Radiation Protection - TOCOTRIENOL Tocotrienol.org

Human Studies
Article	Study objective/ Findings
Clinical evaluation of photoprotective effect by a topical antioxidants combination (tocopherols and tocotrienols) Pedreli, V.F., et.al (2011). J Eur Acad Dermatol Venereol.	In view of experimental evidence for the photoprotective properties of these antioxidants, we evaluated in 30 patients with photosensitivity, the prophylactic efficacy of a new topical agent, containing tocopherols 10% and tocotrienols 0.3%, compared with retinol, simple vehicle and untreated areas. The use of a new topical formulation containing significant concentrations of tocotrienols and tocopherols represents a promising strategy to reduce the photo-induced skin damage.

Human Studies

Article

Study objective/ Findings

Clinical evaluation of photoprotective effect by a topical antioxidants combination (tocopherols and tocotrienols)

Pedreli, V.F., et.al (2011). J Eur Acad Dermatol Venereol.

In view of experimental evidence for the photoprotective properties of these antioxidants, we evaluated in 30 patients with photosensitivity, the prophylactic efficacy of a new topical agent, containing tocopherols 10% and tocotrienols 0.3%, compared with retinol, simple vehicle and untreated areas. The use of a new topical formulation containing significant concentrations of tocotrienols and tocopherols represents a promising strategy to reduce the photo-induced skin damage.

