Tocotrienol-rich Fraction Of Palm Oil Research Articles

Palm Oil Derived Tocotrienol-Rich Fraction Attenuates Vascular Dementia in Type 2 Diabetic Rats.

Vascular dementia (VaD) is a serious global health issue and type 2 diabetes mellitus (T2DM) patients are at higher risk. Palm oil tocotrienol-rich fraction (TRF) exhibits neuroprotective properties; however, its effect on VaD is not reported. Hence, we evaluated TRF effectiveness in T2DM-induced VaD rats. Rats were given a single dose of streptozotocin (STZ) and nicotinamide (NA) to develop T2DM. Seven days later, diabetic rats were given TRF doses of 30, 60, and 120 mg/kg orally for 21 days. The Morris water maze (MWM) test was performed for memory assessment. Biochemical parameters such as blood glucose, plasma homocysteine (HCY) level, acetylcholinesterase (AChE) activity, reduced glutathione (GSH), superoxide dismutase (SOD) level, and histopathological changes in brain hippocampus and immunohistochemistry for platelet-derived growth factor-C (PDGF-C) expression were evaluated. VaD rats had significantly reduced memory, higher plasma HCY, increased AChE activity, and decreased GSH and SOD levels. However, treatment with TRF significantly attenuated the biochemical parameters and prevented memory loss. Moreover, histopathological changes were attenuated and there was increased PDGF-C expression in the hippocampus of VaD rats treated with TRF, indicating neuroprotective action. In conclusion, this research paves the way for future studies and benefits in understanding the potential effects of TRF in VaD rats.

Novel self-nanoemulsifying drug-delivery system enhances antileukemic properties of all-trans retinoic acid.

Aim: All-trans retinoic acid (ATRA) shows erratic oral bioavailability when administered orally against leukemia, which can be solved through its incorporation in self-nanoemulsifying drug-delivery systems (SEDDS). The SEDDS developed contained a hydrophobic ion pair between benzathine (BZT) and ATRA and was enriched with tocotrienols by the input of a palm oil tocotrienol rich fraction (TRF) in its composition. Results: SEDDS-TRF-ATRA-BZT allowed the formation of emulsions with nanometric size that retained ATRA within their core after dispersion. Pharmacokinetic parameters after oral administration of SEDDS-TRF-ATRA-BZT in mice were improved compared with what was seen for an ATRA solution. Moreover, SEDDS-TRF-ATRA-BZT had improved activity against HL-60 cells compared with SEDDS without TRF. Conclusion: SEDDS-TRF-ATRA-BZT is a promising therapeutic choice over ATRA conventional medicine.

Tocotrienol-Rich Fraction (TRF) Improves the Viability of Wild-Type Saccharomyces cerevisiae in the Initial Stationary Phase

Palm oil tocotrienol-rich fraction (TRF) containing majorly of αα, β, γ andδδ-tocotrienols and some αα-tocopherols, was reported to have anti-ageing effects in both human and non-human model organisms, but still remains unexplored in the yeast model. It was reported to have the ability to extend the lifespan of several organisms. Chronological lifespan is one of the means to measure ageing in yeasts. The effect of TRF on the viability of three strains of Saccharomyces cerevisiae (wild-type, CTT1ΔΔand GPx2Δ) was studied. Phenotypic growth analysis of all strains was carried out for 15 hours by measuring the absorbance at OD600nm and cell counting. The optimum dose of TRF was optimised by determining the number of colony-forming unit by the wild-type strain at the end of a 24-hour treatment with TRF (ranging from 0μg/ml to 300μg/ml). TRF at 300 μg/mL showed the best result, and selected as a working dose. Treatment of cells with 300 μg/mL of TRF improved the viability of the wild-type strain in the initial stationary phase, but not on the knockout strains. These finding suggests that TRF has a potential in prolonging the chronological lifespan of S. cerevisiae, and perhaps other organisms as well.

Tocotrienol-Rich Fraction of Palm Oil Improves Behavioral Impairments and Regulates Metabolic Pathways in AβPP/PS1 Mice.

