fraction Prevents Cell Cycle Arrest and Elongates Telomere Length in Senescent Human Diploid Fibroblasts Makpol, S., et.al (2011). J Biomed Biotechnol.	This study determined the molecular mechanisms of tocotrienol-rich fraction (TRF) in preventing cellular senescence of human diploid fibroblasts (HDFs). Primary culture of HDFs at various passages were incubated with 0.5 mg/mL TRF for 24 h. Telomere shortening with decreased telomerase activity was observed in senescent HDFs while the levels of damaged DNA and number of cells in G₀/G₁ phase were increased and S phase cells were decreased. Incubation with TRF reversed the morphology of senescent HDFs to resemble that of young cells with decreased activity of SA-β-gal, damaged DNA, and cells in G₀/G₁ phase while cells in the S phase were increased. Elongated telomere length and restoration of telomerase activity were observed in TRF-treated senescent HDFs. These findings confirmed the ability of tocotrienol-rich fraction in preventing HDFs cellular ageing by restoring telomere length and telomerase activity, reducing damaged DNA, and reversing cell cycle arrest associated with senescence.
Mechanisms Mediating the Synergistic Anticancer Effects of Combined γ-Tocotrienol and Celecoxib Treatment Shirode, A. B., et.al (2011). J Bioanal Biomed.	To characterize the intracellular signaling mechanisms mediating the synergistic anticancer effects of combined γ-tocotrienol and celecoxib treatment in neoplastic +SA mouse mammary epithelial cells in vitro. The synergistic anticancer effects of combined low dose γ-tocotrienol and celecoxib treatment in +SA mammary tumor cells are mediated by COX-2-dependent mechanisms associated with a suppression in PGE₂ levels, as well as, COX-2-independent mechanisms associated with a reduction in ErbB2-4 receptor levels, activation, and subsequent reduction in downstream Akt and NFκB mitogenic signaling.
δ-tocotrienol protects mouse and human hematopoietic progenitors from γ-irradiation through extracellular signal-regulated kinase/mammalian target of rapamycin signaling Li, X. H., et.al (2010). Haematologica.	Natural δ-tocotrienol has been demonstrated to significantly enhanced survival in total body lethally irradiated mice. The data indicate that δ-tocotrienol protects mouse bone marrow and human CD34⁺ cells from radiation-induced damage through extracellular signal-related kinase activation-associated mammalian target of rapamycin survival pathways.
Tocotrienol-Rich Fraction Prevents Cell Cycle Arrest and Elongates Telomere Length in Senescent Human Diploid Fibroblasts Makpol, S., et.al (2011). J Biomed Biotechnol.	This study determined the molecular mechanisms of tocotrienol-rich fraction (TRF) in preventing cellular senescence of human diploid fibroblasts (HDFs). Primary culture of HDFs at various passages were incubated with 0.5 mg/mL TRF for 24 h. Telomere shortening with decreased telomerase activity was observed in senescent HDFs while the levels of damaged DNA and number of cells in G₀/G₁ phase were increased and S phase cells were decreased. Incubation with TRF reversed the morphology of senescent HDFs to resemble that of young cells with decreased activity of SA-β-gal, damaged DNA, and cells in G₀/G₁ phase while cells in the S phase were increased. Elongated telomere length and restoration of telomerase activity were observed in TRF-treated senescent HDFs. These findings confirmed the ability of tocotrienol-rich fraction in preventing HDFs cellular ageing by restoring telomere length and telomerase activity, reducing damaged DNA, and reversing cell cycle arrest associated with senescence.
Mechanisms Mediating the Synergistic Anticancer Effects of Combined γ-Tocotrienol and Celecoxib Treatment Shirode & Sylvester (2011). J Bioanal Biomed.	To characterize the intracellular signaling mechanisms mediating the synergistic anticancer effects of combined γ-tocotrienol and celecoxib treatment in neoplastic +SA mouse mammary epithelial cells in vitro. The synergistic anticancer effects of combined low dose γ-tocotrienol and celecoxib treatment in +SA mammary tumor cells are mediated by COX-2-dependent mechanisms associated with a suppression in PGE₂ levels, as well as, COX-2-independent mechanisms associated with a reduction in ErbB2-4 receptor levels, activation, and subsequent reduction in downstream Akt and NFκB mitogenic signaling.
δ-tocotrienol protects mouse and human hematopoietic progenitors from γ-irradiation through extracellular signal-regulated kinase/mammalian target of rapamycin signaling Li, X.H., et.al (2010). Haematologica.	In this study, natural δ-tocotrienol has been demonstrated significantly enhanced survival in total body lethally irradiated mice. The effects and mechanisms of δ-tocotrienol on hematopoietic progenitor cell survival after γ-irradiation in both in vivo and in vitro experiments. Our data indicate that δ-tocotrienol protects mouse bone marrow and human CD34⁺ cells from radiation-induced damage through extracellular signal-related kinase activation-associated mammalian target of rapamycin survival pathways.
Delta-gamma-tocotrienol-mediated cell cycle arrest and apoptosis in human melanoma cells. Fernandes, N.V., et.al (2010). Anticancer Res.	The rate-limiting enzyme of the mevalonate pathway, 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase, provides essential intermediates for the prenylation or dolichylation of growth-related proteins. d-δ-Tocotrienol, a post-transcriptional downregulator of HMG CoA reductase, suppresses the proliferation of murine B16 melanoma cells. Dietary d-δ- tocotrienol suppresses the growth of implanted B16 melanomas without toxicity to host mice. d-δ-Tocotrienol induced dose-dependent suppression of cell proliferation with 50% inhibitory concentrations (IC50) of 37.5±1.4 (A2058) and 22.3±1.8 (A375) μmol/l, respectively (data are reported as mean±standard deviation). d-δ-Tocotrienol-mediated cell cycle arrest at the G1 phase was accompanied by reduced expression of cyclin-dependent kinase 4. Concomitantly, d- δ-tocotrienol induced caspase-3 activation and apoptosis. The impact of d-δ-tocotrienol on A2058 cell proliferation was potentiated by lovastatin (IC50=3.1±0.5 μmol/l), a competitive inhibitor of HMG CoA reductase. Conclusion: d-δ-Tocotrienol may have potential application in melanoma chemoprevention and/or therapy.
γ-Tocotrienol but Not γ-Tocopherol Blocks STAT3 Cell Signaling Pathway through Induction of Protein-tyrosine Phosphatase SHP-1 and Sensitizes Tumor Cells to Chemotherapeutic Agents Kannappan, R., et.al (2010). J Biol Chem.	In this study, γ-T3 modulate the STAT3 cell signaling pathway, closely linked to inflammation and tumorigenesis is investigated. We found that γ-T3 but not γ-tocopherol, the most common saturated form of vitamin E, inhibited constitutive activation of STAT3 in a dose- and time-dependent manner, and this inhibition was not cell type-specific. γ-T3 also inhibited STAT3 DNA binding. This correlated with inhibition of Src kinase and JAK1 and JAK2 kinases. Pervanadate reversed the γ-T3-induced down-regulation of STAT3 activation, suggesting the involvement of a protein-tyrosine phosphatase. When examined further, it is found that γ-T3 induced the expression of the tyrosine phosphatase SHP-1, and gene silencing of the SHP-1 by small interfering RNA abolished the ability of γ-T3 to inhibit STAT3 activation, suggesting a vital role for SHP-1 in the action of γ-T3. Also γ-T3 down-modulated activation of STAT3 and induced SHP-1 in vivo. Eventually, γ-T3 down-regulated the expression of STAT3-regulated antiapoptotic (Bcl-2, Bcl-xL, and Mcl-1), proliferative (cyclin D1), and angiogenic (VEGF) gene products; and this correlated with suppression of proliferation, the accumulation of cells in sub-G(1) phase of the cell cycle, and induction of apoptosis. This vitamin also sensitized the tumor cells to the apoptotic effects of thalidomide and bortezomib. Overall, the results suggest that γ-T3 is a novel blocker of STAT3 activation pathway both in vitro and in vivo and thus may have potential in prevention and treatment of cancers.
The tocotrienol-rich fraction from rice bran enhances cisplatin-induced cytotoxicity in human mesothelioma H28 cells. Nakashima, K, et.al (2010). Phytother Res.	Resistance to chemotherapy (chemoresistance) is a serious problem in malignant mesothelioma, a highly aggressive neoplasm. Gamma-tocotrienol (gamma-T3) can sensitize various cancerous cells to chemotherapeutic agents by inhibiting pathways that lead to treatment resistance. In this study, the modulating effect of tocotrienol-rich fraction (TRF) from rice bran, which is abundant in gamma-T3, on chemoresistance in human MM H28 cells was investigated. TRF treatment caused a marked reduction in the viability of H28 cells in a dose-dependent manner, while cisplatin treatment had no effect on the cells, indicating that H28 cells are resistant to cisplatin. A significant increase in cytotoxicity was observed in H28 cells treated with TRF, and this effect was enhanced by the combination treatment with cisplatin. The cytotoxic effect was closely related to the inhibition of phosphatidylinositol 3-kinase (PI3K)-AKT signaling. Inactivation of Akt signaling by TRF or the combination with cisplatin mitigated cisplatin-induced activation of Akt, resulting in reducing the chemoresistance H28 cells to cisplatin. Reduced cell viability and attenuated chemoresistance of the H28 cells against cisplatin were also observed following the use of a PI3K inhibitor, LY294002. These results suggest that the combination therapy of cisplatin with TRF is a plausible strategy for achieving tolerance for the chemotherapeutic agent in MM therapy.