In-vivo / Animal Studies
Article	Study objectives/ Findings
Gamma-Tocotrienol Protects the Intestine from Radiation Potentially by Accelerating Mesenchymal Immune Cell Recovery Garg, S., et al., (2019) Antioxidants	Total body irradiation can affect various organs especially the intestine, which is highly sensitive to ionizing radiation. In this study, mice that were treated with gamma-tocotrienol 24 hours before exposed to total body irradiation showed a reduction in intestinal cell-death. In conclusion, the study found that gamma-tocotrienol suppresses radiation-induced structural and functional damage in the intestine, potentially by upregulating occludin level and by facilitating the recovery of mesenchymal immune cells after total body irradiation.
Enhanced Survival in Mice Exposed to Ionizing Radiation by Combination of Gamma-Tocotrienol and Simvastatin. Pathak, R., et al (2019). Military Medicine	This study aimed to investigate the synergistic action of Vitamin E (gamma-tocotrienol) in combination with a cholesterol-lowering drug (Statins) in protection against radiation. It is observed that the combination of both gamma-tocotrienol and Statins provide better radiation protection compared to each drug alone. Optimization fo doses of Statins and gamma-tocotrienol is needed to achieve significantly higher protection.
Nanoencapsulation of coenzyme Q10 and vitamin E acetate protects against UVB radiation-induced skin injury in mice. Pegoraro, N.S., et.al (2017). Colloids Surf B Biointerfaces.	This study aimed to investigate the feasibility of producing semisolid formulations based on nanocapsule suspensions containing the association of the coenzyme Q10 and vitamin E acetate by adding gellan gum (2%) to the suspensions. Furthermore, we studied their application as an alternative for the treatment of inflammation induced by ultraviolet B (UVB) radiation. For this, an animal model of injury induced by UVB-radiation was employed. All semisolids presented pH close to 5.5, drug content above 95% and mean diameter on the nanometric range, after redispersion in water. Besides, the semisolids presented non-Newtonian flow with pseudoplastic behavior and suitable spreadability factor values. The results also showed that the semisolid containing coenzyme Q10-loaded nanocapsules with higher vitamin E acetate concentration reduced in 73±8% the UVB radiation-induced ear edema. Moreover, all formulations tested were able to reduce inflammation parameters evaluated through MPO activity and histological procedure on injured tissue and the semisolids containing the nanoencapsulated coenzyme Q10 reduced oxidative parameters assessment through the non-protein thiols levels and lipid peroxidation. This way, the semisolids based on nanocapsules may be considered a promising approach for the treatment and prevention of skin inflammation diseases.
Development of Orally Administered γ-Tocotrienol (GT3) Nanoemulsion for Radioprotection. Ledet, G.A., et.al (2016). Int J Mol Sci.	The purpose of this study was two-fold: (1) to formulate γ-tocotrienol (GT3) in a nanoemulsion formulation as a prophylactic orally administered radioprotective agent; and (2) to optimize the storage conditions to preserve the structural integrity of both the formulation and the compound. γ-tocotrienol was incorporated into a nanoemulsion and lyophilized with lactose. Ultra performance liquid chromatography-mass spectroscopy (UPLC-MS) was used to monitor the chemical stability of GT3 over time, the particle size and ζ potential, and scanning electron microscopy (SEM) were used to study the physical stability of the nanoemulsion. Radioprotective and toxicity studies were performed in mice. The liquid formulation exhibited GT3 degradation at all storage temperatures. Lyophilization, in the presence of lactose, significantly reduced GT3 degradation. Both the liquid and lyophilized nanoemulsions had stable particle size and ζ potential when stored at 4 °C. Toxicity studies of the nanoemulsion resulted in no observable toxicity in mice at an oral dose of 600 mg/kg GT3. The nano-formulated GT3 (300 mg/kg) demonstrated enhanced survival efficacy compared to GT3 alone (200 and 400 mg/kg) in CD2F1 mice exposed to total body gamma radiation. The optimal long-term storage of formulated GT3 is as a powder at -20 °C to preserve drug and formulation integrity. Formulation of GT3 as a nanoemulsion for oral delivery as a prophylactic radioprotectant shows promise and warrants further investigation.
The potentiation of the radioprotective efficacy of two medical countermeasures, gamma-tocotrienol and amifostine, by a combination prophylactic modality. Singh V.K. et.al (2016). Radiat Prot Dosimetry.	This study was designed to evaluate the possible potentiation of survival protection afforded by relatively low-dose amifostine prophylaxis against total body irradiation in combination with a protective, less toxic agent, gamma-tocotrienol (GT3). Mice were administered amifostine and/or GT3, then exposed to 9.2 Gy ⁶⁰Co γ-irradiation and monitored for survival for 30 days. To investigate cytokine stimulation, mice were administered amifostine or GT3; serum samples were collected and analyzed for cytokines. Survival studies show single treatments of GT3 or amifostine significantly improved survival, compared to the vehicle, and combination treatments resulted in significantly higher survival compared to single treatments. In vivo studies with GT3 confirmed prior work indicating GT3 induces granulocyte colony-stimulating factor (G-CSF). This approach, the prophylactic combination of amifostine and GT3, which act through different mechanisms, shows promise and should be investigated further as a potential countermeasure for acute radiation syndrome.
Progenitor Cell Mobilization by Gamma-tocotrienol: A Promising Radiation Countermeasure. Singh, V.K., et.al (2016). Health Phys.	This article reviews studies of progenitor mobilization with gamma-tocotrienol (GT3), a tocol under advanced development as a radiation countermeasure for acute radiation syndrome (ARS). GT3 protects mice against high doses of ionizing radiation and induces high levels of granulocyte colony-stimulating factor (G-CSF). GT3-induced G-CSF in conjunction with AMD3100 (a chemokine receptor antagonist clinically used to improve the yield of mobilized progenitors) mobilizes progenitors; these mobilized progenitors mitigate injury when infused to mice exposed to acute, high-dose ionizing radiation. The administration of a G-CSF antibody to GT3-injected donor mice abrogated the radiomitigative efficacy of blood or peripheral blood mononuclear cells (PBMC) in irradiated recipient mice. The efficacy of GT3-injected donor mice blood or PBMC was comparable to a recently published article involving blood or mononuclear cells obtained from mice injected with G-CSF. The injected progenitors were found to localize in various tissues of irradiated hosts. The authors demonstrate the efficacy of a bridging therapy in a preclinical animal model that allows the lymphohematopoietic system of severely immunocompromised mice to recover. This suggests that GT3 is a highly effective agent for radioprotection and mobilizing progenitors with significant therapeutic potential. Therefore, GT3 may be considered for further translational development and ultimately for use in humans.
Gamma-Tocotrienol Modulates Radiation-Induced MicroRNA Expression in Mouse Spleen. Ghosh, S.P., et.al (2016). Radiat Res.	In this study, it is shown that GT3 differentially modulates the miR footprint in the spleen of irradiated mice compared to controls at early times (day 1), as well as later times (day 4 and 15) after total-body irradiation. It is observed that miR expression was altered in a dose- and time-dependent manner in GT3-pretreated spleen tissues from total-body irradiated mice. GT3 appeared to affect the expression of a number of radiation-modulated miRs known to be involved in hematopoiesis and lymphogenesis. Moreover, GT3 pretreatment also suppressed the upregulation of radiation-induced p53, suggesting the function of GT3 in the prevention of radiation-induced damage to the spleen. In addition, it is shown that GT3 significantly reduced serum levels of Flt3L, a biomarker of radiation-induced bone marrow aplasia. Further in silico analyses of the effect of GT3 implied the association of p38 MAPK, ERK and insulin signaling pathways. Our study provides initial insight into the mechanism by which GT3 mediates protection of spleen after total-body irradiation.
Radioprotective Efficacy of Gamma-Tocotrienol in Nonhuman Primates. Singh, V.K., et.al (2016). Radiat Res.	Gamma-tocotrienol (GT3) has demonstrated strong radioprotective efficacy in the mouse model, indicating the need for further evaluation in a large animal model. In this study, GT3 pharmacokinetics (PK) and efficacy at different doses of cobalt-60 gamma radiation (0.6 Gy/min) using the nonhuman primate (NHP) model are evaluated. The PK results demonstrated increased area under the curve with increasing drug dose and half-life of GT3. GT3 treatment resulted in reduced group mean neutropenia by 3-5 days and thrombocytopenia by 1-5 days. At 5.8 and 6.5 Gy total-body irradiation, GT3 treatment completely prevented thrombocytopenia. The capability of GT3 to reduce severity and duration of neutropenia and thrombocytopenia was dose dependent; 75 mg/kg treatment was more effective than 37.5 mg/kg treatment after a 5.8 Gy dose. However, the higher GT3 dose (75 mg/kg) was associated with higher frequency of adverse skin effects (small abscess) at the injection site. GT3 treatment of irradiated NHPs caused no significant difference in animal survival at 60 days postirradiation, however, low mortality was observed in irradiated, vehicle-treated groups as well. The data from this pilot study further elucidate the role and pharmacokinetics of GT3 in hematopoietic recovery after irradiation in a NHP model, and demonstrate the potential of GT3 as a promising radioprotector.
Thrombomodulin contributes to gamma-tocotrienol mediated lethality protection and hematopoietic cell recovery in irradiated mice. Pathak, R., et.al (2015). PLoS One.	Systemic administration of recombinant thrombomodulin (TM) confers radiation protection partly by accelerating hematopoietic recovery. The uniquely potent radioprotector gamma tocotrienol (GT3), in addition to being a strong antioxidant, inhibits the enzyme hydroxy-methyl-glutaryl-coenzyme A reductase (HMGCR) and thereby likely modulates the expression of TM. We hypothesized that the mechanism underlying the exceptional radioprotective properties of GT3 partly depends on the presence of endothelial TM. In vitro studies confirmed that ionizing radiation suppresses endothelial TM (about 40% at 4 hr after 5 Gy γ-irradiation) and that GT3 induces TM expression (about 2 fold at the mRNA level after 5 μM GT3 treatment for 4 hr). In vivo survival studies showed that GT3 was significantly more effective as a radioprotector in TM wild type (TM^+/+) mice than in mice with low TM function (TM^Pro/-). After exposure to 9 Gy TBI, GT3 pre-treatment conferred 85% survival in TM^+/+ mice compared to only 50% in TM^Pro/-. Thus, GT3-mediated radiation lethality protection is partly dependent on endothelial TM. Significant post-TBI recovery of hematopoietic cells, particularly leukocytes, was observed in TM^+/+ mice (p = 0.003), but not in TM^Pro/- mice, despite the fact that GT3 induced higher levels of granulocyte colony stimulating factor (G-CSF) in TM^Pro/- mice (p = 0.0001). These data demonstrate a critical, G-CSF-independent, role for endothelial TM in GT3-mediated lethality protection and hematopoietic recovery after exposure to TBI and may point to new strategies to enhance the efficacy of current medical countermeasures in radiological/nuclear emergencies.
A tocotrienol-enriched formulation protects against radiation-induced changes in cardiac mitochondria without modifying late cardiac function or structure. Sridharan, V., et.al (2015) Radiat Res.	Radiation-induced heart disease (RIHD) is a common and sometimes severe late side effect of radiation therapy for intrathoracic and chest wall tumors. It has been previously shown that local heart irradiation in a rat model caused prolonged changes in mitochondrial respiration and increased susceptibility to mitochondrial permeability transition pore (mPTP) opening. Because tocotrienols are known to protect against oxidative stress-induced mitochondrial dysfunction, this study examines the effects of tocotrienols on radiation-induced alterations in mitochondria, and structural and functional manifestations of RIHD. Male Sprague-Dawley rats received image-guided localized X irradiation to the heart to a total dose of 21 Gy. Twenty-four hours before irradiation, rats received a tocotrienol-enriched formulation or vehicle by oral gavage. Mitochondrial function and mitochondrial membrane parameters were studied at 2 weeks and 28 weeks after irradiation. In addition, cardiac function and histology were examined at 28 weeks. A single oral dose of the tocotrienol-enriched formulation preserved Bax/Bcl2 ratios and prevented mPTP opening and radiation-induced alterations in succinate-driven mitochondrial respiration. Nevertheless, the late effects of local heart irradiation pertaining to myocardial function and structure were not modified. Our studies suggest that a single dose of tocotrienols protects against radiation-induced mitochondrial changes, but these effects are not sufficient against long-term alterations in cardiac function or remodeling.
Progenitors Mobilized by Gamma-Tocotrienol as an Effective Radiation Countermeasure Singh, V.K., et.al (2014). PLoS One.	The purpose of this study was to elucidate the role of gamma-tocotrienol (GT3)-mobilized progenitors in mitigating damage to mice exposed to a supralethal dose of cobalt-60 gamma-radiation. CD2F1 mice were transfused 24 h post-irradiation with whole blood or isolated peripheral blood mononuclear cells (PBMC) from donors that had received GT3 72 h prior to blood collection and recipient mice were monitored for 30 days. To understand the role of GT3-induced granulocyte colony-stimulating factor (G-CSF) in mobilizing progenitors, donor mice were administered a neutralizing antibody specific to G-CSF or its isotype before blood collection. Bacterial translocation from gut to heart, spleen and liver of irradiated recipient mice was evaluated by bacterial culture on enriched and selective agar media. Endotoxin in serum samples also was measured. It is also analyzed the colony-forming units in the spleens of irradiated mice. The results demonstrate that whole blood or PBMC from GT3-administered mice mitigated radiation injury when administered 24 h post-irradiation. Furthermore, administration of a G-CSF antibody to GT3-injected mice abrogated the efficacy of blood or PBMC obtained from such donors. Additionally, GT3-mobilized PBMC inhibited the translocation of intestinal bacteria to the heart, spleen, and liver, and increased colony forming unit-spleen (CFU-S) numbers in irradiated mice. Our data suggests that GT3 induces G-CSF, which mobilizes progenitors and these progenitors mitigate radiation injury in recipient mice. This approach using mobilized progenitor cells from GT3-injected donors could be a potential treatment for humans exposed to high doses of radiation.