We have recently shown that the tocotrienol-rich fraction (TRF) of palm oil, a mixture of vitamin E analogs, improves amyloid pathology in vitro and in vivo. However, precise mechanisms remain unknown. In this study, we examined the effects of long-term (10 months) TRF treatment on behavioral impairments and brain metabolites in (15 months old) AβPP/PS1 double transgenic (Tg) Alzheimer’s disease (AD) mice. The open field test, Morris water maze, and novel object recognition tasks revealed improved exploratory activity, spatial learning, and recognition memory, respectively, in TRF-treated Tg mice. Brain metabolite profiling of wild-type and Tg mice treated with and without TRF was performed using ultrahigh performance liquid chromatography (UHPLC) coupled to high-resolution accurate mass (HRAM)-orbitrap tandem mass spectrometry (MS/MS). Metabolic pathway analysis found perturbed metabolic pathways that linked to AD. TRF treatment partly ameliorated metabolic perturbations in Tg mouse hippocampus. The mechanism of this pre-emptive activity may occur via modulation of metabolic pathways dependent on Aβ interaction or independent of Aβ interaction.

Altering Gut Microbiota to Enhance Therapeutic Efficacy in Resistant Colon Cancer by Natural Agents

Tumor recurrence, a common phenomenon in allmalignancies, is observed in nearly 50% of patients with colorectal cancer(CRC) and is thought to be due to re‐emergence of chemotherapy‐resistant cancerstem cells (CSCs), accompanied by dysfunction of the gut microbiota, resultingin alterations in microbial metabolites. Thus,development of preventive/therapeutic strategies that specifically target CSCs and are non‐toxic should be effective in reducing the risk of relapse. We have examinedthe effectiveness of the combination of ETO‐Curcumin (ETO‐Cur; curcumincomplexed with essential turmeric oil) and TRF, tocotrienol‐rich fractionof palm oil in inhibiting the growth of SCID micexenograft of chemo‐resistant (CR) colon cancer cells and whether thisinhibition could be attributed to alterations in gut and tumor microbiome andtheir metabolites. Indeed, feeding of ETO‐Cur (5 mg/kg) together with TRF (2mg/kg) or the vehicle (control) in SCID mice, initiated after 20 days of inoculationof CR colon cancer cells and continued for 45 days resulted in inhibition of growth of colon tumor that became apparent after about 30 days of treatment. At 40‐ and 45 days, differences in tumor growth between the two groups were highly significant. This inhibition of growth was associated with a significant down‐regulation of β‐catenin and TNF‐α in the tumor and alterations in gutmicrobiota. The latter was evidenced by an overlap of 340 bacterial species between the control and ETO‐Cur/TRF‐treated mice, and 216 and 120 species being different in ETO‐Cur/TRF treated and control mice, respectively. ETO‐Cur/TRF treatment also caused a significant reduction in the expression of 7‐α‐dehydroxylase, an enzyme that catalyzes the conversion of primary to secondary bile acids. Since secondary bile acids, specifically deoxycholic acid (DCA) and lithocholic acid(LCA) are known for their co‐carcinogenic activity and to induce CSCs in the colon, we postulate that the decreased conversion of primary bile acids in the gut to secondary bile acids DCA and/or LCA by ETO‐Cur/TRF may have contributed to the suppression of growth of colon tumors.Support or Funding InformationNIH and Department of Veterans AffairsThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Gamma-Tocotrienol Stimulates the Proliferation, Differentiation, and Mineralization in Osteoblastic MC3T3-E1 Cells