Effect of Delta-Tocotrienol on Melanin Content and Enzymes for Melanin Synthesis in Mouse Melanoma Cells Michihara, A., et.al (2010). Biol. Pharm. Bull.	The dose-dependent effect of d-tocotrienol long term (48, 72 h) on the melanin content of cells treated with d-tocotrienol, and whether cells treated with d-tocotrienol for long a time show cytotoxicity was investigated. It is also examined whether other enzymes responsible for melanin biosynthesis, tyrosinaserelated protein-1 (TRP-1) and -2 (TRP-2), are involved in the decrease in melanin levels. Protein levels in cells treated with 25 or 50 mM d-tocotrienol for 48 h or 72 h were similar to those in control cells. Melanin content decreased by 44 (25 mM d-tocotrienol) to 50% (50 mM) at 48 h, and by 14 to 21% at 72 h, compared to control levels. Tyrosinase activity, amounts of tyrosinase and TRP-1 decreased dependent on dose : by 50 (25 mM d-tocotrienol) to 75% (50 mM), 20 to 45% and 42 to 82% at 48 h, and by 25 to 50%, 75 to 80% and 78 to 77% at 72 h, respectively. Although the amount of TRP-2 increased by 20% on treatment with 25 mM d-tocotrienol for 48 h, it decreased by 52% on treatment with 50 mM d-tocotrienol for 48 h. The amount of TRP-2 dose-dependently decreased by 55% and 75% on 72 h by treatment with 25 and 50 mM d-tocotrienol, respectively. From these findings, d-tocotrienol at up to 50 mM dose-dependently caused a reduction in melanin content by the decrease of TRP-1 and TRP-2 as well as tyrosinase, and no cytotoxicity.
Gamma-tocotrienol promotes TRAIL-induced apoptosis through reactive oxygen species/extracellular signal-regulated kinase/ p53-mediated upregulation of death receptors. Kannappan, R., et.al (2010). Mol Cancer Ther.	The ability of tocotrienol (T3), an unsaturated vitamin E present in palm oil, rice bran, barley, oats, and wheat germ, to sensitize tumor cells to TRAIL was investigated. The results show that gamma-T3 sensitizes tumor cells to TRAIL by upregulating DRs through the ROS/ERK/p53 pathway and by downregulating cell survival proteins.
Gamma-tocotrienol modulates the paracrine secretion of VEGF induced by cobalt (II) chloride via ERK signaling pathway in gastric adenocarcinoma SGC-7901 cell line. Bi, S., et.al (2010). Toxicology.	In this study, the effect of gamma-tocotrienol, a natural product commonly found in palm oil and rice bran, on the accumulation of HIF-1alpha protein and the paracrine secretion of VEGF in human gastric adenocarcinoma SGC-7901 cell line induced by cobalt(II) chloride (as a hypoxia mimic) was investigated. These results showed that cobalt (II) chloride induced the high expression of VEGF in SGC-7901 cells at dose of 150 micromol/L for 24h. Both basal level and cobalt(II) chloride-induced HIF-1alpha protein accumulation and VEGF paracrine secretion were inhibited in SGC-7901 cells treated with gamma-tocotrienol at 60 micromol/L treatment for 24 h. U0126, a MEK1/2 inhibitor, decreased the expression of HIF-1alpha protein and the paracrine secretion of VEGF under normoxic and hypoxic conditions. In this study, gamma-tocotrienol also significantly inhibited the hypoxia-stimulated expression of phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK1/2). The mechanism seems to involve in inhibiting hypoxia-mediated activation of p-ERK1/2, it leads to a marked decrease in hypoxia-induced HIF-1alpha protein accumulation and VEGF secretion. These data suggest that HIF-1alpha/VEGF could be a promising target for gamma-tocotrienol in an effective method of chemoprevention and chemotherapy in human gastric cancer.
Synergistic anticancer effects of combined γ-tocotrienol and celecoxib treatment are associated with suppression in Akt and NFκB signaling Shirode, A.B., et.al (2010). Biomed Pharmacother.	The selective cyclooxygenase (COX)-2 inhibitor, celecoxib, and the vitamin E isoform, γ-tocotrienol, both display potent anticancer activity. However, high dose clinical use of selective COX-2 inhibitors has been limited by gastrointestinal and cardiovascular toxicity, whereas limited absorption and transport of γ-tocotrienol by the body has made it difficult to obtain and sustain therapeutic levels in the blood and target tissues. Studies were conducted to characterize the synergistic anticancer antiproliferative effects of combined low dose celecoxib and γ-tocotrienol treatment on mammary tumor cells in culture. The highly malignant mouse +SA mammary epithelial cells were maintained in culture on serum-free defined control or treatment media. Treatment effects on COX-1, COX-2, Akt, NFκB and prostaglandin E₂ (PGE₂) synthesis was assessed following a 3- or 4-day culture period. Treatment with 3–4 μM γ-tocotrienol or 7.5–10 μM celecoxib alone significantly inhibited +SA cell growth in a dose-responsive manner. However, combined treatment with subeffective doses of γ-tocotrienol (0.25 μM) and celecoxib (2.5 μM) resulted in a synergistic antiproliferative effect, as determined by isobologram analysis, and this growth inhibitor effect was associated with a reduction in PGE₂ synthesis, and decrease in COX-2, phospho-Akt (active), and phospho-NFκB (active) levels. These results demonstrate that the synergistic anticancer effects of combined celecoxib and γ-tocotrienol therapy are mediated by COX-2 dependent and independent mechanisms. These findings also suggest that combination therapy with these agents may provide enhanced therapeutic response in breast cancer patients, while avoiding the toxicity associated with high-dose COX-2 inhibitor monotherapy.
Preparation, characterization and anticancer effects of simvastatin-tocotrienol lipid nanoparticles. Shirode, A.H., et.al (2010). Int J Pharm.	The objective of the present work was to prepare and characterize lipid nanoparticles that combined simvastatin and tocotrienol rich fraction (TRF) as potential anticancer therapy. The entrapment of simvastatin in the oily nanocompartments, which were formed by TRF inclusion into the solid matrix of the nanoparticles, was verified by its high entrapment efficiency and the absence of endothermic or crystalline peaks when blends were analyzed by DSC and PXRD, respectively. The release of simvastatin from the nanoparticles in sink conditions was characterized by an initial burst release of approximately 20% in 10h followed by a plateau. No significant change in particle size (approximately 100 nm) was observed after storage for six months. The anticancer activity of the nanoparticles was verified in vitro by observing their antiproliferative effects on malignant +SA mammary epithelial cells. The IC(50) of the reference alpha-tocopherol nanoparticles was 17.7 microM whereas the IC(50) of the simvastatin/TRF nanoparticles was 0.52 microM, which confirmed the potency of the combined treatment and its potential in cancer therapy.
Inhibitory effects of gamma-tocotrienol on invasion and metastasis of human gastric adenocarcinoma SGC-7901 cells. Liu, H.K., et.al (2010). J Nutr Biochem.	In this study, the invasion and metastatic capacities of gastric adenocarcinoma SGC-7901 cells and the correlation with antimetastasis mechanisms induced by gamma-tocotrienol were explored. The results showed the inhibitory effects of gamma-tocotrienol at doses of 15, 30, 45 and 60 mumol/L for 48 h on cell migration and cell matrigel invasion; activities of matrix metalloproteinase (MMPs) increased in SGC-7901 cells when compared to the control group (P<.05 or P<.01). An increasing trend in the chemotactic responses to fibronectin (FN) in SGC-7901 cells was found in the gamma-tocotrienol treatments. SGC-7901 cell attachment decreased in the gamma-tocotrienol-treated groups in comparison with the control group (P<.01). The mRNA expressions of MMP-2 and MMP-9 showed that gamma-tocotrienol significantly reduced the matrigel invasion capability through down-regulation of the mRNA expressions of MMP-2 and MMP-9 (P<.01), and up-regulation of tissue inhibitor of metalloproteinase-1 (TIMP-1) and TIMP-2 in SGC-7901 cells by treatment with gamma-tocotrienol for 48 h (P<.05). gamma-Tocotrienol also significantly increased the mRNA expression of nm23-H1 in SGC-7901 cells (P<.01). These findings suggest a potential mechanism of gamma-tocotrienol-mediated antitumor metastasis activity and indicate the role of vitamin E as potential chemopreventative agents against gastric cancer.
Tocotrienol inhibited growth and induced apoptosis in human HeLa cells through the cell cycle signaling Wu & Ng (2010). Integr Cancer Ther.	Tocotrienols of palm oil have been shown to possess potent neuroprotective, antioxidative, anticancer, and cholesterol-lowering activities. In this study, the authors examined the antiproliferative effects of alpha-, gamma- and delta-tocotrienols (alphaT3, gammaT3, and deltaT3), and alpha-tocopherol (alphaT) in human cervical carcinoma (HeLa) cells. Their mechanism(s) of action on cell cycle signaling pathway were also investigated.