Gamma tocotrienol, a potent radioprotector, preferentially upregulates expression of anti-apoptotic genes to promote intestinal cell survival Suman, S., et.al (2013). Food and Chemial Toxicology.	Gamma tocotrienol (GT3) has been reported as a potent ameliorator of radiation-induced gastrointestinal (GI) toxicity when administered prophylactically. This study aimed to evaluate the role of GT3 mediated pro- and anti-apoptotic gene regulation in protecting mice from radiation-induced GI damage. Our study demonstrated that GT3-mediated protection of intestinal cells from a GI-toxic dose of radiation occurred via upregulation of antiapoptotic and downregulation of pro-apoptotic factors, both at the transcript as well as at the protein levels.
Granulocyte colony-stimulating factor antibody abrogates radioprotective efficacy of gamma-tocotrienol, a promising radiation countermeasure. Kulkarni, S., et.al (2013). Cytokine.	This study aimed to determine the role of granulocyte colony-stimulating factor (G-CSF), induced by a promising radiation countermeasure, gamma tocotrienol (GT3), in protecting mice from lethal doses of ionizing radiation. CD2F1 mice were injected with an optimal dose of GT3 and a G-CSF antibody, and their 30-d survival was monitored. An appropriate antibody isotype was used as a control. Multiplex Luminex was used to analyze GT3-induced cytokines. G-CSF neutralization by exogenous administration of a G-CSF antibody was confirmed by analyzing serum cytokine levels. Our results demonstrate that GT3 significantly protected mice against ionizing radiation, and induced high levels of G-CSF in peripheral blood 24h after administration. Injection of a G-CSF neutralizing antibody to the GT3-treated mice resulted in complete neutralization of G-CSF and abrogation of its protective efficacy. Administration of a G-CSF antibody did not affect levels of other cytokines induced by GT3. Histopathology of bone marrow from GT3-treated and -irradiated mice demonstrated protection of the hematopoietic tissue, and also that such protection was abrogated by administering a G-CSF antibody. Our results suggest that induction of high levels of G-CSF by GT3 administration is responsible for its protective efficacy against radiation injury.
Mechanism of radioprotection by delta-tocotrienol: pharmacokinetics, pharmacodynamics and modulation of signaling pathways. Satyamitra, M., et.al (2012). Br J Radiol.	The objective of this study was to investigate the correlation between in vivo δ-tocotrienol (DT3) pharmacokinetics, pharmacodynamics and radiation protection, and to evaluate the effect of DT3 pre-treatment on radiation-induced alterations in apoptotic and autophagic pathways. These data indicate that DT3 stimulates multilineage haematopoiesis, protects against radiation-induced apoptosis downstream of the mitochondria and stimulates cytoprotective autophagy. Apart from a potent antioxidant activity, DT3 may elicit survival advantage following irradiation by enhancing haematopoiesis and modulating signalling pathways.
Hematopoietic recovery and amelioration of radiation-induced lethality by the vitamin E isoform delta-tocotrienol. Satyamitra, M.M., et.al (2011). Radiat Res.	δ-Tocotrienol (DT3), a vitamin E isoform, is associated with strong antioxidant and immunomodulatory properties. We confirmed the potent antioxidant activity in membrane systems and showed that DT3 is an effective radiation protector and mitigator. DT3 (4 μM, P < 0.001) inhibited lipid peroxidation in mouse liver microsomes and nitric oxide (NO) formation (20 μM DT3, P < 0.01) in RAW264.7 cells, a murine alveolar macrophage line. In CD2F1 mice exposed to lethal total-body radiation from a (60)Co γ-radiation source, a single subcutaneous (s.c.) injection of DT3 before or after irradiation produced a significant increase in 30-day survival. DT3 was effective from 18.75 to 300 mg/kg (–24 h, P < 0.001). A single dose of 150 or 300 mg/kg DT3 given 24 h before irradiation (radioprotection) resulted in dose reduction factors (DRFs) of 1.19 and 1.27, respectively (P < 0.001). Further, DT3 reduced radiation lethality when administered 2, 6 or 12 h after irradiation, and 150 mg/kg DT3 administered 2 h after exposure conferred a DRF of 1.1 (mitigation). The optimum schedule of 300 mg/kg DT3 24 h prior to 7 Gy significantly reduced pancytopenia compared to irradiated controls (P < 0.05). The large therapeutic potential of and multi-lineage hematopoietic recovery for DT3 warrants further studies.
Pentoxifylline Enhances the Radioprotective Properties of γ-Tocotrienol: Differential Effects on the Hematopoietic, Gastrointestinal and Vascular Systems Berbee, M., et.al (2011). Radiat Res.	The vitamin E analog γ-tocotrienol (GT3) is a potent radioprotector and mitigator. This study was performed to (a) determine whether the efficacy of GT3 can be enhanced by the addition of the phosphodiesterase inhibitor pentoxifylline (PTX) and (b) to obtain information about the mechanism of action. Mice were injected subcutaneously with vehicle, GT3 [400 mg/kg 24 h before total-body irradiation (TBI)], PTX (200 mg/kg 30 min before TBI), or GT3+PTX before being exposed to 8.5–13 Gy TBI. Overall lethality, survival time and intestinal, hematopoietic and vascular injury were assessed. Cytokine levels in the bone marrow microenvironment were measured, and the requirement for endothelial nitric oxide synthase (eNOS) was studied in eNOS-deficient mice. GT3+PTX significantly improved survival compared to GT3 alone and provided full protection against lethality even after exposure to 12.5 Gy. GT3+PTX improved bone marrow CFUs, spleen colony counts and platelet recovery compared to GT3 alone. GT3 and GT3+PTX increased bone marrow plasma G-CSF levels as well as the availability of IL-1α, IL-6 and IL-9 in the early postirradiation phase. GT3 and GT3+PTX were equally effective in ameliorating intestinal injury and vascular peroxynitrite production. Survival studies in eNOS-deficient mice and appropriate controls revealed that eNOS was not required for protection against lethality after TBI. Combined treatment with GT3 and PTX increased postirradiation survival over that with GT3 alone by a mechanism that may depend on induction of hematopoietic stimuli. GT3+PTX did not reduce GI toxicity or vascular oxidative stress compared to GT3 alone. The radioprotective effect of either drug alone or both drugs in combination does not require the presence of eNOS.
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.	Exposure to γ-radiation causes rapid hematopoietic cell apoptosis and bone marrow suppression. However, there are no approved radiation countermeasures for the acute radiation syndrome. In this study, we demonstrated that natural δ-tocotrienol, one of the isomers of vitamin E, significantly enhanced survival in total body lethally irradiated mice. We explored 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.
Gamma-tocotrienol protects hematopoietic stem and progenitor cells in mice after total-body irradiation. Kulkami, S., et.al (2010). Radiat Res.	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(+), Lin(-)) 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 (Lin(-), 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 (naïve). GT3-treated irradiated mice maintained higher numbers of colonies (86% and 80% compared to naïve mice), thereby preserving the self-renewable 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.
γ-Tocotrienol Ameliorates Intestinal Radiation Injury and Reduces Vascular Oxidative Stress after Total-Body Irradiation by an HMG-CoA Reductase-Dependent Mechanism Berbee, M., et.al (2009). Radiat Res.	Analogs of vitamin E (tocols) are under development as radioprophylactic agents because of their high efficacy and lack of toxicity. Gamma-tocotrienol (GT3) is of particular interest because, in addition to being an antioxidant, it also inhibits 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase and accumulates to greater extent in endothelial cells than other tocols. We addressed in vivo whether HMG-CoA reductase inhibition contributes to the radioprotection conferred by GT3. Groups of mice were treated with vehicle, mevalonate (the product of the reaction catalyzed by HMG-CoA reductase), GT3 alone or GT3 in combination with mevalonate. Lethality and standard parameters of injury to the hematopoietic, intestinal and vascular/endothelial systems were assessed after exposure to total-body irradiation. GT3 improved post-irradiation survival and decreased radiation-induced vascular oxidative stress, an effect that was reversible by mevalonate. GT3 also enhanced hematopoietic recovery, reduced intestinal radiation injury, and accelerated the recovery of soluble markers of endothelial function. These parameters were not reversed by mevalonate co-administration. Our data confirm GT3’s radioprophylactic properties against hematopoietic injury and, for the first time, demonstrate benefits in terms of protection against gastrointestinal and vascular injury. The radioprotective efficacy of GT3 against vascular injury is related to its properties as an HMG-CoA reductase inhibitor.
Dietary tocotrienol reduces UVB-induced skin damage and sesamin enhances tocotrienol effects in hairless mice. Yamada, Y., et.al (2008). J Nutr Sci Vitaminol.	The present studies were undertaken to determine whether dietary tocotrienols and those with sesamin could protect the skin from damage induced by UVB irradiation in hairless mice fed four diets: a vitamin E-free diet, a 50mg/kg alpha-tocopherol) diet and a 229 mg/kg T-mix with 2 g/kg sesamin diet. The results suggest that dietary tocotrienols protect the skin more strongly than alpha-tocopherol against damage induced by UVB and sesamin enhances tocotrienol effects. The results suggest that dietary tocotrienols protect the skin more strongly than alpha-tocopherol against damage induced UVB and sesamin enhances tocotrienol effects.
Efficacy of topically applied tocopherols and tocotrienols in protection of murine skin from oxidative damage induced by UV-irradiation. Weber, C., et.al (1997). Free Radic Biol Med.	To assess the efficacy of various forms of vitamin E in protection of skin from UV-light-induced oxidative stress, vitamin E (tocotrienol-rich fraction of palm oil, TRF) was applied to mouse skin and the contents of antioxidants before and after exposure to UV-light were measured. Four polypropylene plastic rings (1 cm2) were glued onto the animals’ backs, and 20 microliters 5% TRF in polyethylene glycol-400 (PEG) was applied to the skin circumscribed by two rings and 20 microliters PEG to the other two rings. After 2 h, the skin was washed and half of the sites were exposed to UV-irradiation (2.8 mW/cm2 for 29 mi: 3 MED). TRF treatment (n = 19 mice) increased mouse skin alpha-tocopherol 28 +/- 16-fold, alpha-tocotrienol 80 +/- 50-fold, gamma-tocopherol 130 +/- 108-fold, and gamma-tocotrienol 51 +/- 36-fold. A significantly higher percentage of alpha-tocopherol was present in the skin as compared with that in the applied TRF. After UV-irradiation, all vitamin E forms decreased significantly (p < .01), while a larger proportion of the vitamin E remained in PEG-treated (approximately 80%) compared with TRF-treated (approximately 40%) skin. Nonetheless, vitamin E concentrations in irradiated TRF-treated skin were significantly higher than in the nonirradiated PEG-treated (control) skin (p < .01). Thus, UV-irradiation of skin destroys its antioxidants: however, prior application of TRF to mouse skin results in preservation of vitamin E.
Efficacy of topically applied tocopherols and tocotrienols in protection of murine skin from oxidative damage induced by UV-irradiation. Weber, C., et.al (1997). Free Radic Biol Med.	To assess the efficacy of various forms of vitamin E in protection of skin from UV-light-induced oxidative stress, vitamin E (tocotrienol-rich fraction of palm oil, TRF) was applied to mouse skin and the contents of antioxidants before and after exposure to UV-light were measured. Four polypropylene plastic rings (1 cm2) were glued onto the animals’ backs, and 20 microliters 5% TRF in polyethylene glycol-400 (PEG) was applied to the skin circumscribed by two rings and 20 microliters PEG to the other two rings. After 2 h, the skin was washed and half of the sites were exposed to UV-irradiation (2.8 mW/cm2 for 29 mi: 3 MED). TRF treatment (n = 19 mice) increased mouse skin alpha-tocopherol 28 +/- 16-fold, alpha-tocotrienol 80 +/- 50-fold, gamma-tocopherol 130 +/- 108-fold, and gamma-tocotrienol 51 +/- 36-fold. A significantly higher percentage of alpha-tocopherol was present in the skin as compared with that in the applied TRF. After UV-irradiation, all vitamin E forms decreased significantly (p < .01), while a larger proportion of the vitamin E remained in PEG-treated (approximately 80%) compared with TRF-treated (approximately 40%) skin. Nonetheless, vitamin E concentrations in irradiated TRF-treated skin were significantly higher than in the nonirradiated PEG-treated (control) skin (p < .01). Thus, UV-irradiation of skin destroys its antioxidants: however, prior application of TRF to mouse skin results in preservation of vitamin E.
Ozone depletes tocopherols and tocotrienols topically applied to murine skin. Thiele, J.J., et.al (1997). FEBS Lett.	To evaluate ozone damage to hairless mouse skin, two parameters of oxidative damage, vitamin E depletion and malondialdehyde (MDA) production, were measured in vitamin E-enriched and in control skin from mice exposed to ozone (10 ppm). A 5% vitamin E solution (tocotrienol-rich fraction, TRF) in polyethylene glycol (PEG) was applied to 2 sites on the back of hairless mice, PEG to 2 sites. After 2 h, the sites were washed, one of each pair of sites covered and the mice exposed ozone for 2 h. Ozone exposure (compared with covered sites) increased epidermal MDA in PEG-treated sites, while vitamin E was unchanged. In contrast, ozone exposure significantly depleted vitamin E in TRF-treated sites, while significant MDA accumulation was prevented. This is the first demonstration that ozone exposure causes damage to cutaneous lipids, an effect which can be attenuated by vitamin E application.