Gamma-tocotrienol, a major component of tocotrienol-rich fraction of palm oil, has been suggested to exhibit bone protective effectsin vivo. However, the effects ofγ-tocotrienol on osteoblast cells are still unclear. In this study, the effects ofγ-tocotrienol on the proliferation, differentiation, and mineralization in osteoblastic MC3T3-E1 cells were investigated. Our results showed thatγ-tocotrienol (2–8 μmol/L) significantly improved the cell proliferation (p<0.05), but it did not affect cell cycle progression.γ-Tocotrienol significantly increased alkaline phosphatase (ALP) activity (p<0.05), secretion levels of osteocalcin (OC) and osteonectin (ON), and mRNA levels of collagen type I (Col I) of MC3T3-E1 cells. Meanwhile, we found thatγ-tocotrienol is promoted in differentiation MC3T3-E1 cells by upregulation of the expression of Runx2 protein. Moreover, the number of bone nodules increased over 2.5-fold in cells treated withγ-tocotrienol (2–8 μmol/L) for 24 d compared to control group. These results indicated thatγ-tocotrienol at low dose levels, especially 4 μmol/L, could markedly enhance the osteoblastic function by increasing the proliferation, differentiation, and mineralization of osteoblastic MC3T3-E1 cells. Moreover, our data also indicated that Runx2 protein may be involved in these effects. Further studies are needed to determine the potential ofγ-tocotrienol as an antiosteoporotic agent.

The Effect of Tocotrienol-Rich Fraction on Oxidative Liver Damage Induced by Fenitrothion

Exposure to organophosphate pesticide including fenitrothion (FNT) has led to many adverse effects on human health. However, a potent antioxidant like palm oil tocotrienol-rich fraction (TRF) can reduce oxidative damage in various pathological conditions, could also reduce the adverse effects by FNT. The aim of this study was to evaluate the effect of TRF on oxidative liver damage in FNT induce hepatotoxicity in experimental rats. A total of 40 male Sprague-Dawley rats were randomly divided into four groups of 10, namely control, TRF, FNT and TRF+FNT group. TRF (200 mg/kg body weight) and FNT (20 m/kg body weight) were administered through oral gavage for 28 days. Corn oil which served as vehicle was given orally to the control group. At the end of the study period, liver and blood was taken for oxidative damage and biochemical evaluation and histological observation. TRF supplementation prevents oxidative liver damage by reducing the hepatic malondialdehyde (MDA) and protein carbonyl (PCO) level significantly. Besides, TRF also restored the endogenous antioxidants particularly reduced glutathione (GSH), glutathione peroxidase (GPx) and ferric reducing/antioxidant power (FRAP). TRF also prevent liver damage by reducing the liver enzymes, alanine aminotransferase (ALT) and aspartate aminotransferase (AST). The attenuation of liver damage by TRF was also showed histologically. In conclusion, TRF supplementation showed a potential in preventing oxidative liver damage in FNT-treated rats by reducing the oxidative damage and improving the antioxidant status.

Comparing palm oil tocotrienol rich fraction with α-tocopherol supplementation on oxidative stress in healthy older adults

Vitamin E is a fat-soluble compound and powerful antioxidant that have been shown to protect the cell membranes against damage caused by free radicals. Human vitamin E supplementation studies are usually limited to α-tocopherol but currently tocotrienols are also available. This study aims to compare the effects of tocotrienol rich fraction (TRF) with α-tocopherol (α-TF) supplementation on oxidative stress in healthy male and female older adults aged 50-55 years old. A total of 71 subjects both male and female aged between 50 and 55 years were divided into groups receiving placebo (n=23), α-TF (n=24) and TRF (n=24) for six months. Blood was taken at baseline (month 0), 3 months and 6 months osf supplementation for determination of plasma malondialdehyde (MDA), protein carbonyl, total DNA damage, vitamin D concentration and vitamin E isomers. α-TF supplementation reduced plasma MDA and protein carbonyl in female subjects after 3 and 6 months. TRF supplementation reduced MDA levels in both males and females as early as 3 months while DNA damage was reduced in females only at 6 months. Supplementation with α-TF and TRF increased plasma vitamin D concentration in both males and females after 6 months, but vitamin D concentration in male subjects were significantly higher compared to female subjects in TRF group. Vitamin E isomer determination showed α-TF, α-tocotrienol and γ-tocotrienol were increased in both male and female subjects. In conclusion, TRF supplementation effects were different from α-TF in reducing oxidative stress markers and vitamin D levels with a more pronounced effect in female subjects.