Synergistic Actions of Atorvastatin with γ-Tocotrienol and Celecoxib against Human Colon Cancer HT29 and HCT116 Cells Yang, Z.H., et.al (2010). Int J Cancer.	The synergistic actions of atorvastatin (ATST) with γ-tocotrienol (γ-TT) and celecoxib (CXIB) were studied in human colon cancer cell lines HT29 and HCT116. The synergistic inhibition of cell growth by ATST and γ-TT was demonstrated by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay and isobologram analysis. δ-TT exhibited a similar inhibitory action when combined with ATST. Mevalonate and geranylgeranyl pyrophosphate eliminated most of the growth inhibitory effect of ATST, but only marginally decreased that of γ-TT; whereas farnesyl pyrophosphate and squalene exhibited little effect on the inhibitory action of ATST and γ-TT, indicating protein geranylgeranylation, but not farnesylation are involved in the inhibition of colon cancer cell growth. Both mevalonate and squalene restored the cellular cholesterol level that was reduced by ATST treatment, but only mevalonate eliminated the cell growth inhibitory effect, suggesting that the cholesterol level in cells does not play an essential role in inhibiting cancer cell growth. Protein level of HMG-CoA reductase increased after ATST treatment, and the presence of γ-TT attenuated the elevated level of HMG-CoA reductase. ATST also decreased membrane-bound RhoA, possibly due to a reduced level of protein geranylgeranylation; addition of γ-TT enhanced this effect. The mediation of HMG-CoA reductase and RhoA provides a possible mechanism for the synergistic action of ATST and γ-TT. The triple combination of ATST, γ-TT, and CXIB showed a synergistic inhibition of cancer cell growth in MTT assays. The synergistic action of these three compounds was also illustrated by their induction of G₀/G₁ phase cell cycle arrest and apoptosis.
Tocotrienols are good adjuvants for developing cancer vaccines Abdul Hafid, S.R., et.al (2010). BMC Cancer.	Tocotrienol-rich fraction (TRF), a non-toxic natural compound, as an adjuvant to enhance the effectiveness of dendritic cells (DC) vaccines in treating mouse mammary cancers. In the mouse model, six-week-old female BALB/c mice were injected subcutaneously with DC and supplemented with oral TRF daily (DC+TRF) and DC pulsed with tumour lysate from 4T1 cells (DC+TL). Experimental mice were also injected with DC pulsed with tumour lysate and supplemented daily with oral TRF (DC+TL+TRF) while two groups of animal which were supplemented daily with carrier oil (control) and with TRF (TRF). After three times vaccination, mice were inoculated with 4T1 cells in the mammary breast pad to induce tumour. The study shows that TRF has the potential to be an adjuvant to augment DC based immunotherapy.
Tumor regression after systemic administration of tocotrienol entrapped in tumor-targeted vesicles. Fu, J.Y., et.al (2009). J Control Release.	The therapeutic potential of tocotrienol, an extract of vitamin E with anti-cancer properties, is hampered by its failure to specifically reach tumors after intravenous administration, without secondary effects on normal tissues. We hypothesize that the encapsulation of tocotrienol-rich fraction (TRF) within vesicles bearing transferrin, whose receptors are overexpressed on many cancer cells, could result in a selective delivery to tumors after intravenous administration. The objectives of this study are therefore to prepare and characterize transferrin-targeted vesicles encapsulating TRF, and to evaluate their therapeutic efficacy in vitro and in vivo. The entrapment of TRF in transferrin-bearing vesicles led to a 3-fold higher TRF uptake and more than 100-fold improved cytotoxicity in A431 (epidermoid carcinoma), T98G (glioblastoma) and A2780 (ovarian carcinoma) cell lines compared to TRF solution. The intravenous administration of TRF encapsulated in transferrin-bearing vesicles led to tumor regression and improvement of animal survival in a murine xenograft model, contrary to that observed with controls. The treatment was well tolerated by the animals. This work corresponds to the first preparation of a tumor-targeted delivery system able to encapsulate tocotrienol. Our findings show that TRF encapsulated in transferrin-bearing vesicles is a highly promising therapeutic system, leading to tumor regression after intravenous administration without visible toxicity.
Evidence of gamma-tocotrienol as an apoptosis-inducing, invasion-suppresing, and chemotherapy drug-sensitizing agent in human melanoma cells. Chang, P.N., et.al (2009). Nutr Cancer.	To date, the most effective cure for metastatic melanoma remains the surgical resection of the primary tumor. Recently, tocotrienol-rich-fraction has shown antiproliferative effect on cancer cells. To elucidate this anticancer property in malignant melanoma, this study aimed, first, to identify the most potent isomer for eliminating melanoma cells and second to decipher the molecular pathway responsible for its activity. Results showed that the inhibitory effect of gamma-tocotrienol was most potent, which resulted in induction of apoptosis as evidenced by activation of procaspases and the accumulation of sub-G1 cell population. Examination of the prosurvival genes revealed that the gamma-tocotrienol-induced cell death was associated with suppression of NF-kappaB, EGF-R, and Id family proteins. Meanwhile, gamma-tocotrienol treatment also resulted in induction of JNK signaling pathway, and inhibition of JNK activity by selective inhibitor was able to partially block the effect of gamma-tocotrienol. Interestingly, gamma-tocotrienol treatment led to suppression of mesenchymal markers and the restoration of E-cadherin and gamma-catenin expression, which was associated with suppression of cell invasion capability. Furthermore, synergistic effect was observed when cells were cotreated with gamma-tocotrienol and chemotherapy drugs. Together, our results demonstrated for the first time the anti-invasion and chemonsensitization effect of gamma-tocotrienol against human malignant melanoma cells.
Suppression of tumor growth by palm tocotrienols via the attenuation of angiogenesis. Wong, W-Y., et.al (2009). Nutr Cancer.	In this study, the antiangiogenic effects of TRF (tocotrienol-rich fraction) was examined. In vitro investigations of the antiangiogenic activities of TRF, delta-tocotrienol (deltaT3), and alpha-tocopherol (alphaToc) were carried out in human umbilical vein endothelial cells (HUVEC). TRF and deltaT3 significantly inhibited cell proliferation from 4 microg/ml onward (P < 0.05). Cell migration was inhibited the most by deltaT3 at 12 microg/ml. Anti-angiogenic properties of TRF were carried out further in vivo using the chick embryo chorioallantoic membrane (CAM) assay and BALB/c mice model. TRF at 200 microg/ml reduced the vascular network on CAM. TRF treatment of 1 mg/mouse significantly reduced 4T1 tumor volume in BALB/c mice. TRF significantly reduced serum vascular endothelial growth factor (VEGF) level in BALB/c mice. In conclusion, this study showed that palm tocotrienols exhibit anti-angiogenic properties that may assist in tumor regression.
Suppression in mevalonate synthesis mediates antitumor effects of combined statin and gamma-tocotrienol treatment. Wali, V.B., et.al (2009). Lipids.	Statins directly inhibit 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR) activity, while gamma-tocotrienol, an isoform of vitamin E, enhances the degradation and reduces cellular levels of HMGR in various tumor cell lines. Since treatment with statins or gamma-tocotrienol alone induced a dose-responsive inhibition, whereas combined treatment with subeffective doses of these agents resulted in a synergistic inhibition in +SA mammary tumor cell growth, studies were conducted to investigate the role of the HMGR pathway in mediating the antiproliferative effects of combined low dose statin and gamma-tocotrienol. Treatment with 8 microM simvastatin inhibited cell growth and isoprenylation of Rap1A and Rab6, and supplementation with 2 microM mevalonate reversed these effects. However, the growth inhibitory effects of 4 microM gamma-tocotrienol were not dependent upon suppression in mevalonate synthesis. Treatment with subeffective doses of simvastatin (0.25 microM), lovastatin (0.25 microM), mevastatin (0.25 microM), pravastatin (10 microM), or gamma-tocotrienol (2 muM) alone had no effect on protein prenylation or mitogenic signaling, whereas combined treatment with these agents resulted in a significant inhibition in +SA cell growth, and a corresponding decrease in total HMGR, Rap1A and Rab6 prenylation, and MAPK signaling, and mevalonate supplementation reversed these effects. These findings demonstrate that the synergistic antiproliferative effects of combined low dose statin and gamma-tocotrienol treatment are directly related to an inhibition in HMGR activity and subsequent suppression in mevalonate synthesis.
A redox-silent analogue of tocotrienol acts as potential cytotoxic agent against human mesothelioma cells. Kashiwagi, K, et.al (2009). Life Sci.	The effect of tocotrienol (T3) on killing of chemoresistant mesothelioma cell (H28) was investigated. Tocotrienol can be a new effective therapeutic agent against chemoresistant mesothelioma cells.