In-vivo / Animal Studies

Article

Study objectives/ Findings

Gamma-Tocotrienol Protects the Intestine from Radiation Potentially by Accelerating Mesenchymal Immune Cell Recovery

Garg, S., et al., (2019) Antioxidants

Total body irradiation can affect various organs especially the intestine, which is highly sensitive to ionizing radiation. In this study, mice that were treated with gamma-tocotrienol 24 hours before exposed to total body irradiation showed a reduction in intestinal cell-death. In conclusion, the study found that gamma-tocotrienol suppresses radiation-induced structural and functional damage in the intestine, potentially by upregulating occludin level and by facilitating the recovery of mesenchymal immune cells after total body irradiation.

Enhanced Survival in Mice Exposed to Ionizing Radiation by Combination of Gamma-Tocotrienol and Simvastatin.

Pathak, R., et al (2019). Military Medicine

This study aimed to investigate the synergistic action of Vitamin E (gamma-tocotrienol) in combination with a cholesterol-lowering drug (Statins) in protection against radiation. It is observed that the combination of both gamma-tocotrienol and Statins provide better radiation protection compared to each drug alone. Optimization fo doses of Statins and gamma-tocotrienol is needed to achieve significantly higher protection.

Nanoencapsulation of coenzyme Q10 and vitamin E acetate protects against UVB radiation-induced skin injury in mice.

Pegoraro, N.S., et.al (2017). Colloids Surf B Biointerfaces.

This study aimed to investigate the feasibility of producing semisolid formulations based on nanocapsule suspensions containing the association of the coenzyme Q10 and vitamin E acetate by adding gellan gum (2%) to the suspensions. Furthermore, we studied their application as an alternative for the treatment of inflammation induced by ultraviolet B (UVB) radiation. For this, an animal model of injury induced by UVB-radiation was employed. All semisolids presented pH close to 5.5, drug content above 95% and mean diameter on the nanometric range, after redispersion in water. Besides, the semisolids presented non-Newtonian flow with pseudoplastic behavior and suitable spreadability factor values. The results also showed that the semisolid containing coenzyme Q10-loaded nanocapsules with higher vitamin E acetate concentration reduced in 73±8% the UVB radiation-induced ear edema. Moreover, all formulations tested were able to reduce inflammation parameters evaluated through MPO activity and histological procedure on injured tissue and the semisolids containing the nanoencapsulated coenzyme Q10 reduced oxidative parameters assessment through the non-protein thiols levels and lipid peroxidation. This way, the semisolids based on nanocapsules may be considered a promising approach for the treatment and prevention of skin inflammation diseases.

Development of Orally Administered γ-Tocotrienol (GT3) Nanoemulsion for Radioprotection.

Ledet, G.A., et.al (2016). Int J Mol Sci.

The purpose of this study was two-fold: (1) to formulate γ-tocotrienol (GT3) in a nanoemulsion formulation as a prophylactic orally administered radioprotective agent; and (2) to optimize the storage conditions to preserve the structural integrity of both the formulation and the compound. γ-tocotrienol was incorporated into a nanoemulsion and lyophilized with lactose. Ultra performance liquid chromatography-mass spectroscopy (UPLC-MS) was used to monitor the chemical stability of GT3 over time, the particle size and ζ potential, and scanning electron microscopy (SEM) were used to study the physical stability of the nanoemulsion. Radioprotective and toxicity studies were performed in mice. The liquid formulation exhibited GT3 degradation at all storage temperatures. Lyophilization, in the presence of lactose, significantly reduced GT3 degradation. Both the liquid and lyophilized nanoemulsions had stable particle size and ζ potential when stored at 4 °C. Toxicity studies of the nanoemulsion resulted in no observable toxicity in mice at an oral dose of 600 mg/kg GT3. The nano-formulated GT3 (300 mg/kg) demonstrated enhanced survival efficacy compared to GT3 alone (200 and 400 mg/kg) in CD2F1 mice exposed to total body gamma radiation. The optimal long-term storage of formulated GT3 is as a powder at -20 °C to preserve drug and formulation integrity. Formulation of GT3 as a nanoemulsion for oral delivery as a prophylactic radioprotectant shows promise and warrants further investigation.

The potentiation of the radioprotective efficacy of two medical countermeasures, gamma-tocotrienol and amifostine, by a combination prophylactic modality.

Singh V.K. et.al (2016). Radiat Prot Dosimetry.

This study was designed to evaluate the possible potentiation of survival protection afforded by relatively low-dose amifostine prophylaxis against total body irradiation in combination with a protective, less toxic agent, gamma-tocotrienol (GT3). Mice were administered amifostine and/or GT3, then exposed to 9.2 Gy ⁶⁰Co γ-irradiation and monitored for survival for 30 days. To investigate cytokine stimulation, mice were administered amifostine or GT3; serum samples were collected and analyzed for cytokines. Survival studies show single treatments of GT3 or amifostine significantly improved survival, compared to the vehicle, and combination treatments resulted in significantly higher survival compared to single treatments. In vivo studies with GT3 confirmed prior work indicating GT3 induces granulocyte colony-stimulating factor (G-CSF). This approach, the prophylactic combination of amifostine and GT3, which act through different mechanisms, shows promise and should be investigated further as a potential countermeasure for acute radiation syndrome.

Progenitor Cell Mobilization by Gamma-tocotrienol: A Promising Radiation Countermeasure.

Singh, V.K., et.al (2016). Health Phys.

This article reviews studies of progenitor mobilization with gamma-tocotrienol (GT3), a tocol under advanced development as a radiation countermeasure for acute radiation syndrome (ARS). GT3 protects mice against high doses of ionizing radiation and induces high levels of granulocyte colony-stimulating factor (G-CSF). GT3-induced G-CSF in conjunction with AMD3100 (a chemokine receptor antagonist clinically used to improve the yield of mobilized progenitors) mobilizes progenitors; these mobilized progenitors mitigate injury when infused to mice exposed to acute, high-dose ionizing radiation. The administration of a G-CSF antibody to GT3-injected donor mice abrogated the radiomitigative efficacy of blood or peripheral blood mononuclear cells (PBMC) in irradiated recipient mice. The efficacy of GT3-injected donor mice blood or PBMC was comparable to a recently published article involving blood or mononuclear cells obtained from mice injected with G-CSF. The injected progenitors were found to localize in various tissues of irradiated hosts. The authors demonstrate the efficacy of a bridging therapy in a preclinical animal model that allows the lymphohematopoietic system of severely immunocompromised mice to recover. This suggests that GT3 is a highly effective agent for radioprotection and mobilizing progenitors with significant therapeutic potential. Therefore, GT3 may be considered for further translational development and ultimately for use in humans.

Gamma-Tocotrienol Modulates Radiation-Induced MicroRNA Expression in Mouse Spleen.

Ghosh, S.P., et.al (2016). Radiat Res.

In this study, it is shown that GT3 differentially modulates the miR footprint in the spleen of irradiated mice compared to controls at early times (day 1), as well as later times (day 4 and 15) after total-body irradiation. It is observed that miR expression was altered in a dose- and time-dependent manner in GT3-pretreated spleen tissues from total-body irradiated mice. GT3 appeared to affect the expression of a number of radiation-modulated miRs known to be involved in hematopoiesis and lymphogenesis. Moreover, GT3 pretreatment also suppressed the upregulation of radiation-induced p53, suggesting the function of GT3 in the prevention of radiation-induced damage to the spleen. In addition, it is shown that GT3 significantly reduced serum levels of Flt3L, a biomarker of radiation-induced bone marrow aplasia. Further in silico analyses of the effect of GT3 implied the association of p38 MAPK, ERK and insulin signaling pathways. Our study provides initial insight into the mechanism by which GT3 mediates protection of spleen after total-body irradiation.

Radioprotective Efficacy of Gamma-Tocotrienol in Nonhuman Primates.

Singh, V.K., et.al (2016). Radiat Res.

Gamma-tocotrienol (GT3) has demonstrated strong radioprotective efficacy in the mouse model, indicating the need for further evaluation in a large animal model. In this study, GT3 pharmacokinetics (PK) and efficacy at different doses of cobalt-60 gamma radiation (0.6 Gy/min) using the nonhuman primate (NHP) model are evaluated. The PK results demonstrated increased area under the curve with increasing drug dose and half-life of GT3. GT3 treatment resulted in reduced group mean neutropenia by 3-5 days and thrombocytopenia by 1-5 days. At 5.8 and 6.5 Gy total-body irradiation, GT3 treatment completely prevented thrombocytopenia. The capability of GT3 to reduce severity and duration of neutropenia and thrombocytopenia was dose dependent; 75 mg/kg treatment was more effective than 37.5 mg/kg treatment after a 5.8 Gy dose. However, the higher GT3 dose (75 mg/kg) was associated with higher frequency of adverse skin effects (small abscess) at the injection site. GT3 treatment of irradiated NHPs caused no significant difference in animal survival at 60 days postirradiation, however, low mortality was observed in irradiated, vehicle-treated groups as well. The data from this pilot study further elucidate the role and pharmacokinetics of GT3 in hematopoietic recovery after irradiation in a NHP model, and demonstrate the potential of GT3 as a promising radioprotector.

Thrombomodulin contributes to gamma-tocotrienol mediated lethality protection and hematopoietic cell recovery in irradiated mice.

Pathak, R., et.al (2015). PLoS One.

Systemic administration of recombinant thrombomodulin (TM) confers radiation protection partly by accelerating hematopoietic recovery. The uniquely potent radioprotector gamma tocotrienol (GT3), in addition to being a strong antioxidant, inhibits the enzyme hydroxy-methyl-glutaryl-coenzyme A reductase (HMGCR) and thereby likely modulates the expression of TM. We hypothesized that the mechanism underlying the exceptional radioprotective properties of GT3 partly depends on the presence of endothelial TM.