Palm oil tocotrienol-rich fraction attenuates testicular toxicity induced by fenitrothion via an oxidative stress mechanism

Palm oil tocotrienol rich fraction reduced the testicular toxicity effects of fenitrothion by its antioxidant properties, involved in the synthesis of DNA repairing enzymes as well as blocking the HSP70 gene transcription.

Fenitrothion alters sperm characteristics in rats: ameliorating effects of palm oil tocotrienol-rich fraction.

Exposure to organophosphate insecticides such as fenitrothion (FNT) in agriculture and public health has been reported to affect sperm quality. Antioxidants may have a potential to reduce spermatotoxic effects induced by organophosphate. The present study was carried out to evaluate the effects of palm oil tocotrienol-rich fraction (TRF) in reducing the detrimental effects occurring in spermatozoa of FNT-treated rats. Adult male Sprague-Dawley rats were divided into four equal groups: a control group and groups of rats treated orally with palm oil TRF (200 mg/kg), FNT (20 mg/kg) and palm oil TRF (200 mg/kg) combined with FNT (20 mg/kg). The sperm characteristics, DNA damage, superoxide dismutase (SOD) activity, and levels of reduced glutathione (GSH), malondialdehyde (MDA), and protein carbonyl (PC) were evaluated. Supplementation with TRF attenuated the detrimental effects of FNT by significantly increasing the sperm counts, motility, and viability and decreased the abnormal sperm morphology. The SOD activity and GSH level were significantly increased, whereas the MDA and PC levels were significantly decreased in the TRF+FNT group compared with the rats receiving FNT alone. TRF significantly decreased the DNA damage in the sperm of FNT-treated rats. A significant correlation between abnormal sperm morphology and DNA damage was found in all groups. TRF showed the potential to reduce the detrimental effects occurring in spermatozoa of FNT-treated rats.

Fenitrothion Alters Sperm Characteristics in Rats: Ameliorating Effects of Palm Oil Tocotrienol-Rich Fraction

Fenitrothion Alters Sperm Characteristics in Rats: Ameliorating Effects of Palm Oil Tocotrienol-Rich Fraction

Effect of PEG surface conformation on anticancer activity and blood circulation of nanoemulsions loaded with tocotrienol-rich fraction of palm oil.

Tocotrienol-rich fraction of palm oil, which contains the isomers of vitamin E, was shown to possess potent anticancer activity against mammary adenocarcinoma cell lines. Its clinical use, however, is limited by poor oral bioavailability and short half-life. Previously, we developed tocotrienol-rich lipid nanoemulsions for intravenous administration. 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). The effect of PEG surface topography on the antiproliferative activity of nanoemulsions against mammary adenocarcinoma cells, their susceptibility to protein adsorption, and its effect on blood hemolysis and circulation time was investigated. Nanoemulsions PEGylated with poloxamer or PEG2000-DSPE were stable under physical stress. Poloxamer nanoemulsion, however, displayed higher uptake and potency against MCF-7 tumor cells in 2D and 3D culture and increased hemolytic effect and susceptibility to IgG adsorption, which was reflected in its rapid clearance and short circulation half-life (1.7 h). Conversely, PEGylation with PEG2000-DSPE led to a 7-fold increase in mean residence time (12.3 h) after IV injection in rats. 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.

γ-Tocotrienol Induces Paraptosis-Like Cell Death in Human Colon Carcinoma SW620 Cells

Colorectal cancer is one of the most serious illnesses among diagnosed cancer. As a new type of anti-cancer composition from tocotrienol-rich fraction of palm oil, γ-tocotrienol is widely used in anti-cancer research. The objectives of this study were to investigate the effects of γ-tocotrienol on human colon cancer SW620 and HCT-8 cells. We showed that treatment with different concentrations of γ-tocotrienol resulted in a dose dependent inhibition of cell growth. Cell death induced by γ-tocotrienol was mediated by a paraptosis-like cell death in SW620 and HCT-8 cells. Real-time RT-PCR and western blot analyses showed that γ-tocotrienol inhibited the expression level of β-catenin, cyclin D1 and c-jun. These data suggest that a paraptosis-like cell death induced by γ-tocotrienol in SW620 cells is associated with the suppression of the Wnt signaling pathway, which offers a novel tool for treating apoptosis-resistance colon cancer.