Gamma-tocotrienol suppresses prostate cancer cell proliferation and invasion through multiple-signaling pathways. Yap, W.N., et.al (2008). Br J Cancer.	Tocotrienol-rich fraction (TRF) has demonstrated antiproliferative effect on prostate cancer (PCa) cells. To elucidate this anticancer property in PCa cells, this study aimed, first, to identify the most potent isomer for eliminating PCa cells; and second, to decipher the molecular pathway responsible for its activity. Results showed that the inhibitory effect of γ-tocotrienol was most potent, which resulted in induction of apoptosis as evidenced by activation of pro-caspases and the presence of sub-G₁ cell population. Examination of the pro-survival genes revealed that the γ-tocotrienol-induced cell death was associated with suppression of NF-κB, EGF-R and Id family proteins (Id1 and Id3). Meanwhile, γ-tocotrienol treatment also resulted in the induction of JNK-signalling pathway and inhibition of JNK activity by a specific inhibitor (SP600125) was able to partially block the effect of γ-tocotrienol. Interestingly, γ-tocotrienol treatment led to suppression of mesenchymal markers and the restoration of E-cadherin and γ-catenin expression, which was associated with suppression of cell invasion capability. Furthermore, a synergistic effect was observed when cells were co-treated with γ-tocotrienol and Docetaxel. Our results suggested that the antiproliferative effect of γ-tocotrienol act through multiple-signalling pathways, and demonstrated for the first time the anti-invasion and chemosensitisation effect of γ-tocotrienol against PCa cells.
Tocotrienol Inhibits Secretion of Angiogenic Factors from Human Colorectal Adenocarcinoma Cells by Suppressing Hypoxia-Inducible Factor-1α Shibata, A., et.al (2008). American Society for Nutrition.	Tocotrienol (T3), unsaturated vitamin E, has recently gained considerable attention as a potent antiangiogenic agent minimizing tumor growth, the exact intracellular mechanisms of which remain poorly understood. Because hypoxia-inducible factor-1α (HIF-1α), its downstream target vascular endothelial growth factor (VEGF), and other angiogenic factors such as interleukin-8 (IL-8) and cyclooxygenase 2 (COX-2) play critical roles in neovascularization, we tested the hypothesis that the inhibitory effect of T3 on tumor angiogenesis is via regulation of these angiogenic factors. 2 cancer cell lines, human colorectal adenocarcinoma cells (DLD-1) and human hepatoma cells (HepG2). T3 isomers (2 μmol/L) inhibited hypoxia-induced VEGF secretion from DLD-1, with δ-T3 showing potent inhibition. δ-T3 suppressed hypoxia-induced VEGF and IL-8 expression in DLD-1 at both mRNA and protein levels, and it was found the inhibitory mechanism of δ-T3 by reducing HIF-1α protein expression or increasing HIF-1αdegradation. Also, δ-T3 (2 μmol/L) did not affect hypoxia-induced COX-2 mRNA expression; however, δ-T3 tended to suppress (P = 0.044) hypoxia-induced COX-2 protein expression, implying a possible post-transcriptional mechanism by δ-T3. Overall, the results confirmed that T3 has an inhibitory effect on angiogenic factor secretion from cancer cells and revealed the possible mechanisms, providing new information about the antiangiogenic effects of T3.
Gamma-tocotrienol-induced apoptosis in human gastric cancer SGC-7901 cells is associated with a suppression in mitogen-activated protein kinase signaling. Wang, S.W., et.al (2008). Br J Nutr.	Tocotrienols have been shown to inhibit proliferation and induce apoptosis in cancer cells. However, the molecular mechanisms involved in tocotrienol-induced apoptosis are still unclear. In the present study, gamma-tocotrienol induced apoptosis in human gastric adenocarcinoma SGC-7901 cell line through down regulation of the extracellular signal-regulated kinase (ERK) signalling pathway. Furthermore, gamma-tocotrienol-induced apoptosis was accompanied by down regulation of Bcl-2, up regulation of Bax, activation of caspase-3, and subsequent poly (ADP-ribose) polymerase cleavage. These results indicated that up or down regulation of Bcl-2 family proteins play a major role in the initiation of gamma-tocotrienol-induced apoptosis as an activator of caspase-3. Gamma-tocotrienol also down regulated the activation of the Raf-ERK signalling pathway, and down regulated c-Myc by decreasing the expressions of Raf-1 and p-ERK1/2 proteins. The results suggest that key regulators in tocotrienol-induced apoptosis may be Bcl-2 families and caspase-3 in SGC-7901 cells through down regulation of the Raf-ERK signalling pathway.
γ-Tocotrienol Inhibits Nuclear Factor-κB Signaling Pathway through Inhibition of Receptor-interacting Protein and TAK1 Leading to Suppression of Antiapoptotic Gene Products and Potentiation of Apoptosis Kwang S.A., et.al (2007). J Bio Chem.	The effect of γ-tocotrienol on the NF-κB pathway in tumorigenesis was investigated. Overall, the results demonstrate that γ-tocotrienol inhibited the NF-κB activation pathway, leading to down-regulation of various gene products and potentiation of apoptosis.
Apoptosis induction by gamma-tocotrienol in human hepatoma Hep3B cells. Sakai, M., et.al (2006) J Nutr Biochem..	The antitumor activity of tocotrienol (T3) on human hepatoma Hep3B cells was evaluated. At first, the effect of T3 on the proliferation of human hepatoma Hep3B cells and found that gamma-T3 inhibited cell proliferation at lower concentrations and shorter treatment times than alpha-T3 was examined. Then, the effect of gamma-T3 apoptosis induction and found that gamma-T3 induced poly (ADP-ribose) polymerase (PARP) cleavage and stimulated a rise in caspase-3 activity was examined. In addition, gamma-T3 stimulated a rise in caspase-8 and caspase-9 activities. It was found that gamma-T3-induced apoptotic cell death was accompanied by up-regulation of Bax and a rise in the fragments of Bid and caspase-8. These data indicate that gamma-T3 induced apoptosis in Hep3B cells and that caspase-8 and caspase-9 were involved in apoptosis induction. Moreover, these results suggest that Bax and Bid regulated apoptosis induction by gamma-T3.
Down-regulation of telomerase activity in DLD-1 human colorectal adenocarcinoma cells by tocotrienol. Eitsuka, T., et.al (2006). Biochem Biophys Res Commun.	As high telomerase activity is detected in most cancer cells, inhibition of telomerase by drug or dietary food components is a new strategy for cancer prevention. Here, we investigated the inhibitory effect of vitamin E, with particular emphasis on tocotrienol (unsaturated vitamin E), on human telomerase in cell-culture study. As results, tocotrienol inhibited telomerase activity of DLD-1 human colorectal adenocarcinoma cells in time- and dose-dependent manner, interestingly, with delta-tocotrienol exhibiting the highest inhibitory activity. Tocotrienol inhibited protein kinase C activity, resulting in down-regulation of c-myc and human telomerase reverse transcriptase (hTERT) expression, thereby reducing telomerase activity. In contrast to tocotrienol, tocopherol showed very weak telomerase inhibition. These results provide novel evidence for the first time indicating that tocotrienol acts as a potent candidate regulator of telomerase and supporting the anti-proliferative function of tocotrienol.
Tocotrienol inhibits proliferation of human Tenon’s fibroblasts in vitro: A comparative study with vitamin E forms and mitomycin C. Meyenberg, A., et.al (2005). Graefes Arch Clin Exp Ophthalmol.	The potential of the vitamin E compound alpha-tocotrienol as antifibrotic agent in vitro was evaluated. Of the vitamin E forms tested, only alpha-tocotrienol significantly inhibited growth at non-toxic concentrations. In this in vitro study, antiproliferative effects of mitomycin C were stronger than those of alpha-tocotrienol.
Effects of tocotrienols on cell viability and apoptosis in normal murine liver cells ( BNL CL2) and liver cancer cells (BNL1ME A.7R.1) in vitro. Har C.H., et.al (2005). Asia Pac J Clin Nutr.	The effects of tocotrienols on murine liver cell viability and their apoptotic events were studied over a dose range of 0-32 microg mL(-1). Normal murine liver cells (BNL CL.2) and murine liver cancer cells (BNL 1ME A.7R.1) were treated with tocotrienols (T(3)), alpha tocopherol (alpha-T) and the chemo drug, Doxorubicin (Doxo, as a positive control). The results suggest that tocotrienols were able to reduce the cell viability in the murine liver cancer cells at a dose of 8-32 microg mL(-1) and that this decrease in percentage cell viability may be due to apoptosis.
