Publications

Gamma-Tocotrienol controls proliferation, modulates expression of cell cycle regulatory proteins and up-regulates quinone reductase NQO2 in MCF-7 breast cancer cells

Hsieh TC, Elangovan S, Wu JM.

Anticancer Res. 2010 Jul;30(7):2869-74

Background: Tocotrienols, a subgroup of the vitamin E family, have demonstrated antioxidant and anticancer properties. Differential growth responses among different types of tocotrienols have been observed in breast cancer cells; however, specific bioactivity of each individual tocotrienol remains to be elucidated.

Materials And Methods: In this study, the effects of gamma-tocotrienol were examined with regard to its ability to suppress cell proliferation via modulation of cell cycle regulatory protein expression, and also from the perspective of control of cellular oxidoreductive status through regulation of detoxification enzymes, e.g., quinone reductase NQO2, using estrogen receptor-positive MCF-7 human breast cancer cells.

Results: It was shown that treatment by gamma-tocotrienol suppressed MCF-7 cell proliferation in a dose- and time-dependent manner. Growth suppression by gamma-tocotrienol was accompanied by changes in the levels of cell cycle regulatory proteins, notably, Rb/E2F complex, cyclin D1/cdk4 and cyclin B1/cdk1, as exemplified by loss of cyclin D1, inhibition of specific Rb phosphorylation (pRb-p at Thr821), and by the time- and dose-dependent increase in the expression of NQO2.

Conclusion: By exerting control on expression of specific cell cycle regulatory proteins in concomitance with suppression of cell proliferation, as well as the induction of NQO2, gamma-tocotrienol offers promise as an added chemopreventive and/or chemotherapeutic agent against breast cancer carcinogenesis.

Tocochromanol functions in plants: Antioxidation and beyond

Falk J, Munné-Bosch S.

J Exp Bot. 2010 Jun;61(6):1549-66.

Tocopherols and tocotrienols, collectively known as tocochromanols, are lipid-soluble molecules that belong to the group of vitamin E compounds and are essential in the human diet. Not surprisingly, most of what is known about the biological functions of tocochromanols comes from studies of mammalian systems, yet they have been shown to be synthesized only by photosynthetic organisms. The last decade has seen a radical change in the appreciation of the biological role of tocochromanols in plants thanks to a detailed characterization of mutant and transgenic plants, including several Arabidopsis thaliana mutants, the sucrose export defective1 (sxd1) maize mutant, and some transgenic potato and tobacco lines altered in tocochromanol biosynthesis. Recent findings indicate that tocopherols may play important roles in plants beyond their antioxidant function in photosynthetic membranes. Plants deficient in tocopherols show alterations in germination and export of photoassimilates, and growth, leaf senescence, and plant responses to abiotic stresses, thus suggesting that tocopherols may influence a number of physiological processes in plants. Thus, in this review not only the antioxidant function of tocochromanols in plants, but also these new emerging possible roles will be considered. Particular attention will be paid to specific roles attributed to different tocopherol homologues (particularly alpha- and gamma-tocopherol) and the possible functions of tocotrienols, which in contrast to tocopherols are only present in a range of unrelated plant groups and are almost exclusively found in seeds and fruits.

Alpha-Tocopherol attenuates the cytotoxic effect of delta-tocotrienol in human colorectal adenocarcinoma cells

Shibata A, Nakagawa K, Sookwong P, Tsuduki T, Asai A, Miyazawa T.

Biochem Biophys Res Commun. 2010 Jun 25;397(2):214-9. Epub 2010 May 20.

Recent studies have demonstrated that tocotrienol (T3) is superior to tocopherol (Toc) for cancer chemoprevention. However, there is little information on whether Toc influences the anticancer property of T3. In this study, we investigated the influence of Toc on the cytotoxic effects of delta-T3 in DLD-1 human colorectal adenocarcinoma cells. Toc, especially alpha-Toc, attenuated delta-T3-induced cytotoxicity and apoptosis in DLD-1 cells, whereas Toc alone did not exhibit any cytotoxic effect. delta-T3-induced cell cycle arrest and proapoptotic gene/protein expression (e.g., p21, p27, and caspases) were abrogated by alpha-Toc. Furthermore, coadministration of alpha-Toc decreased delta-T3 uptake into DLD-1 cells in a dose-dependent manner. These results indicate that alpha-Toc is not only less cytotoxic to cancer cells, but it also reduces the cytotoxicity of delta-T3 by inhibiting its cellular uptake.

The value of tocotrienols in the prevention and treatment of cancer

Sylvester PW, Kaddoumi A, Nazzal S, El Sayed KA.