Antioxidant Activity of Tocotrienol Rich Fraction Prevents Fenitrothion-induced Renal Damage in Rats

Fenitrothion (FNT) is an organophosphate compound widely used as pesticide in Malaysia. The present study aims to investigate effects of palm oil tocotrienol rich fraction (TRF) on the renal damage of FNT-treated rats. A total of 40 male Sprague Dawley rats were divided into 4 groups randomly, the control, TRF, FNT and FNT+TRF groups. FNT (20 mg/kg b.w.) and TRF (200 mg/kg b.w.) were given orally for 28 days continuously. Rats from the FNT+TRF group were supplemented with TRF 30 minutes prior to administration of FNT. Rats were sacrificed after 28 days, and the kidneys were removed for determination of oxidative stress and histological analysis. Plasma was collected for determination of blood creatinine and urea level. Statistical analysis showed that palm oil TRF has a protective effect against renal oxidative damage induced by FNT. In the FNT+TRF group, malondialdehyde and protein carbonyl levels were significantly lower, while the glutathione level as well as superoxide dismutase and catalase activities were significantly higher compared with the FNT-treated group (p<0.05). As for renal function, there was a markedly lower urea level (p<0.05) in the FNT+TRF group compared with the FNT-treated group, but there was no significant difference in creatinine level. Besides, total protein also showed no significant difference for all groups of rats (p>0.05). Histological evaluation also revealed that the FNT+TRF group had less glomerulus and renal tubule damage than the FNT-treated group. In conclusion, palm oil TRF was able to reduce oxidative stress and renal damage in FNT-treated rats.

Tocotrienol Rich Fraction Prevents Fenitrothion Induced Pancreatic Damage by Restoring Antioxidant Status

Fenitrothion (FNT) is extensively used as pesticide and may induce oxidative stress in various organs. Tocotrienol, a form of vitamin E found in palm oil, reduces oxidative impairments in pathological conditions. This study aims to investigate the effects of palm oil tocotrienol rich fraction (TRF) on fenitrothion-induced oxidative damage in rat pancreas. Forty male Sprague-Dawley rats were divided into four groups: control group, FNT group, TRF group and FNT+TRF group. Regimens FNT (20 mg kg(-1) b.wt.) and TRF (200 mg kg(-1) b.wt.) were force-fed for 28 consecutive days with control group only receiving corn oil. Chronic administration of fenitrothion significantly (p < 0.05) induced oxidative damage in pancreas of rats with elevated malondialdehyde and protein carbonyl level. Depletion of glutathione and significant (p < 0.05) reduction in antioxidant enzyme activities in pancreas homogenate additionally suggested induction of oxidative stress. Despite these changes in pancreas of intoxicated rats, no significant (p < 0.05) changes in blood glucose and pancreas histology were observed. Co-administration of FNT with TRF alleviated these oxidative changes and significantly (p < 0.05) restored antioxidant status. Enzymatic activities of Superoxide Dismutase (SOD) and Catalase (CAT) were normalized. In conclusion, tocotrienol rich fraction of palm oil prevents fenitrothion-induced pancreatic oxidative damage in rats.