Induction of cytotoxicity in human lung adenocarcinoma cells by 6-O-carboxypropyl-alpha-tocotrienol, a redox silent derivative of alpha-tocotrienol Yano, Y., et.al (2005). Int J Cancer.	Tocotrienols are one of the most potent anticancer agents of all natural compounds and the anticancer property may be related to the inactivation of Ras family molecules. The anticancer potential of tocotrienols, however, is weakened due to its short elimination half life in vivo. To overcome the disadvantage and reinforce the anticancer activity in tocotrienols, a redox-silent analogue of alpha-tocotrienol (T3), 6-O-carboxypropyl-alpha-tocotrienol (T3E) is synthesized. IT is estimated the possibility of T3E as a new anticancer agent against lung adenocarcinoma showing poor prognosis based on the mutation of ras gene. T3E showed cytotoxicity against A549 cells, a human lung adenocarcinoma cell line with a ras gene mutation, in a dose-dependent manner (0-40 microM), whereas T3 and a redox-silent analogue of alpha-tocopherol (T), 6-O-carboxypropyl-alpha-tocopherol (TE), showed much less cytotoxicity in cells within 40 microM. T3E cytotoxicity was based on the accumulation of cells in the G1-phase of the cell-cycle and the subsequent induction of apoptosis. Similar to this event, 24-hr treatment of A549 cells with 40 microM T3E caused the inhibition of Ras farnesylation, and a marked decrease in the levels of cyclin D required for G1/S progression in the cell-cycle and Bcl-xL, a key anti-apoptotic molecule. Moreover, the T3E-dependent inhibition of RhoA geranyl-geranylation is an inducing factor for the occurrence of apoptosis in A549 cells. The results suggest that T3E suppresses Ras and RhoA prenylation, leading to negative growth control against A549 cells. In conclusion, a redox-silent analogue of T3, T3E may be a new candidate as an anticancer agent against lung adenocarcinoma showing poor prognosis based on the mutation of ras genes.
Induction of Apoptosis by Tocotrienol in Rat Hepatoma dRLh-84 Cells Sakai, M., et.al (2004). Anticancer Res.	To evaluate the antitumor activities of tocopherol (Toc) and tocotrienol (T3) derivatives. At first, the effect of these vitamin E homologues on the proliferation of rat normal hepatocyte RLN-10 and hepatoma dRLh-84 cells and found that especially T3 inhibited cell proliferation in dRLh-84 cells was evaluated. Then, the effect of vitamin E homologues on apoptosis induction and found that T3 induced DNA fragmentation and stimulated a rise of caspase-3 activity was examined. In addition, T3 stimulated a rise in caspase- 8 activity, while a caspase-8 inhibitor suppressed apoptosis induction by T3. The incorporation of vitamin E homologues into dRLh-84 cells. T3 was incorporated more quickly, compared to Toc. These results indicated that T3 induces apoptosis in dRLh-84 cells and that caspase-8 is involved in this apoptosis induction. The difference in terms of apoptosis induction by vitamin E homologues seems to be related to their different rates of cellular incorporation.
Tocotrienol-rich fraction of palm oil activates p53, modulates Bax/Bcl2 ratio and induces apoptosis independent of cell cycle association. Agarwal, M.K., et.al (2004). Cell Cycle.	Several chemopreventive agents have been shown to utilize p53 pathway in negative regulation of cell growth, using human colon carcinoma RKO cells which express wild type p53, the effect of TRF on components of p53 signaling network was investigated. Data suggest that TRF-induced apoptosis in colon carcinoma cells is mediated by p53 signaling network which appears to be independent of cell cycle association.
Anti-angiogenic Potential of Tocotrienol in vitro Miyazawa, T., et.al (2003). Biochemistry.	The physiological activity of vitamin E has been recognized to be associated with its well-defined antioxidant property, especially against lipid peroxidation in biological membranes. Among the vitamin E group, alpha-tocopherol is considered to be the most active form in vivo. However, a recent study suggested that tocotrienol may be a better antioxidant than tocopherol. Moreover, tocotrienol has been reported to have an anti-tumor effect, indicating that tocotrienol may serve as an effective agent in the prevention and/or treatment of cancer.On the other hand, focusing on cancer prevention, anti-angiogenic therapy is now a recognized new strategy. Since oxidative stress has been implicated in angiogenesis, it is believed that vitamin E may prevent angiogenesis. Angiogenesis, the process of forming new blood vessels from an existing vascular bed, normally involves a series of steps, which include endothelial cell activation, breakdown of the basement membrane, migration, proliferation, and tube formation of the endothelial cell. Thus, the purpose of this study was to obtain direct evidence on the effects of vitamin E, especially for tocotrienol, on proliferation, migration, and tube formation of bovine aortic endothelial cells (BAEC), the key steps of angiogenesis.
Inhibition of tumour promotion by various palm-oil tocotrienols. Goh, S.H., et.al (1994). Int J Cancer.	Inhibition of tumour promotion by various vitamin E compounds (tocopherols and tocotrienols) and some of their dimers was examined by an in vitro assay utilizing the activation of Epstein-Barr virus (EBV) early antigen (EA) expression in EBV-genome-carrying human lymphoblastoid cells. The results reveal that gamma- and delta-tocotrienols derived from palm oil exhibit a strong activity against tumour promotion by inhibiting EBV EA expression in Raji cells induced by 12-O-tetradecanoylphorbol-13-acetate (TPA). However, alpha- and gamma-tocopherols and dimers of gamma-tocotrienol or gamma-tocopherol lack this activity.

Review Article
Article	Study objectives/ findings
Tocotrienols Regulate Bone Loss through Suppression on Osteoclast Differentiation and Activity: A Systematic Review Radzi, et al., (2018). Current Drug Target.	There are accumulating studies reporting vitamin E in general exhibits bone protective effects. This systematic review, however discusses the effects of a group of vitamin E isomers, tocotrienols in preventing bone loss through osteoclast differentiation and activity suppression. This review is aimed to discuss the literature reporting the effects of tocotrienols on osteoclasts, the cells specialized for resorbing bone. Literature search for relevant studies was conducted using SCOPUS and PUBMED MEDLINE. The inclusion criteria were original research articles published that reported the effect of any tocotrienol isomers or treatment with mixture containing tocotrienols on osteoclasts. Out of the total 22 studies from the literature search, only 11 of them were identified as relevant, which comprised of eight animal studies, two in vitro studies and only one combination of both. The in vivo studies indicated that tocotrienols improve the bone health and reduce bone loss via inhibition of osteoclast formation and resorption activity, which could be through regulation of RANKL and OPG expression as seen from their levels in the sera. This is well supported by data from the in vitro studies demonstrating the suppression of osteoclast formation and resorption activity following treatment with tocotrienol isomers. Thus, tocotrienols are suggested to be potential antioxidants for prevention and treatment of bone-related diseases characterized by increased bone loss.
Potential roles of vitamin E in age-related changes in skeletal muscle health Chung, E., et al., (2017). Nutrition Research	Skeletal muscle disorders including sarcopenia are prevalent during the complex biological process of aging. Loss of muscle mass and strength commonly seen in sarcopenia is induced by impaired neuromuscular innervation, transition of skeletal muscle fiber type, and reduced muscle regenerative capacity, all attributable to chronic inflammation, oxidative stress, and mitochondrial dysfunction. Current literature suggests that vitamin E molecules (α-, β-, γ-, δ-tocopherols and the corresponding tocotrienols) with their antioxidant and anti-inflammatory capabilities may mitigate age-associated skeletal dysfunction and enhance muscle regeneration, thus attenuating sarcopenia. Preclinical and human experimental studies show that vitamin E benefits myoblast proliferation, differentiation, survival, membrane repair, mitochondrial efficiency, muscle mass, muscle contractile properties, and exercise capacity. Limited number of human cross-sectional observational studies reveal positive associations between serum tocopherol level and muscle strength. Several factors, including difficulties in validating vitamin E intake and deficiency, variations in muscle-protective activity and metabolism of diverse forms of vitamin E, and lack of understanding of the mechanisms of action, preclude randomized clinical trials of vitamin E in people with sarcopenia. Future research should consider longterm clinical trials of with adequate sample size, advanced imaging technology and omics approaches to investigate underlying mechanisms and assess clinically meaningful parameters such as muscle strength, physical performance, and muscle mass in sarcopenia prevention and/or treatment.
Tocopherol and Tocotrienol: Therapeutic Potential in Animal Models of Stress Azlina, M. F. N., et al., (2017). Current Drug Targets.	Scientific reports had shown that stress is related to numerous pathological changes in the body. These pathological changes can bring about numerous diseases and can significantly cause negative effects in an individual. These include gastric ulcer, liver pathology and neurobehavioral changes. A common pathogenesis in many diseases related to stress involves oxidative damage. Therefore, the administration of antioxidants such as vitamin E is a reasonable therapeutic approach. However, there is conflicting evidence about antioxidant supplementation. The aim of this work was to summarize documented reports on the effects of tocopherol and tocotrienol on various pathological changes induced by stress. This review will reveal the scientific evidence of enteral supplementation of vitamin E in the forms of tocotrienol and tocopherol in animal models of stress. These models mimic the stress endured by critically ill patients in a clinical setting and psychological stress in individuals. Positive outcomes from enteral feeding of vitamin E in reducing the occurrence of stress-induced pathological changes are discussed in this review. These positive findings include their ability to reduced stress-induced gastric ulcers, elevated liver enzymes and improved locomotors activity. Evidences showing tocotrienol and tocopherol effects are not just related to its ability to reduce oxidative stress but also acting on other mechanism are discussed.