In vitro studies confirmed that ionizing radiation suppresses endothelial TM (about 40% at 4 hr after 5 Gy γ-irradiation) and that GT3 induces TM expression (about 2 fold at the mRNA level after 5 μM GT3 treatment for 4 hr). In vivo survival studies showed that GT3 was significantly more effective as a radioprotector in TM wild type (TM^+/+) mice than in mice with low TM function (TM^Pro/-). After exposure to 9 Gy TBI, GT3 pre-treatment conferred 85% survival in TM^+/+ mice compared to only 50% in TM^Pro/-. Thus, GT3-mediated radiation lethality protection is partly dependent on endothelial TM. Significant post-TBI recovery of hematopoietic cells, particularly leukocytes, was observed in TM^+/+ mice (p = 0.003), but not in TM^Pro/- mice, despite the fact that GT3 induced higher levels of granulocyte colony stimulating factor (G-CSF) in TM^Pro/- mice (p = 0.0001). These data demonstrate a critical, G-CSF-independent, role for endothelial TM in GT3-mediated lethality protection and hematopoietic recovery after exposure to TBI and may point to new strategies to enhance the efficacy of current medical countermeasures in radiological/nuclear emergencies.

A tocotrienol-enriched formulation protects against radiation-induced changes in cardiac mitochondria without modifying late cardiac function or structure.

Sridharan, V., et.al (2015) Radiat Res.

Radiation-induced heart disease (RIHD) is a common and sometimes severe late side effect of radiation therapy for intrathoracic and chest wall tumors. It has been previously shown that local heart irradiation in a rat model caused prolonged changes in mitochondrial respiration and increased susceptibility to mitochondrial permeability transition pore (mPTP) opening. Because tocotrienols are known to protect against oxidative stress-induced mitochondrial dysfunction, this study examines the effects of tocotrienols on radiation-induced alterations in mitochondria, and structural and functional manifestations of RIHD. Male Sprague-Dawley rats received image-guided localized X irradiation to the heart to a total dose of 21 Gy. Twenty-four hours before irradiation, rats received a tocotrienol-enriched formulation or vehicle by oral gavage. Mitochondrial function and mitochondrial membrane parameters were studied at 2 weeks and 28 weeks after irradiation. In addition, cardiac function and histology were examined at 28 weeks. A single oral dose of the tocotrienol-enriched formulation preserved Bax/Bcl2 ratios and prevented mPTP opening and radiation-induced alterations in succinate-driven mitochondrial respiration. Nevertheless, the late effects of local heart irradiation pertaining to myocardial function and structure were not modified. Our studies suggest that a single dose of tocotrienols protects against radiation-induced mitochondrial changes, but these effects are not sufficient against long-term alterations in cardiac function or remodeling.

Progenitors Mobilized by Gamma-Tocotrienol as an Effective Radiation Countermeasure

Singh, V.K., et.al (2014). PLoS One.

The purpose of this study was to elucidate the role of gamma-tocotrienol (GT3)-mobilized progenitors in mitigating damage to mice exposed to a supralethal dose of cobalt-60 gamma-radiation. CD2F1 mice were transfused 24 h post-irradiation with whole blood or isolated peripheral blood mononuclear cells (PBMC) from donors that had received GT3 72 h prior to blood collection and recipient mice were monitored for 30 days. To understand the role of GT3-induced granulocyte colony-stimulating factor (G-CSF) in mobilizing progenitors, donor mice were administered a neutralizing antibody specific to G-CSF or its isotype before blood collection. Bacterial translocation from gut to heart, spleen and liver of irradiated recipient mice was evaluated by bacterial culture on enriched and selective agar media. Endotoxin in serum samples also was measured. It is also analyzed the colony-forming units in the spleens of irradiated mice. The results demonstrate that whole blood or PBMC from GT3-administered mice mitigated radiation injury when administered 24 h post-irradiation. Furthermore, administration of a G-CSF antibody to GT3-injected mice abrogated the efficacy of blood or PBMC obtained from such donors. Additionally, GT3-mobilized PBMC inhibited the translocation of intestinal bacteria to the heart, spleen, and liver, and increased colony forming unit-spleen (CFU-S) numbers in irradiated mice. Our data suggests that GT3 induces G-CSF, which mobilizes progenitors and these progenitors mitigate radiation injury in recipient mice. This approach using mobilized progenitor cells from GT3-injected donors could be a potential treatment for humans exposed to high doses of radiation.

Gamma tocotrienol, a potent radioprotector, preferentially upregulates expression of anti-apoptotic genes to promote intestinal cell survival

Suman, S., et.al (2013). Food and Chemial Toxicology.

Gamma tocotrienol (GT3) has been reported as a potent ameliorator of radiation-induced gastrointestinal (GI) toxicity when administered prophylactically. This study aimed to evaluate the role of GT3 mediated pro- and anti-apoptotic gene regulation in protecting mice from radiation-induced GI damage. Our study demonstrated that GT3-mediated protection of intestinal cells from a GI-toxic dose of radiation occurred via upregulation of antiapoptotic and downregulation of pro-apoptotic factors, both at the transcript as well as at the protein levels.

Granulocyte colony-stimulating factor antibody abrogates radioprotective efficacy of gamma-tocotrienol, a promising radiation countermeasure.

Kulkarni, S., et.al (2013). Cytokine.

This study aimed to determine the role of granulocyte colony-stimulating factor (G-CSF), induced by a promising radiation countermeasure, gamma tocotrienol (GT3), in protecting mice from lethal doses of ionizing radiation. CD2F1 mice were injected with an optimal dose of GT3 and a G-CSF antibody, and their 30-d survival was monitored. An appropriate antibody isotype was used as a control. Multiplex Luminex was used to analyze GT3-induced cytokines. G-CSF neutralization by exogenous administration of a G-CSF antibody was confirmed by analyzing serum cytokine levels. Our results demonstrate that GT3 significantly protected mice against ionizing radiation, and induced high levels of G-CSF in peripheral blood 24h after administration. Injection of a G-CSF neutralizing antibody to the GT3-treated mice resulted in complete neutralization of G-CSF and abrogation of its protective efficacy. Administration of a G-CSF antibody did not affect levels of other cytokines induced by GT3. Histopathology of bone marrow from GT3-treated and -irradiated mice demonstrated protection of the hematopoietic tissue, and also that such protection was abrogated by administering a G-CSF antibody. Our results suggest that induction of high levels of G-CSF by GT3 administration is responsible for its protective efficacy against radiation injury.

Mechanism of radioprotection by delta-tocotrienol: pharmacokinetics, pharmacodynamics and modulation of signaling pathways.

Satyamitra, M., et.al (2012). Br J Radiol.

The objective of this study was to investigate the correlation between in vivo δ-tocotrienol (DT3) pharmacokinetics, pharmacodynamics and radiation protection, and to evaluate the effect of DT3 pre-treatment on radiation-induced alterations in apoptotic and autophagic pathways. These data indicate that DT3 stimulates multilineage haematopoiesis, protects against radiation-induced apoptosis downstream of the mitochondria and stimulates cytoprotective autophagy. Apart from a potent antioxidant activity, DT3 may elicit survival advantage following irradiation by enhancing haematopoiesis and modulating signalling pathways.

Hematopoietic recovery and amelioration of radiation-induced lethality by the vitamin E isoform delta-tocotrienol.

Satyamitra, M.M., et.al (2011). Radiat Res.

δ-Tocotrienol (DT3), a vitamin E isoform, is associated with strong antioxidant and immunomodulatory properties. We confirmed the potent antioxidant activity in membrane systems and showed that DT3 is an effective radiation protector and mitigator. DT3 (4 μM, P < 0.001) inhibited lipid peroxidation in mouse liver microsomes and nitric oxide (NO) formation (20 μM DT3, P < 0.01) in RAW264.7 cells, a murine alveolar macrophage line. In CD2F1 mice exposed to lethal total-body radiation from a (60)Co γ-radiation source, a single subcutaneous (s.c.) injection of DT3 before or after irradiation produced a significant increase in 30-day survival. DT3 was effective from 18.75 to 300 mg/kg (–24 h, P < 0.001). A single dose of 150 or 300 mg/kg DT3 given 24 h before irradiation (radioprotection) resulted in dose reduction factors (DRFs) of 1.19 and 1.27, respectively (P < 0.001). Further, DT3 reduced radiation lethality when administered 2, 6 or 12 h after irradiation, and 150 mg/kg DT3 administered 2 h after exposure conferred a DRF of 1.1 (mitigation). The optimum schedule of 300 mg/kg DT3 24 h prior to 7 Gy significantly reduced pancytopenia compared to irradiated controls (P < 0.05). The large therapeutic potential of and multi-lineage hematopoietic recovery for DT3 warrants further studies.

Pentoxifylline Enhances the Radioprotective Properties of γ-Tocotrienol: Differential Effects on the Hematopoietic, Gastrointestinal and Vascular Systems

Berbee, M., et.al (2011). Radiat Res.