Assessment of the protective effects of oral tocotrienols in arginine chronic-like pancreatitis

Tocotrienols exhibit anti-inflammatory properties over macrophages and promote cytotoxicity in activated pancreatic stellate cells, suggesting that they may limit chronic pancreatitis progression. We aimed to quantitate the effect of oral tocotrienols on a rat model of chronic pancreatic injury. Chronic-like pancreatitis was induced by repeated arginine pancreatitis. Palm oil tocotrienol-rich fraction (TRF) was given by gavage before and after pancreatitis inductions. Amylase and hydroxyproline were determined in pancreatic homogenates; collagen, fibronectin, α-smooth muscle actin (SMA), glial fibrillary acidic protein (GFAP), and phosphorylated Smad3 were assessed by Western blotting. Transforming growth factor (TGF)-β1 was measured in plasma. Morphological assessment included light microscopy, fibrosis area fraction, and collagen network fractal analysis. Arginine pancreatitis induced pancreatic atrophy and increased hydroxyproline that ameliorated after TRF. Arginine increased TGF-β1 (185 ± 40 vs. 15 ± 2 ng/ml; P <0.01) that was blunted by TRF (53 ± 19; P < 0.01). TRF reduced protease and Smad3 activation, collagen, and fibronectin. α-SMA increased and GFAP diminished in arginine pancreatitis, consistent with long-term stellate cell activation, and TRF reverted these changes to basal. Arginine pancreatitis increased fibrosis area fraction (4.5 ± 0.3% vs. 0.2 ± 0.2%), collagen network complexity (fractal dimension 1.52 ± 0.03 vs. 1.42 ± 0.01; P < 0.001), and inhomogeneity (lacunarity 0.63 ± 0.03 vs. 0.40 ± 0.02; P < 0.001), which were all reduced by TRF (1.3 ± 0.4%, 1.43 ± 0.02%, and 0.51 ± 0.03%, respectively; P < 0.01). Best correlation coefficients were obtained when comparing fibrosis area fraction with lacunarity (r = 0.88) and both parameters with pancreatic weight (r = -0.91 and -0.79, respectively). TRF administered only before pancreatitis best, but not fully, recapitulated the beneficial effects of TRF. Tocotrienols improve quantitative measures of chronic pancreatic damage. They may be of benefit in human chronic pancreatitis.

γ-Tocotrienol inhibits cell viability through suppression of β-catenin/Tcf signaling in human colon carcinoma HT-29 cells

γ-Tocotrienol, a major component of the tocotrienol-rich fraction of palm oil, has been suggested to have antioxidant and anticancer activity as well as potent chemopreventive effects on tumor cells. 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. We found that γ-tocotrienol could strongly suppress the transcriptional activity of β-catenin/Tcf signaling pathway in HT-29 cells. γ-Tocotrienol inhibited the expression level of total β-catenin protein but did not significantly affect the phosphorylated β-catenin level. Meanwhile, γ-tocotrienol down-regulated the protein level of nuclear β-catenin and induced its redistribution to cell membrane. Furthermore, γ-tocotrienol suppressed the expression of downstream target genes such as c-myc, cyclin D1 and survivin. 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.

Palm oil tocotrienol rich fraction reduces extracellular matrix production by inhibiting transforming growth factor-β1 in human intestinal fibroblasts

Extracellular matrix deposition is key event for the development of bowel stenosis in Crohn's disease patients. Transforming growth factor-β plays a key role in this process. We aimed at characterizing the effects of tocotrienol rich fraction on ECM proteins production and molecules that regulate the synthesis and degradation of extracellular matrix, matrix metalloproteinase-3 and tissue inhibitor of metalloproteinases-1, in human intestinal fibroblasts, and at elucidating whether the effects of tocotrienol rich fraction (TRF) are mediated through inhibition of TGF-β1. HIF were isolated from colonic or ileal tissue from Crohn's disease patients and control subjects, and were treated with TRF from palm oil either alone or in combination with TGF-β1. Procollagen 1, procollagen 3, TIMP-1 and MMP-3 production, and Smad3 phosphorylation were analyzed by Western-blotting. TRF significantly diminished procollagen 1 and 3 synthesis in HIF. Treatment of HIF with TRF increased MMP-3 production but did not modify TIMP-1. TGF-β1 induced Smad3 phosphorylation and enhanced procollagen 1 and 3 and TIMP-1 production. Pre-treatment of HIF with TRF prevented Smad3 phosphorylation and minimized the increase in collagen 1 and 3 production caused by TGF-β1. TRF has anti-fibrogenic effects on HIF, decreasing ECM production and increasing its degradation. This effect is mediated, at least in part, by inhibition of TGF-β1.