Tocotrienols for bone health: a translational approach Shen, C. L., et al., (2017). Annals of the New York Academy of Sciences	Osteoporosis, a degenerative bone disease, is characterized by low bone mass and microstructural deterioration of bone tissue resulting in aggravated bone fragility and susceptibility to fractures. The trend of extended life expectancy is accompanied by a rise in the prevalence of osteoporosis and concomitant complications in the elderly population. Epidemiological evidence has shown an association between vitamin E consumption and the prevention of age-related bone loss in elderly women and men. Animal studies show that ingestion of vitamin E, especially tocotrienols, may benefit bone health in terms of maintaining higher bone mineral density and improving bone microstructure and quality. The beneficial effects of tocotrienols on bone health appear to be mediated via antioxidant/anti-inflammatory pathways and/or 3-hydroxy-3-methylglutaryl coenzyme A mechanisms. This review discuss (1) an overview of the prevalence and etiology of osteoporosis, (2) types of vitamin E (tocopherols versus tocotrienols), (3) findings of tocotrienols and bone health from published in vitro and animal studies, (4) possible mechanisms involved in bone protection, and (5) challenges and future direction for research.
Tocopherols and Tocotrienols in Common and Emerging Dietary Sources: Occurrence, Applications, and Health Benefits. Shahidi & de Camargo. (2016). Int J Mol Sci.	Edible oils are the major natural dietary sources of tocopherols and tocotrienols, collectively known as tocols. Plant foods with low lipid content usually have negligible quantities of tocols. However, seeds and other plant food processing by-products may serve as alternative sources of edible oils with considerable contents of tocopherols and tocotrienols. Tocopherols are among the most important lipid-soluble antioxidants in food as well as in human and animal tissues. Tocopherols are found in lipid-rich regions of cells (e.g., mitochondrial membranes), fat depots, and lipoproteins such as low-density lipoprotein cholesterol. Their health benefits may also be explained by regulation of gene expression, signal transduction, and modulation of cell functions. Potential health benefits of tocols include prevention of certain types of cancer, heart disease, and other chronic ailments. Although deficiencies of tocopherol are uncommon, a continuous intake from common and novel dietary sources of tocopherols and tocotrienols is advantageous. Thus, this contribution will focus on the relevant literature on common and emerging edible oils as a source of tocols. Potential application and health effects as well as the impact of new cultivars as sources of edible oils and their processing discards are presented. Future trends and drawbacks are also briefly covered.
Tocotrienol and Its Role in Chronic Diseases. Chin, K.Y., et.al (2016). Adv Exp Med Biol.	This review aimed to discuss the health beneficial effects of tocotrienol, specifically in preventing or treating hyperlipidaemia, diabetes mellitus, osteoporosis and cancer with respect to these properties. Evidence from in vitro, in vivo and human studies has been examined. It is revealed that tocotrienol shows promising effects in preventing or treating the health conditions previously mentioned in in vivo and in vitro models. In some cases, alpha-tocopherol attenuates the biological activity of tocotrienol. Except for its cholesterol-lowering effects, data on the health-promoting effects of tocotrienol in human are limited. As a conclusion, the encouraging results on the health beneficial effects of tocotrienol should motivate researchers to explore its potential use in human.
Synergistic Anticancer Effect of Tocotrienol Combined with Chemotherapeutic Agents or Dietary Components: A Review. Eitsuka, T., et.al (2016). Int J Mol Sci.	Tocotrienol (T3), unsaturated vitamin E, is gaining a lot of attention owing to its potent anticancer effect, since its efficacy is much greater than that of tocopherol (Toc). Various factors are known to be involved in such antitumor action, including cell cycle arrest, apoptosis induction, antiangiogenesis, anti-metastasis, nuclear factor-κB suppression, and telomerase inhibition. Owing to a difference in the affinity of T3 and Toc for the α-tocopherol transfer protein, the bioavailability of orally ingested T3 is lower than that of Toc. Furthermore, cellular uptake of T3 is interrupted by coadministration of α-Toc in vitro and in vivo. Based on this, several studies are in progress to screen for molecules that can synergize with T3 in order to augment its potency. Combinations of T3 with chemotherapeutic drugs (e.g., statins, celecoxib, and gefitinib) or dietary components (e.g., polyphenols, sesamin, and ferulic acid) exhibit synergistic actions on cancer cell growth and signaling pathways. In this review, we summarize the current status of synergistic effects of T3 and an array of agents on cancer cells, and discuss their molecular mechanisms of action. These combination strategies would encourage further investigation and application in cancer prevention and therapy.
Tocotrienol and cancer metastasis. De Silva, et.al (2016). Biofactors.	Tumor metastasis involves some of the most complex and dynamic processes in cancer, often leading to poor quality of life and inevitable death. The search for therapeutic compounds and treatment strategies to prevent and/or manage metastasis is the ultimate challenge to fight cancer. In the past two decades, research focus on vitamin E has had a shift from saturated tocopherols to unsaturated tocotrienols (T3). Despite sharing structural similarities with tocopherols, T3 strive to gain scientific prominence due to their anti-cancer effects. Recent studies have shed some light on the anti-metastatic properties of T3. In this review, the roles of T3 in each step of the metastatic process are discussed. During the invasion process, signaling pathways that regulate the extracellular matrix and tumor cell motility have been reported to be modulated by T3. Although studies on T3 and tumor cell migration are fairly limited, they were shown to play a vital role in the suppression of angiogenesis. Furthermore, the anti-inflammatory effect of T3 could be highly promising in the regulation of tumor microenvironment, which is crucial in supporting tumor growth in distant organs.
Effect of PEG surface conformation on anticancer activity and blood circulation of nanoemulsions loaded with tocotrienol-rich fraction of palm oil. Alayoubi, A., et.al (2013). AAPS.	The objective of this study was to investigate the effect of surface grafted polyethylene glycol (PEG) on the properties of the nanoemulsions. PEGylation was achieved by the addition of equimolar PEG groups using poloxamer or 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene glycol)2000] (PEG2000-DSPE). Reduced activity in vitro and improved circulation time suggested strong shielding of plasma proteins from the droplets. Differences between the nanoemulsions were attributed to polymer imbibitions and the differences in PEG conformation and density on the surface of the droplets.
Tocoperhols and tocotrienols as anticancer treatment for lung cancer: future nutrition. Zarogoulidis, P., et.al (2013). J Thorac Dis.	Nutrition has been known for ages to shield the immune system against several formulations that deregulate normal DNA repair mechanisms, and induce tumorigenesis. Vitamins and in specific Vit E and its members tocopherols (α-, β-, γ-, δ-) and tocotrienols (α-, β-, γ-, δ-) have demonstrated strong association with the prevention of cancer and inhibition of tumor, both in vitro and in vivo. Vitamin E has also demonstrated effective role against chemotherapy resistant cancer cell evolution and a protective role in acute interstitial disease. Several formulations of Vitamin E have been investigated conjugated with different carriers as nano-formulations and administered in different forms. Additionally, several tumorigenic pathways have been investigated separately in an effort to identify which member of Vitamin E inhibits efficiently every pathway. Vitamin E presented efficiency against specific subhistology types of lung cancer. Finally, in the current work up to date information regarding novel formulations with Vitamin E and inhibition pathways are going to be presented and commented.
Redox-inactive analogue of tocotrienol as a potential anti-cancer agent. Yano, T., et.al (2013). Anticancer Agents Med Chem.	Vitamins are prominent among natural or endogenous compounds that are considered to be beneficial for both prevention and therapy of various human ailments. The vitamin E group of compounds composed of tocopherol and tocotrienol isoforms, has been subsequently proven to have health benefits including antioxidant and related protective properties. However, individual isoforms exhibit a wide-range of antioxidant potencies. Tocotrienol (T3) displays powerful anticancer activity that is often not exhibited by tocopherols, by modulating multiple intracellular signaling pathways associated with tumor cell proliferation and survival. The anticancer effect of T3 remains not fully understood but generally is mediated independently of its antioxidant activity. Further we have synthesized a new redox-inactive analogue of T3, 6-O-carboxypropyl-α-tocotrienol (T3E) showing considerable promise for stronger anticancer potency than its mother compound. In this mini-review, we particularly focus upon the anticancer action of the above active components of vitamin E and describe current research on the anticancer effects of T3 irrespective of antioxidant activity.
Redox-inactive analogue of tocotrienol as a potential anti-cancer agent. Yano, T., et.al (2012). Anticancer Agents Med Chem.	Vitamins are prominent among natural or endogenous compounds that are considered to be beneficial for both prevention and therapy of various human ailments. The vitamin E group of compounds composed of tocopherol and tocotrienol isoforms, has been subsequently proven to have health benefits including antioxidant and related protective properties. However, individual isoforms exhibit a wide-range of antioxidant potencies. Tocotrienol (T3) displays powerful anticancer activity that is often not exhibited by tocopherols, by modulating multiple intracellular signaling pathways associated with tumor cell proliferation and survival. The anticancer effect of T3 remains not fully understood but generally is mediated independently of its antioxidant activity. Further we have synthesized a new redox-inactive analogue of T3, 6-O-carboxypropyl-α-tocotrienol (T3E) showing considerable promise for stronger anticancer potency than its mother compound. In this mini-review, we particularly focus upon the anticancer action of the above active components of vitamin E and describe current research on the anticancer effects of T3 irrespective of antioxidant activity.