J Am Coll Nutr. 2010 Jun;29(3 Suppl):324S-333S.

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 oftocotrienols, investigators have developed novel prodrug derivatives and nanoparticle delivery systems that greatly enhance tocotrienolbioavailability 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 oftocotrienols in the prevention and treatment of cancer.

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Mixed tocotrienols inhibit prostate carcinogenesis in TRAMP mice

Barve A, Khor TO, Reuhl K, Reddy B, Newmark H, Kong AN.

Nutr Cancer. 2010;62(6):789-94. June

The biological activities of tocotrienols are receiving increasing attention. Herein, we report the efficacy of a mixed-tocotrienol diet against prostate tumorigenesis in the transgenic adenocarcinoma mouse prostate (TRAMP) mouse model. Male TRAMP mice, 8 wk old, were fed 0.1%, 0.3%, or 1% mixed tocotrienols in AIN- 76A diet up to 24 wk old. Likewise, a positive control group consisting of male TRAMP mice and a negative control group consisting of wild-type nontransgenic mice were fed regular AIN-76A diet up to 24 wk old. Our results show that mixed-tocotrienol-fed groups had a lower incidence of tumor formation along with a significant reduction in the average wet weight of genitourinary apparatus. Furthermore, mixed tocotrienols significantly reduced the levels of high-grade neoplastic lesions as compared to the positive controls. This decrease in levels of high-grade neoplastic lesions was found to be associated with increased expression of proapoptotic proteins BAD (Bcl(2) antagonist of cell death) and cleaved caspase-3 and cell cycle regulatory proteins cyclin dependent kinase inhibitors p21 and p27. In contrast, the expression of cyclins A and E were found to be decreased in mixed-tocotrienol groups. Taken together, our results show that by modulating cell cycle regulatory proteins and increasing expression of proapoptotic proteins, mixed tocotrienols suppress prostate tumorigenesis in the TRAMP mice.

Gamma-tocotrienol protects hematopoietic stem and progenitor cells in mice after total-body irradiation

Shilpa Kulkarni, Sanchita P. Ghosh, Merriline Satyamitra, Steven Mog, Kevin Hieber, Lyudmila Romanyukha, Kristen Gambles, Raymond Toles, Tzu-Cheg Kao, Martin Hauer-Jensenb and K. Sree Kumara

Radiat Res. 2010 Jun;173(6):738-47.

We analyzed the radioprotective effects of gamma-tocotrienol (GT3) on hematopoietic stem cells (HSCs) and progenitor cells (HPCs) in sublethally irradiated mice. Flow cytometry analysis indicated that radiation depleted HPCs (c-Kit+, Lin2) to 40% at days 2 and 4 after total-body irradiation (TBI) in all treatment groups. The HPC numbers in GT3-treated mice recovered almost completely (90%) at day 7 but remained depleted in vehicle-treated mice (30%) even at day 13 after TBI. An in vitro colony-forming assay on sorted HSCs (Lin2, Sca1+, c-Kit+) indicated that TBI reduced the number of colonies to 40% and 50% at day 17 and 60, respectively, in vehicle-treated groups compared to unirradiated controls (naive). GT3-treated irradiated mice maintained higher numbers of colonies (86% and 80% compared to naive mice), thereby preserving the selfrenewable capacity of HSCs. Histopathology of sternal bone marrow indicated more regenerative microfoci for myeloid cells and megakaryocytes and higher overall cellularity in GT3- treated mice compared to vehicle controls at days 7 and 13 after TBI. GT3 treatment also reduced the frequency of micronucleated erythrocytes significantly in irradiated mice. Our results demonstrate that GT3 protected hematopoietic tissue by preserving the HSCs and HPCs and by preventing persistent DNA damage.

Palm oil–derived natural vitamin E α-tocotrienol in brain health and disease

Chandan K. Sen, PhD, Cameron Rink, PhD, and Savita Khanna, PhD

J Am Coll Nutr. 2010 June ; 29(3 Suppl): 314S–323S.