Synthesis of Fluorescent Analogues of the Anticancer Natural Products 4-Hydroxyphenylmethylene Hydantoin and δ-Tocotrienol

4-Hydroxyphenylmethylene hydantoin (PMH, 1), isolated from the Red Sea sponge Hemimycale arabica, and delta-tocotrienol (3), isolated from the tocotrienol-rich fraction of palm oil, are important antimetastatic and antiproliferative natural products that proved effective against metastatic prostate and breast cancers, respectively. New fluorescent derivatives of PMH (2) and delta-tocotrienol (4) were synthesized by Steglich esterification. Both 2 and 4 retained good anti-migratory and antiproliferative activities, respectively. Fluorescent analogues 2 and 4 can be used for the identification of molecular targets of 1 and 3 in tumor cell cultures.

Abstract 4442: Novel mechanism of gamma-tocotrienol-induced apoptosis in breast cancer cells: endoplasmic reticulum stress-dependent increase in death receptor signaling pathways

Abstract Tocotrienols are naturally occurring vitamin E compounds found in the seed endosperms of monocots and enriched in palm oil. Tocotrienols have gained substantial attention for their distinct functions in health and disease including their neuroprotective, cholesterol lowering and anti-cancer properties. Our study focused on investigating the mechanisms involved in anti-cancer actions by tocotrienols using the BALB/c 66cl-4-GFP syngeneic mouse mammary tumor model and human MCF-7 and MDA-MB-231 breast cancer cells in cell culture. In the 66cl-4-GFP implanted mammary tumor model, dietary delivery of gamma-tocotrienol (250 mg/kg diet) inhibited tumor growth (41% inhibition compared to the control diet). Immunohistochemical analyses of tumor sections from gamma-tocotrienol treated mice showed decreased proliferation (Ki-67) and increased apoptosis (TUNEL). Gamma-tocotrienol inhibited colony formation by 66cl-4 as well as human MCF-7 and MDA-MB-231 breast cancer cells. Anti-proliferative effects of gamma-tocotrienol were determined to be derived primarily from treatment-induced apoptosis. Mechanistic studies using human breast cancer cells, showed gamma-tocotrienol to cleave caspases 8, 9, and 3, suggesting the involvement of death receptor signaling. Western blot and RT-PCR analyses showed that death receptor 5 (DR5) was upregulated at both the protein level as well as mRNA level after gamma-tocotrienol treatment. Silencing of DR5 using small interfering RNA (siRNA) attenuated gamma-tocotrienol induced apoptosis. Studies investigated C/EBP homologous protein (CHOP), a transcription factor that serves as an indicator for endoplasmic reticulum (ER) stress. In our study, CHOP expression was highly induced by gamma-tocotrienol treatment, and shown to directly bind to the promoter region of DR5 gene by chromatin immunoprecipitation analyses. Silencing CHOP using siRNA partially blocked gamma-tocotrienol induced apoptosis and DR5 upregulation. Functional knock-down of JNK and p38MAPK using siRNA suppressed the upregulation of CHOP as well as DR5, suggesting upregulation of CHOP-mediated DR5 expression by gamma-tocotrienol treatment is mediated by JNK and p38 MAPK. Additional analyses showed that not only gamma-tocotrienol but also δ-tocotrienol and the tocotrienol-rich fraction of palm oil increased the level of CHOP, Grp78, ATF-4, and Xbp-1 splicing, all of which are recognized biomarkers of ER stress. In conclusion, tocotrienols trigger apoptosis by inducing ER stress which leads to JNK and p38 MAPK-dependent CHOP and DR5 upregulation. Research was supported by The Clayton Foundation for Research, NIH Grant CA59739 and NIEHS Center Grant ES007784. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4442.