Tocotrienol as a potential anticancer agent. Ling M.T., et.al (2012). Carcinogenesis.	Vitamin E is composed of two structurally similar compounds: tocopherols (Ts) and tocotrienols (T3). Despite being overshadowed by TP over the past few decades, T3 is now considered to be a promising anticancer agent due to its potent effects against a wide range of cancers. A growing body of evidence suggests that in addition to its antioxidative and pro-apoptotic functions, T3 possesses a number of anticancer properties that make it superior to T. These include the inhibition of epithelial-to-mesenchymal transitions, the suppression of vascular endothelial growth factor tumor angiogenic pathway and the induction of antitumor immunity. More recently, T3, but not T, has been shown to have chemosensitization and anti-cancer stem cell effects, further demonstrating the potential of T3 as an effective anticancer therapeutic agent. With most of the previous clinical studies on T producing disappointing results, research has now focused on testing T3 as the next generation vitamin E for chemoprevention and cancer treatment. This review will summarize recent developments in the understanding of the anticancer effects of T3. The current progress in clinical trials involving T3 as an adjuvant to conventional cancer therapy will be discussed as well.
Tocotrienols fight cancer by targeting multiple cell signaling pathways Kannappan, R., et.al (2012). Genes Nutr.	Cancer cells are distinguished by several distinct characteristics, such as self-sufficiency in growth signal, resistance to growth inhibition, limitless replicative potential, evasion of apoptosis, sustained angiogenesis, and tissue invasion and metastasis. Tumor cells acquire these properties due to the dysregulation of multiple genes and associated cell signaling pathways, most of which are linked to inflammation. For that reason, rationally designed drugs that target a single gene product are unlikely to be of use in preventing or treating cancer. Moreover, targeted drugs can cause serious and even life-threatening side effects. Therefore, there is an urgent need for safe and effective promiscuous (multitargeted) drugs. “Mother Nature” produces numerous such compounds that regulate multiple cell signaling pathways, are cost effective, exhibit low toxicity, and are readily available. One among these is tocotrienol, a member of the vitamin E family, which has exhibited anticancer properties. This review summarizes data from in vitro and in vivo studies of the effects of tocotrienol on nuclear factor-κB, signal transducer and activator of transcription (STAT) 3, death receptors, apoptosis, nuclear factor (erythroid-derived 2)-like 2 (Nrf2), hypoxia-inducible factor (HIF) 1, growth factor receptor kinases, and angiogenic pathways.
Why tocotrienols work better: insights into the in vitro anti-cancer mechanism of vitamin E Viola, V., et.al (2012). Genes Nutr.	The selective constraint of liver uptake and the sustained metabolism of tocotrienols (T3) demonstrate the need for a prompt detoxification of this class of lipophilic vitamers, and thus the potential for cytotoxic effects in hepatic and extra-hepatic tissues. Hypomethylated (γ and δ) forms of T3 show the highest in vitro and in vivo metabolism and are also the most potent natural xenobiotics of the entire vitamin E family of compounds. These stimulate a stress response with the induction of detoxification and antioxidant genes. Depending on the intensity of this response, these genes may confer cell protection or alternatively they stimulate a senescence-like phenotype with cell cycle inhibition or even mitochondrial toxicity and apoptosis. In cancer cells, the uptake rate and thus the cell content of these vitamers is again higher for the hypomethylated forms, and it is the critical factor that drives the dichotomy between protection and toxicity responses to different T3 forms and doses. These aspects suggest the potential for marked biological activity of hypomethylated “highly metabolized” T3 that may result in cytoprotection and cancer prevention or even chemotherapeutic effects. Cytotoxicity and metabolism of hypomethylated T3 have been extensively investigated in vitro using different cell model systems that will be discussed in this review paper as regard molecular mechanisms and possible relevance in cancer therapy.
Synergistic anticancer effects of combined γ-tocotrienol with statin or receptor tyrosine kinase inhibitor treatment Sylvester, P.W. (2012). Genes Nutr.	Systemic chemotherapy is the only current method of treatment that provides some chance for long-term survival in patients with advanced or metastatic cancer. γ-Tocotrienol is a natural form of vitamin E found in high concentrations in palm oil and displays potent anticancer effects, but limited absorption and transport of by the body has made it difficult to obtain and sustain therapeutic levels in the blood and target tissues. Statins are inhibitors of 3-hydroxy-3-methylglutaryl-coenzyme A (HMGCoA) reductase and are an example of a promising cancer chemotherapeutic agent whose clinical usefulness has been limited due to high-dose toxicity. Similarly, erlotinib and gefitinib are anticancer agents that inhibit the activation of individual HER/ErbB receptor subtypes, but have shown limited clinical success because of heterodimerization between different EGF receptor family members that can rescue cancer cells from agents directed against a single receptor subtype. Recent studies have investigated the anticancer effectiveness of low-dose treatment of various statins or EGF receptor inhibitors alone and in combination with γ-tocotrienol on highly malignant +SA mouse mammary epithelial cells in vitro. Combined treatment with subeffective doses of γ-tocotrienol with these other chemotherapeutic agents resulted in a synergistic inhibition of +SA cell growth and viability. These findings strongly suggest that combined treatment of γ-tocotrienol with other anticancer agents may not only provide an enhanced therapeutic response but also provide a means to avoid the toxicity, low bioavailability, or limited therapeutic action associated with high-dose monotherapy.
Delivery of the vitamin E compound tocotrienol to cancer cells. Dufes, C. (2011). Ther Deliv.	Tocotrienol, a member of the vitamin E family of compounds, is currently receiving increased attention owing to its highly promising anticancer effects. However, its potential in cancer therapy is limited by its poor bioavailability and its inability to specifically reach tumors at therapeutic concentrations after intravenous administration. In order to address these problems, various delivery strategies have been proposed, such as the inclusion of tocotrienol in gamma-cyclodextrins, prodrugs and emulsions, and entrapment in lipid nanoparticles and vesicles. Among these approaches, we have demonstrated that the entrapment of tocotrienol within vesicles bearing transferrin, whose receptors are overexpressed on numerous cancer cells, significantly improved the uptake by cancer cells overexpressing transferrin receptors. Consequently, the intravenous administration of tocotrienol entrapped in transferrin-bearing vesicles led to tumor regression and even complete tumor suppression in some cases in a murine tumor model, as well as improvement of animal survival. Transferrin-bearing vesicles are therefore highly promising for the delivery of tocotrienol to cancer cells in vitro and in vivo, and should be further investigated to optimize the anticancer therapeutic effect of tocotrienol.
Tocotrienol combination therapy results in synergistic anticancer response. Sylvester, P.W., et.al (2011). Front Biosci.	Vitamin E represents a family of compounds that is divided into two subgroups called tocopherols and tocotrienols, which act as important antioxidants that regulate peroxidation reactions and control free-radical production within the body. However, many of the biological effects of vitamin E are mediated independently of its antioxidant activity. Although tocopherols and tocotrienols have the same basic chemical structure characterized by a long phytyl chain attached to a chromane ring, only tocotrienols display potent anticancer activity, by modulating multiple intracellular signaling pathways associated with tumor cell proliferation and survival, and combination therapy with other chemotherapeutic agents result in a synergistic anticancer response. Combination therapy is most effective when tocotrienols are combined with agents that have complementary anticancer mechanisms of action. These findings strongly suggest that the synergistic antiproliferative and apoptotic effects demonstrated by combined low dose treatment of γ-tocotrienol with other chemotherapeutic agents may provide significant health benefits in the prevention and/or treatment of breast cancer in women, while at the same time avoiding tumor resistance and toxic side effects associated with high dose monotherapy.
Tocotrienols, the Vitamin E of the 21^st Century: It’s Potential Against Cancer and Other Chronic Diseases Aggarwal, B.B., et.al (2010). Biochem Pharmacol.	Initially discovered in 1938 as a “fertility factor,” vitamin E now refers to eight different isoforms that belong to two categories, four saturated analogues (α, β, γ, and δ) called tocopherols and four unsaturated analogues referred to as tocotrienols. While the tocopherols have been investigated extensively, little is known about the tocotrienols. Very limited studies suggest that both the molecular and therapeutic targets of the tocotrienols are distinct from those of the tocopherols. For instance, suppression of inflammatory transcription factor NF-κB, which is closely linked to tumorigenesis and inhibition of HMG-CoA reductase, mammalian DNA polymerases and certain protein tyrosine kinases, is unique to the tocotrienols. This review examines in detail the molecular targets of the tocotrienols and their roles in cancer, bone resorption, diabetes, and cardiovascular and neurological diseases at both preclinical and clinical levels. As disappointment with the therapeutic value of the tocopherols grows, the potential of these novel vitamin E analogues awaits further investigation.