The vitamin E analog γ-tocotrienol (GT3) is a potent radioprotector and mitigator. This study was performed to (a) determine whether the efficacy of GT3 can be enhanced by the addition of the phosphodiesterase inhibitor pentoxifylline (PTX) and (b) to obtain information about the mechanism of action. Mice were injected subcutaneously with vehicle, GT3 [400 mg/kg 24 h before total-body irradiation (TBI)], PTX (200 mg/kg 30 min before TBI), or GT3+PTX before being exposed to 8.5–13 Gy TBI. Overall lethality, survival time and intestinal, hematopoietic and vascular injury were assessed. Cytokine levels in the bone marrow microenvironment were measured, and the requirement for endothelial nitric oxide synthase (eNOS) was studied in eNOS-deficient mice. GT3+PTX significantly improved survival compared to GT3 alone and provided full protection against lethality even after exposure to 12.5 Gy. GT3+PTX improved bone marrow CFUs, spleen colony counts and platelet recovery compared to GT3 alone. GT3 and GT3+PTX increased bone marrow plasma G-CSF levels as well as the availability of IL-1α, IL-6 and IL-9 in the early postirradiation phase. GT3 and GT3+PTX were equally effective in ameliorating intestinal injury and vascular peroxynitrite production. Survival studies in eNOS-deficient mice and appropriate controls revealed that eNOS was not required for protection against lethality after TBI. Combined treatment with GT3 and PTX increased postirradiation survival over that with GT3 alone by a mechanism that may depend on induction of hematopoietic stimuli. GT3+PTX did not reduce GI toxicity or vascular oxidative stress compared to GT3 alone. The radioprotective effect of either drug alone or both drugs in combination does not require the presence of eNOS.

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.

Exposure to γ-radiation causes rapid hematopoietic cell apoptosis and bone marrow suppression. However, there are no approved radiation countermeasures for the acute radiation syndrome. In this study, we demonstrated that natural δ-tocotrienol, one of the isomers of vitamin E, significantly enhanced survival in total body lethally irradiated mice. We explored 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.

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

Kulkami, S., et.al (2010). Radiat Res.

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(+), Lin(-)) 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 (Lin(-), 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 (naïve). GT3-treated irradiated mice maintained higher numbers of colonies (86% and 80% compared to naïve mice), thereby preserving the self-renewable 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.

γ-Tocotrienol Ameliorates Intestinal Radiation Injury and Reduces Vascular Oxidative Stress after Total-Body Irradiation by an HMG-CoA Reductase-Dependent Mechanism

Berbee, M., et.al (2009). Radiat Res.

Analogs of vitamin E (tocols) are under development as radioprophylactic agents because of their high efficacy and lack of toxicity. Gamma-tocotrienol (GT3) is of particular interest because, in addition to being an antioxidant, it also inhibits 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase and accumulates to greater extent in endothelial cells than other tocols. We addressed in vivo whether HMG-CoA reductase inhibition contributes to the radioprotection conferred by GT3. Groups of mice were treated with vehicle, mevalonate (the product of the reaction catalyzed by HMG-CoA reductase), GT3 alone or GT3 in combination with mevalonate. Lethality and standard parameters of injury to the hematopoietic, intestinal and vascular/endothelial systems were assessed after exposure to total-body irradiation. GT3 improved post-irradiation survival and decreased radiation-induced vascular oxidative stress, an effect that was reversible by mevalonate. GT3 also enhanced hematopoietic recovery, reduced intestinal radiation injury, and accelerated the recovery of soluble markers of endothelial function. These parameters were not reversed by mevalonate co-administration. Our data confirm GT3’s radioprophylactic properties against hematopoietic injury and, for the first time, demonstrate benefits in terms of protection against gastrointestinal and vascular injury. The radioprotective efficacy of GT3 against vascular injury is related to its properties as an HMG-CoA reductase inhibitor.

Dietary tocotrienol reduces UVB-induced skin damage and sesamin enhances tocotrienol effects in hairless mice.

Yamada, Y., et.al (2008). J Nutr Sci Vitaminol.

The present studies were undertaken to determine whether dietary tocotrienols and those with sesamin could protect the skin from damage induced by UVB irradiation in hairless mice fed four diets: a vitamin E-free diet, a 50mg/kg alpha-tocopherol) diet and a 229 mg/kg T-mix with 2 g/kg sesamin diet. The results suggest that dietary tocotrienols protect the skin more strongly than alpha-tocopherol against damage induced by UVB and sesamin enhances tocotrienol effects. The results suggest that dietary tocotrienols protect the skin more strongly than alpha-tocopherol against damage induced UVB and sesamin enhances tocotrienol effects.

Efficacy of topically applied tocopherols and tocotrienols in protection of murine skin from oxidative damage induced by UV-irradiation.

Weber, C., et.al (1997). Free Radic Biol Med.

To assess the efficacy of various forms of vitamin E in protection of skin from UV-light-induced oxidative stress, vitamin E (tocotrienol-rich fraction of palm oil, TRF) was applied to mouse skin and the contents of antioxidants before and after exposure to UV-light were measured. Four polypropylene plastic rings (1 cm2) were glued onto the animals’ backs, and 20 microliters 5% TRF in polyethylene glycol-400 (PEG) was applied to the skin circumscribed by two rings and 20 microliters PEG to the other two rings. After 2 h, the skin was washed and half of the sites were exposed to UV-irradiation (2.8 mW/cm2 for 29 mi: 3 MED). TRF treatment (n = 19 mice) increased mouse skin alpha-tocopherol 28 +/- 16-fold, alpha-tocotrienol 80 +/- 50-fold, gamma-tocopherol 130 +/- 108-fold, and gamma-tocotrienol 51 +/- 36-fold. A significantly higher percentage of alpha-tocopherol was present in the skin as compared with that in the applied TRF. After UV-irradiation, all vitamin E forms decreased significantly (p < .01), while a larger proportion of the vitamin E remained in PEG-treated (approximately 80%) compared with TRF-treated (approximately 40%) skin. Nonetheless, vitamin E concentrations in irradiated TRF-treated skin were significantly higher than in the nonirradiated PEG-treated (control) skin (p < .01). Thus, UV-irradiation of skin destroys its antioxidants: however, prior application of TRF to mouse skin results in preservation of vitamin E.

Efficacy of topically applied tocopherols and tocotrienols in protection of murine skin from oxidative damage induced by UV-irradiation.

Weber, C., et.al (1997). Free Radic Biol Med.

Ozone depletes tocopherols and tocotrienols topically applied to murine skin.

Thiele, J.J., et.al (1997). FEBS Lett.

To evaluate ozone damage to hairless mouse skin, two parameters of oxidative damage, vitamin E depletion and malondialdehyde (MDA) production, were measured in vitamin E-enriched and in control skin from mice exposed to ozone (10 ppm). A 5% vitamin E solution (tocotrienol-rich fraction, TRF) in polyethylene glycol (PEG) was applied to 2 sites on the back of hairless mice, PEG to 2 sites. After 2 h, the sites were washed, one of each pair of sites covered and the mice exposed ozone for 2 h. Ozone exposure (compared with covered sites) increased epidermal MDA in PEG-treated sites, while vitamin E was unchanged. In contrast, ozone exposure significantly depleted vitamin E in TRF-treated sites, while significant MDA accumulation was prevented. This is the first demonstration that ozone exposure causes damage to cutaneous lipids, an effect which can be attenuated by vitamin E application.