A growing body of research supports that members of the vitamin E family are not redundant with respect to their biological function. Palm oil derived from Elaeis guineensis represents the richest source of the lesser characterized vitamin E, α-tocotrienol. One of 8 naturally occurring and chemically distinct vitamin E analogs, α-tocotrienol possesses unique biological activity that is independent of its potent antioxidant capacity. Current developments in α-tocotrienol research demonstrate neuroprotective properties for the lipid-soluble vitamin in brain tissue rich in polyunsaturated fatty acids (PUFAs). Arachidonic acid (AA), one of the most abundant PUFAs of the central nervous system, is highly susceptible to oxidative metabolism under pathologic conditions. Cleaved from the membrane phospholipid bilayer by cytosolic phospholipase A2, AA is metabolized by both enzymatic and nonenzymatic pathways. A number of neurodegenerative conditions in the human brain are associated with disturbed PUFA metabolism of AA, including acute ischemic stroke. Palm oil–derived α-tocotrienol at nanomolar concentrations has been shown to attenuate both enzymatic and nonenzymatic mediators of AA metabolism and neurodegeneration. On a concentration basis, this represents the most potent of all biological functions exhibited by any natural vitamin E molecule. Despite such therapeutic potential, the scientific literature on tocotrienols accounts for roughly 1% of the total literature on vitamin E, thus warranting further investment and investigation

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Role of NF-kappaB in the anti-inflammatory effects of tocotrienols

Kaileh M, Sen R.

J Am Coll Nutr. 2010 Jun;29(3 Suppl):334S-339S.

The NF-kappaB family of transcription factors regulates genes that are critical for inflammation and immunity. In most cells, NF-kappaB function is induced upon activation of cells by various stimuli. However, constitutive NF-kappaB activity is an equally important aspect of NF-kappaB function that is particularly relevant to chronic inflammation and cancer. Here, we provide a brief overview of NF-kappaB biology and discuss the role of NF-kappaB in mediating the anti-inflammatory effects of tocotrienols The NF-kappaB family of transcription factors is a central player in the regulation of inflammation and immune responses. Consequently, NF-kappaB dysregulation has been implicated in diverse human pathologies ranging from autoimmune diseases to cancers. Additionally, there is considerable interest in the contribution of NF-kappaB-mediated chronic inflammation in aging. Because NF-kappaB-dependent gene regulation is important in virtually all mammalian cell types, it is critical to keep in mind some basic features of its functions when considering interventional therapeutics.

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Suppression of gamma-tocotrienol on UVB induced inflammation in HaCaT keratinocytes and HR-1 hairless mice via inflammatory mediators multiple signaling

Shibata A, Nakagawa K, Kawakami Y, Tsuzuki T, Miyazawa T.

J Agric Food Chem. 2010 Jun 9;58(11):7013-20.

Tocopherol (Toc) such as alpha-Toc has been expected to act as photochemopreventive agent of skin, but the effect of the other vitamin E forms [tocotrienols (T3)] has not been fully understood. We evaluated the anti-inflammatory effect of T3 on UVB-induced inflammatory reaction using immortalized human keratinocytes and hairless mice. gamma-T3 suppressed UVB-induced PGE(2) production while similar alpha-Toc doses had no effect. The anti-inflammatory actions of gamma-T3 were explained by its ability to reduce UVB-induced inflammatory gene and protein expression [cyclooxgenase-2 (COX-2), interleukin (IL)-1beta, IL-6, and monocyte chemotactic protein-1]. Western blot analysis revealed gamma-T3 inhibited p38, extracellular signal-regulated kinase, and c-Jun N-terminal kinase/stress-activated protein kinase activation. In HR-1 hairless mice, oral T3 suppressed UVB-induced changes in skin thickness, COX-2 protein expression, and hyperplasia, but alpha-Toc did not. These results suggest T3 has potential use to protect against UVB-induced skin inflammation.

Synergistic anticancer effects of combined gamma-tocotrienol and celecoxib treatment are associated with suppression in Akt and NFkappaB signaling

Shirode AB, Sylvester PW.

Biomed Pharmacother. 2010 May;64(5):327-32. Epub 2009 Nov 14.

The selective cyclooxygenase (COX)-2 inhibitor, celecoxib, and the vitamin E isoform, gamma-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 gamma-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 gamma-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, NFkappaB and prostaglandin E(2) (PGE(2)) synthesis were assessed following a 3- or 4-day culture period. Treatment with 3-4 microM gamma-tocotrienol or 7.5-10 microM celecoxib alone significantly inhibited +SA cell growth in a dose-responsive manner. However, combined treatment with subeffective doses of gamma-tocotrienol (0.25 microM) andcelecoxib (2.5 microM) resulted in a synergistic antiproliferative effect, as determined by isobologram analysis, and this growth inhibitory effect wasassociated with a reduction in PGE(2) synthesis, and decrease in COX-2, phospho-Akt (active), and phospho-NFkappaB (active) levels. These results demonstrate that the synergistic anticancer effects of combined celecoxib and gamma-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.

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