Regulation of survival, proliferation, invasion, angiogenesis, and metastasis of tumor cells through modulation of inflammatory pathways by nutraceuticals. Gupta, S.C., et.al (2010). Cancer Metastasis Rev.	Almost 25 centuries ago, Hippocrates, the father of medicine, proclaimed “Let food be thy medicine and medicine be thy food.” Exploring the association between diet and health continues today. For example, we now know that as many as 35% of all cancers can be prevented by dietary changes. Carcinogenesis is a multistep process involving the transformation, survival, proliferation, invasion, angiogenesis, and metastasis of the tumor and may take up to 30 years. The pathways associated with this process have been linked to chronic inflammation, a major mediator of tumor progression. The human body consists of about 13 trillion cells, almost all of which are turned over within 100 days, indicating that 70,000 cells undergo apoptosis every minute. Thus, apoptosis/cell death is a normal physiological process, and it is rare that a lack of apoptosis kills the patient. Almost 90% of all deaths due to cancer are linked to metastasis of the tumor. How our diet can prevent cancer is the focus of this review. Specifically, we will discuss how nutraceuticals, such as allicin, apigenin, berberine, butein, caffeic acid, capsaicin, catechin gallate, celastrol, curcumin, epigallocatechin gallate, fisetin, flavopiridol, gambogic acid, genistein, plumbagin, quercetin, resveratrol, sanguinarine, silibinin, sulforaphane, taxol, gamma-tocotrienol, and zerumbone, derived from spices, legumes, fruits, nuts, and vegetables, can modulate inflammatory pathways and thus affect the survival, proliferation, invasion, angiogenesis, and metastasis of the tumor. Various cell signaling pathways that are modulated by these agents will also be discussed.
The value of tocotrienols in the prevention and treatment of cancer. Sylvester, P.W., et.al (2010). J Am Coll Nutr.	Tocopherols and tocotrienols represent the 2 subgroups that make up the vitamin E family of compounds, but only tocotrienols display potent anticancer activity. Although in vitro experimental evidence has been very promising, oral supplementation of tocotrienols in animal and human studies has produced inconsistent results. However, recent studies have now clarified the reasons for these discrepancies observed between in vitro and in vivo studies. Oral absorption of tocotrienols into the circulation is mediated in large part by carrier transporter systems that display saturation and apparently down-regulation when exposed to high concentrations of tocotrienols. To circumvent these limitations in oral absorption of tocotrienols, investigators have developed novel prodrug derivatives and nanoparticle delivery systems that greatly enhance tocotrienol bioavailability and therapeutic responsiveness. Additional studies have also demonstrated that combined treatment of tocotrienols with other traditional chemotherapeutic agents results in a synergistic anticancer response, and this synergistic response was further enhanced when these agents were encapsulated in a nanoparticle delivery system. Taken together, these findings clarify the limitations of oral tocotrienol administration and provide novel alternative drug-delivery systems that circumvent these limitations and greatly enhance the therapeutic effectiveness of tocotrienols in the prevention and treatment of cancer.
Cancer-preventive activities of tocopherols and tocotrienols Ju, J.Y., et.al (2010). Carcinogenesis.	The cancer-preventive activity of vitamin E has been studied. Whereas some epidemiological studies have suggested a protective effect of vitamin E against cancer formation, many large-scale intervention studies with α-tocopherol (usually large doses) have not demonstrated a cancer-preventive effect. Studies on α-tocopherol in animal models also have not demonstrated robust cancer prevention effects. One possible explanation for the lack of demonstrable cancer-preventive effects is that high doses of α-tocopherol decrease the blood and tissue levels of δ-tocopherols. It has been suggested that γ-tocopherol, due to its strong anti-inflammatory and other activities, may be the more effective form of vitamin E in cancer prevention. Our recent results have demonstrated that a γ-tocopherol-rich mixture of tocopherols inhibits colon, prostate, mammary and lung tumorigenesis in animal models, suggesting that this mixture may have a high potential for applications in the prevention of human cancer. In this review, we discuss biochemical properties of tocopherols, results of possible cancer-preventive effects in humans and animal models and possible mechanisms involved in the inhibition of carcinogenesis. Based on this information, we propose that a γ-tocopherol-rich mixture of tocopherols is a very promising cancer-preventive agent and warrants extensive future research.
Chemoprevention of tocotrienols: The mechanism of antiproliferative effects. Wada, S. (2009). Forum Nutr.	Tocotrienols have been reported as antitumor agents and widely commercialized as an antioxidant dietary supplement. Tocotrienols have more significant biological activity than tocopherols, although serum level of tocotrienols is much lower than that of tocopherols. This may be because intracellular concentration of tocotrienols was revealed to be significantly higher compared with tocopherols, and tocotrienol accumulation is observed in tumor. Previous reports have suggested antiproliferative effect, induction of apoptosis, modulation of cell cycle, antioxidant activity, inhibition of angiogenesis, and suppression of 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase activity as anticarcinogenesis mechanisms of tocotrienols both in vivo and in vitro. Extension of the duration of host survival was observed in tumor-implanted mice treated with tocotrienol. Tocotrienols induce apoptosis mainly via mitochondria-mediated pathway. Cell cycle arrest is due to suppression of cyclin D by tocotrienols. Tocotrienols also inhibit vascularization-reducing proliferation, migration and tube formation. Malignant proliferation demands elevation of HMG CoA reductase activity, and tocotrienols suppress its activity. Tocotrienol treatment decreases oncogene expression and increases the level of tumor suppressors. Only a few clinical trials to determine the effects of tocotrienol on cancer prevention or treatment have been carried out. There is no convincing or probable evidence of the role of tocotrienols in cancer prevention, while alpha-tocopherol has been suggested to have a limited anti-prostate cancer potential. Neither beneficial activity nor adverse effect of tocotrienol has sufficiently been explored so far. The above-mentioned mechanisms of tocotrienols seem to be promising for cancer prevention; however, further clinical studies are warranted to assess the efficacy and safety of tocotrienol.
Multitargeted therapy of cancer by tocotrienols. Naseretnam, K., et.al (2008). Cancer Lett.	Natural compounds with possible health benefits have become attractive targets for research in areas pertaining to human health. For both prevention and therapy of various human ailments, such compounds are preferred over synthetic ones due to their lesser toxicity. They are also easily absorbed and processed by our body. Vitamins are prominent among natural or endogenous compounds that are considered to be beneficial. The vitamin E group of compounds is among the better known of the vitamins due to their suggested health benefits including antioxidant and related protective properties. Among these, tocotrienols have gained prominence in recent years due to their potential applications and better protective effects in certain systems. These tocotrienols are vitamin E derivatives that are analogs of the more established forms of vitamin E namely tocopherols. In addition to their potent antioxidant activity, tocotrienols have other important functions, especially in maintaining a healthy cardiovascular system and a possible role in protection against cancer and other ailments.
Tumor suppressive effects of tocotrienol in vivo and in vitro. Wada, S., et.al (2005). Cancer Lett.	Tocotrienols have been reported to have higher biological activities than tocopherols. We investigated the antitumor effect of tocotrienols both in vivo and in vitro. Oral administration of tocotrienols resulted in significant suppression of liver and lung carcinogenesis in mice. In human hepatocellular carcinoma HepG2 cells, delta-tocotrienol exerted more significant antiproliferative effect than alpha-, beta-, and gamma-tocotrienols. delta-tocotrienol induced apoptosis, and also tended to induce S phase arrest. On the other hand, gene expression analysis showed that delta-tocotrienol increased CYP1A1 gene, a phase I enzyme. Although further study will be necessary to investigate possible adverse effect, the data obtained in present study suggest that tocotrienols could be promising agents for cancer prevention.
Anti-angiogenic activity of tocotrienol Inokuchi, H., et.al (2003). Biosci.Biotechnol Biochem.	The anti-angiogenic property of vitamin E compounds, with particular emphasis on tocotrienol, has been investigated in vitro. Tocotrienol, but not tocopherol, inhibited both the proliferation and tube formation of bovine aortic endothelial cells, with delta-tocotrienol appearing the highest activity. Also, delta-tocotrienol reduced the vascular endothelial growth factor-stimulated tube formation by human umbilical vein endothelial cells. The findings suggest that tocotrienol has potential use as a therapeutic dietary supplement for minimizing tumor angiogenesis.
Isolation and identification of novel tocotrienols from rice bran with hypocholesterolemic, antioxidant and antitumor properties. Qureshi, A.A., et.al (2000). J Agric Food Chem.	Two novel tocotrienols were isolated from stabilized and heated rice bran, apart from the known alpha-, beta-, gamma-, and delta-tocopherols and tocotrienols. These new tocotrienols were separated by HPLC, using a normal phase silica column. Their structures were determined by ultraviolet, infrared, nuclear magnetic resonance, circular dichroism, and high-resolution mass spectroscopies and established as desmethyl tocotrienol [3, 4-dihydro-2-methyl-2-(4,8,12-trimethyltrideca-3′(E),7′(E), 11′-trienyl)-2H-1-benzopyran-6-ol] and didesmethy tocotrienol [3, 4-dihydro-2-(4,8,12-trimethyltrideca-3′(E),7′(E), 11′-trienyl)-2H-1-benzopyran-6-ol]. These tocotrienols significantly lowered serum total and LDL cholesterol levels and inhibited HMG-CoA reductase activity in chickens. They had much greater in vitro antioxidant activities and greater suppression of B16 melanoma cell proliferation than alpha-tocopherol and known tocotrienols. Results indicated that the number and position of methyl substituents in tocotrienols affect their hypocholesterolemic, antioxidant, and antitumor properties.

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