*In vitro* Studies
Article	Study objectives/ Findings
Radioprotective role of α-tocotrienol against oxidative damage in γ-irradiated human blood cultures Ahmed, M. M., et al., (2018). Bioscience Journal	This study investigate the antimutagenic and antioxidant ability of γ-tocotrienol using cytogenetic study and biochemical analysis in γ-irradiated human blood cultures. The study find γ-tocotrienol treatments reduced the number of radiation-induced micronuclei (MNi) frequencies and nucleuplasmic bridge (N bridge), which are usually increased in radiation cell. There is also an increase in the level of Caspase-3 in the treated cells compared to the untreated cells.
Tocotrienol-rich fraction attenuates UV-induced inflammaging: A bench to bedside study. Yap, W. N. (2017). Journal of Cosmetic Dermatology.	This study aimed to evaluate the possible palliative effects of a new topical nanoemulsion formulation containing tocotrienol-rich fraction (TRF) on UV-induced inflammation (erythema) of human skin. In vitro studies involving HaCaT keratinocytes revealed that TRF possesses marked anti-inflammatory properties, as indicated by the attenuation of UV-induced upregulation of pro-inflammatory cytokines. This study also show that TRF possesses comparable anti-inflammatory effects as diclofenac sodium by treating the sunburn on human skin.
The Vitamin E Analog Gamma-Tocotrienol (GT3) Suppresses Radiation-Induced Cytogenetic Damage. Pathak, R., et.al (2016). Pharm Res.	Ionizing radiation (IR) generates reactive oxygen species (ROS), which cause DNA double-strand breaks (DSBs) that are responsible for cytogenetic alterations. Because antioxidants are potent ROS scavengers, we determined whether the vitamin E isoform γ-tocotrienol (GT3), a radio-protective multifunctional dietary antioxidant, can suppress IR-induced cytogenetic damage. GT3 attenuates radiation-induced cytogenetic damage, possibly by affecting RAD50 expression. GT3 should be explored as a therapeutic to reduce the risk of developing genetic diseases after radiation exposure.
Tocotrienol-Rich Fraction from Rice Bran Demonstrates Potent Radiation Protection Activity. Krager, K.J., et.al (2015). Evid Based Complement Alternat Med.	The vitamin E analogs δ-tocotrienol (DT3) and γ-tocotrienol (GT3) have significant protective and mitigative capacity against the detrimental effects of ionizing radiation (IR). However, the expense of purification limits their potential use. This study examined the tocotrienol-rich fraction of rice bran (TRFRB) isolated from rice bran deodorizer distillate, a rice oil refinement waste product, to determine its protective effects against IR induced oxidative damage and H2O2. Several cell lines were treated with tocotrienols or TRFRB prior to or following exposure to H2O2 or IR. To determine the radioprotective capacity cells were analyzed for morphology, mitochondrial bioenergetics, clonogenic survival, glutathione oxidation, cell cycle, and migration rate. TRFRB displayed similar antioxidant activity compared to pure tocotrienols. Cells pretreated with TRFRB or DT3 exhibited preserved cell morphology and mitochondrial respiration when exposed to H2O2. Oxidized glutathione was decreased in TRFRB treated cells exposed to IR. TRFRB reversed mitochondrial uncoupling and protected cells migration rates following IR exposure. The protective antioxidant capacity of TRFRB treated cells against oxidative injury was similar to that of purified DT3. TRFRB effectively protects normal cells against IR induced injury suggesting that rice bran distillate may be an inexpensive and abundant alternate source.
Development and In Vitro Evaluation of Vitamin E-Enriched Nanoemulsion Vehicles Loaded with Genistein for Chemoprevention Against UVB-Induced Skin Damage. Brownlow, B., et.al (2015). J Pharm Sci.	This report investigates the development of nanoemulsified tocotrienol(T3)-rich fraction of red palm oil (Tocomin®), to yield an optimal NE delivery system for dermal photoprotection (z-average size <150 nm, ζ-potential ≈ -30 mV, polydispersity index < 0.25). Physicochemical characterization and photostability studies indicate NE formulations utilizing surfactant mixture (Smix) of Solutol® HS-15 (SHS15) blended with vitamin E TPGS (TPGS) as cosurfactant was significantly superior to formulations that utilized Lutrol® F68 (LF68) as the cosurfactant. A ratio of 60:40 of SHS15-TPGS-NE was further identified as lead Tocomin® NE topical platform using in vitro pharmaceutical skin reactivity studies that assess cutaneous irritancy and cytotoxicity. Prototype Tocomin® NE loaded with the antiphotocarcinogenic molecule Gen (Gen-Tocomin® NE) showed slow-release profile in both liquid and cream forms. Gen-Tocomin® NE also showed excellent biocompatibility, and provided substantial UVB protection to cultured subcutaneous L929 fibroblasts, indicating the great potential of our Tocomin® NE warranting further prototype development as topical pharmaceutical platform for skin photoprotection applications.
Delta-tocotrienol suppresses radiation-induced microRNA-30 and protects mice and human CD34+ cells from radiation injury. Li, X.H., et.al (2015). PLoS One.	It is reported that microRNA-30c (miR-30c) plays a key role in radiation-induced human cell damage through an apoptotic pathway. Herein we further evaluated radiation-induced miR-30 expression and mechanisms of delta-tocotrienol (DT3), a radiation countermeasure candidate, for regulating miR-30 in a mouse model and human hematopoietic CD34+ cells. CD2F1 mice were exposed to 0 (control) or 7-12.5 Gy total-body gamma-radiation, and CD34+ cells were irradiated with 0, 2 or 4 Gy of radiation. Single doses of DT3 (75 mg/kg, subcutaneous injection for mice or 2 μM for CD34+ cell culture) were administrated 24 h before irradiation and animal survival was monitored for 30 days. Mouse bone marrow (BM), jejunum, kidney, liver and serum as well as CD34+ cells were collected at 1, 4, 8, 24, 48 or 72 h after irradiation to determine apoptotic markers, pro-inflammatory cytokines interleukin (IL)-1β and IL-6, miR-30, and stress response protein expression. Our results showed that radiation-induced IL-1β release and cell damage are pathological states that lead to an early expression and secretion of miR-30b and miR-30c in mouse tissues and serum and in human CD34+ cells. DT3 suppressed IL-1β and miR-30 expression, protected against radiation-induced apoptosis in mouse and human cells, and increased survival of irradiated mice. Furthermore, an anti-IL-1β antibody downregulated radiation-induced NFκBp65 phosphorylation, inhibited miR-30 expression and protected CD34+ cells from radiation exposure. Knockdown of NFκBp65 by small interfering RNA (siRNA) significantly suppressed radiation-induced miR-30 expression in CD34+ cells. Our data suggest that DT3 protects human and mouse cells from radiation damage may through suppression of IL-1β-induced NFκB/miR-30 signaling.
Molecular dynamics guided design of tocoflexol: a new radioprotectant tocotrienol with enhanced bioavailability. Compadre, C.M., et.al (2014). Drug Dev Res.	There is a pressing need to develop safe and effective radioprotector/radiomitigator agents for use in accidental or terrorist-initiated radiological emergencies. Naturally occurring vitamin E family constituents, termed tocols, that include the tocotrienols, are known to have radiation-protection properties. These agents, which work through multiple mechanisms, are promising radioprotectant agents having minimal toxicity. Although α-tocopherol (AT) is the most commonly studied form of vitamin E, the tocotrienols are more potent than AT in providing radioprotection and radiomitigation. Unfortunately, despite their very significant radioprotectant activity, tocotrienols have very short plasma half-lives and require dosing at very high levels to achieve necessary therapeutic benefits. Thus, it would be highly desirable to develop new vitamin E analogues with improved pharmacokinetic properties, specifically increased elimination half-life and increased area under the plasma level versus time curve. The short elimination half-life of the tocotrienols is related to their low affinity for the α-tocopherol transfer protein (ATTP), the protein responsible for maintaining the plasma level of the tocols. Tocotrienols have less affinity for ATTP than does AT, and thus have a longer residence time in the liver, putting them at higher risk for metabolism and biliary excretion. We hypothesized that the low-binding affinity of tocotrienols to ATTP is due to the relatively more rigid tail structure of the tocotrienols in comparison with that of the tocopherols. Therefore, compounds with a more flexible tail would have better binding to ATTP and consequently would have longer elimination half-life and, consequently, an increased exposure to drug, as measured by area under the plasma drug level versus time curve (AUC). This represents an enhanced residence of drug in the systemic circulation. Based on this hypothesis, we developed a new class of vitamin E analogues, the tocoflexols, which maintain the superior bioactivity of the tocotrienols with the potential to achieve the longer half-life and larger AUC of the tocopherols.
Radioprotective efficacy of delta-tocotrienol, a vitamin E isoform, is mediated through granulocyte colony-stimulating factor. Singh V.K., et.al (2014). Life Sci.	The objectives of this study were to determine the cytokine induction by delta tocotrienol (DT3, a promising radiation countermeasure) and to investigate the role of granulocyte colony-stimulating factor (G-CSF) in its radioprotective efficacy against ionizing radiation in mice. Our study findings suggest that G-CSF induced by DT3 mediates its radioprotective efficacy against ionizing radiation in mice.
Delta-tocotrienol protects mice from radiation-induced gastrointestinal injury. Li, X.H., et.al (2013). Radiat Res.	It is recently demonstrated that natural delta-tocotrienol (DT3) significantly enhanced survival in total-body irradiated (TBI) mice, and protected mouse bone marrow cells from radiation-induced damage through Erk activation-associated mTOR survival pathways. Here, it is evaluated the effects and mechanisms of DT3 on survival of radiation-induced mouse acute gastrointestinal syndrome. DT3 (75-100 mg/kg) or vehicle was administered as a single subcutaneous injection to CD2F1 mice 24 h before 10-12 Gy (60)Co total-body irradiation at a dose rate of 0.6 Gy/min and survival was monitored. In a separate group of mice, jejunum sections were stained with hematoxylin and eosin and the surviving crypts in irradiated mice were counted. Apoptosis in intestinal epithelial cells was measured by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) staining and bacterial translocation from gut to heart, spleen and liver in irradiated mice were evaluated. DT3 (75 mg/kg) significantly enhanced survival in mice that received 10, 10.5, 11 or 12 Gy TBI. Administration of DT3 protected intestinal tissue, decreased apoptotic cells in jejunum and inhibited gut bacterial translocation in irradiated mice. Furthermore, DT3 significantly inhibited radiation-induced production of pro-inflammatory factors interleukin-1β and -6 and suppressed expression of protein tyrosine kinase 6 (PTK6), a stress-induced kinase that promotes apoptosis in mouse intestinal cells. The data demonstrate that administration of DT3 protected mice from radiation-induced gastrointestinal system damage.
Mobilization of progenitor cells into peripheral blood by gamma-tocotrienol: a promising radiation countermeasure. Ray, S., et.al (2013). Int Immunopharmacol.	Gamma-tocotrienol (GT3), a vitamin E isoform, is shown to induce high levels of granulocyte colony stimulating factor (G-CSF) in mice. G-CSF is a key cytokine used for stimulation of hematopoiesis, and mobilization of hematopoietic stem and progenitor cells into peripheral blood. GT3 is also shown to induce vascular endothelial growth factor (VEGF), another important cytokine necessary for vasculogenesis and endothelial progenitor mobilization. Since GT3 induces both these cytokines, we tested whether GT3 mobilizes hematopoietic and endothelial progenitors in mice. GT3 (200mg/kg) was injected in 10-week-old CD2F1 mice and mobilization of progenitors in peripheral blood was analyzed at 24, 48, and 72 h post-administration. Circulating hematopoietic progenitor cells (HPCs, Lin (-), cKit(+)), endothelial progenitor cells (EPCs, Lin(-), CD34(+), Flk(+)), and stromal progenitor cells (SPCs, Lin(-), CD29(+), CD105(+)) in peripheral blood mononuclear cells (PBMCs) were analyzed simultaneously by flow cytometry. Mobilized HPCs, EPCs and SPCs in PBMC were also measured by colony-forming unit (CFU) assay in progenitor-specific media. Three groups of mice received vehicle, GT3 and GT3 plus AMD3100, a receptor antagonist used to enhance mobilization. GT3 induced significant mobilization of all three progenitor cell types compared to vehicle in peripheral blood; AMD3100 enhanced GT3-induced mobilization even further. Mobilization of progenitor cells in peripheral blood by GT3 indicates that GT3 can be used as an alternative to G-CSF and VGEF to mobilize HPCs and EPCs.
Mechanisms underlying the radioprotective properties of γ-tocotrienol: comparative gene expression profiling in tocol-treated endothelial cells. Berbee, M., et.al (2012). Genes Nutr.	Among the eight naturally occurring vitamin E analogs, γ-tocotrienol (GT3) is a particularly potent radioprophylactic agent in vivo. Moreover, GT3 protects endothelial cells from radiation injury not only by virtue of its antioxidant properties but also by inhibition of 3-hydroxy-3-methyl-glutaryl-CoA (HMG-CoA) reductase and by improving the availability of the nitric oxide synthase cofactor tetrahydrobiopterin. Nevertheless, the precise mechanisms underlying the superior radioprotective properties of GT3 compared with other tocols are not known. This study, therefore, examined the differences in gene expression profiles between GT3 and its tocopherol counterpart, γ-tocopherol, as well as between GT3 and α-tocopherol in human endothelial cells. Cells were treated with vehicle or the appropriate tocol for 24 h, after which total RNA was isolated and genome-wide gene expression profiles were obtained using the Illumina platform. GT3 was far more potent in inducing gene-expression changes than α-tocopherol or γ-tocopherol. In particular, GT3 induced multiple changes in pathways known to be of importance in the cellular response to radiation exposure. Affected GO functional clusters included response to oxidative stress, response to DNA damage stimuli, cell cycle phase, and regulation of cell death, regulation of cell proliferation, hematopoiesis, and blood vessel development. These results form the basis for further studies to determine the exact importance of differentially affected GO functional clusters in endothelial radioprotection by GT3.
Reduction of radiation-induced vascular nitrosative stress by the vitamin E analog, γ-tocotrienol: evidence of a role for tetrahydrobiopterin Berbee, M., et.al (2011). Int J Radiat Oncol Biol Phys.	The vitamin E analog γ-tocotrienol (GT3) is a powerful radioprotector. GT3 reduces post-radiation vascular peroxynitrite production, an effect dependent on inhibition of hydroxy-methyl-glutaryl coenzyme A (HMG-CoA) reductase. HMG-CoA reductase inhibitors mediate their pleiotropic effects via eNOS that requires the co-factor tetrahydrobiopterin (BH4). This study investigated the effects of radiation on BH4 bioavailability and of GT3 on BH4 metabolism.
A novel mechanism of natural vitamin E tocotrienol activity: involvement of ERβ signal transduction Comitato, R., et.al (2009). Am J Physiol Endocrinol Metab.	Vitamin E is a generic term used to indicate all tocopherol (TOC) and tocotrienol (TT) derivates. In the last few years, several papers have shown that a TT-rich fraction (TTRF) extracted from palm oil inhibits proliferation and induces apoptosis in a large number of cancer cells. However, the molecular mechanism(s) involved in TT action is still unclear. In the present study, we proposed for the first time a novel mechanism for TT activity that involves estrogen receptor (ER) signaling. In silico simulations and in vitro binding analyses indicated a high affinity of TTs for ERβ but not for ERα. In addition, in ERβ-containing MDA-MB-231 breast cancer cells, we demonstrated that TTs increase the ERβ translocation into the nucleus, which in turn activates estrogen-responsive genes (MIC-1, EGR-1 and cathepsin D), as demonstrated by cell preincubation with the ER inhibitor ICI-182,780. Finally, we observed that TT treatment is associated with alteration of cell morphology, DNA fragmentation, and caspase-3 activation. Altogether, these experiments elucidated the molecular mechanism underling γ- and δ-TT effects.

Review Article
Article	Study objectives/ Findings
Antioxidant Tocols as Radiation Countermeasures (Challenges to be Addressed to Use Tocols as Radiation Countermeasures in Humans) Nukala U. et al., (2018). Antioxidants.	Radiation countermeasures fall under three categories, radiation protectors, radiation mitigators, and radiation therapeutics. Radiation protectors are agents that are administered before radiation exposure to protect from radiation-induced injuries by numerous mechanisms, including scavenging free radicals that are generated by initial radiochemical events. Radiation mitigators are agents that are administered after the exposure of radiation but before the onset of symptoms by accelerating the recovery and repair from radiation-induced injuries. Whereas radiation therapeutic agents administered after the onset of symptoms act by regenerating the tissues that are injured by radiation. Vitamin E is an antioxidant that neutralizes free radicals generated by radiation exposure by donating H atoms. The vitamin E family consists of eight different vitamers, including four tocopherols and four tocotrienols. Though alpha-tocopherol was extensively studied in the past, tocotrienols have recently gained attention as radiation countermeasures. Despite several studies performed on tocotrienols, there is no clear evidence on the factors that are responsible for their superior radiation protection properties over tocopherols. Their absorption and bioavailability are also not well understood. In this review, the authors discuss tocopherol’s and tocotrienol’s efficacy as radiation countermeasures and identify the challenges to be addressed to develop them into radiation countermeasures for human use in the event of radiological emergencies.
Vitamin E therapy beyond cancer: Tocopherol versus tocotrienol Peh, H.Y., et.al (2016).	The discovery of vitamin E (α-tocopherol) began in 1922 as a vital component required in reproduction. Today, there are eight naturally occurring vitamin E isoforms, namely α-, β-, γ- and δ-tocopherol and α-, β-, γ- and δ-tocotrienol. Vitamin E is potent antioxidants, capable of neutralizing free radicals directly by donating hydrogen from its chromanol ring. α-Tocopherol is regarded the dominant form in vitamin E as the α-tocopherol transfer protein in the liver binds mainly α-tocopherol, thus preventing its degradation. That contributed to the oversight of tocotrienols and resulted in less than 3% of all vitamin E publications studying tocotrienols. Nevertheless, tocotrienols have been shown to possess superior antioxidant and anti-inflammatory properties over α-tocopherol. In particular, inhibition of 3-hydroxy-3-methylglutaryl-coenzyme A reductase to lower cholesterol, attenuating inflammation via downregulation of transcription factor NF-κB activation, and potent radioprotectant against radiation damage are some properties unique to tocotrienols, not tocopherols. Aside from cancer, vitamin E has also been shown protective in bone, cardiovascular, eye, nephrological and neurological diseases. In light of the different pharmacological properties of tocopherols and tocotrienols, it becomes critical to specify which vitamin E isoform(s) are being studied in any future vitamin E publications. This review provides an update on vitamin E therapeutic potentials, protective effects and modes of action beyond cancer, with comparison of tocopherols against tocotrienols. With the concerted efforts in synthesizing novel vitamin E analogs and clinical pharmacology of vitamin E, it is likely that certain vitamin E isoform(s) will be therapeutic agents against human diseases besides cancer.
γ-Tocotrienol as a Promising Countermeasure for Acute Radiation Syndrome: Current Status. Singh, V.K., et.al (2016). Int J Mol Sci.	The hazard of ionizing radiation exposure due to nuclear accidents or terrorist attacks is ever increasing. Despite decades of research, still, there is a shortage of non-toxic, safe and effective medical countermeasures for radiological and nuclear emergency. To date, the U.S. Food and Drug Administration (U.S. FDA) has approved only two growth factors, Neupogen (granulocyte colony-stimulating factor (G-CSF), filgrastim) and Neulasta (PEGylated G-CSF, pegfilgrastim) for the treatment of hematopoietic acute radiation syndrome (H-ARS) following the Animal Efficacy Rule. Promising radioprotective efficacy results of γ-tocotrienol (GT3; a member of the vitamin E family) in the mouse model encouraged its further evaluation in the nonhuman primate (NHP) model. These studies demonstrated that GT3 significantly aided the recovery of radiation-induced neutropenia and thrombocytopenia compared to the vehicle controls; these results particularly significant after exposure to 5.8 or 6.5 Gray (Gy) whole body γ-irradiation. The stimulatory effect of GT3 on neutrophils and thrombocytes (platelets) was directly and positively correlated with dose; a 75 mg/kg dose was more effective compared to 37.5 mg/kg. GT3 was also effective against 6.5 Gy whole body γ-irradiation for improving neutrophils and thrombocytes. Moreover, a single administration of GT3 without any supportive care was equivalent, in terms of improving hematopoietic recovery, to multiple doses of Neupogen and two doses of Neulasta with full supportive care (including blood products) in the NHP model. GT3 may serve as an ultimate radioprotector for use in humans, particularly for military personnel and first responders. In brief, GT3 is a promising radiation countermeasure that ought to be further developed for U.S. FDA approval for the ARS indication.
Vitamin E: tocopherols and tocotrienols as potential radiation countermeasures Singh, V.K., et.al (2013). J Radiat Res.	Despite the potential devastating health consequences of intense total-body irradiation, and the decades of research, there still remains a dearth of safe and effective radiation countermeasures for emergency, radiological/ nuclear contingencies that have been fully approved and sanctioned for use by the US FDA. Vitamin E is a well-known antioxidant, effective in scavenging free radicals generated by radiation exposure. Vitamin E analogs, collectively known as tocols, have been subject to active investigation for a long time as radioprotectors in patients undergoing radiotherapy and in the context of possible radiation accidents or terrorism scenarios. Eight major isoforms comprise the tocol group: four tocopherols and four tocotrienols. A number of these agents and their derivatives are being investigated actively as radiation countermeasures using animal models, and several appear promising. Although the tocols are well-recognized as potent antioxidants and are generally thought to mediate radioprotection through ‘ free radical quenching’, recent studies have suggested several alternative mechanisms: most notably, an ‘indirect effect’ of tocols in eliciting specific species of radioprotective growth factors/ cytokines such as granulocyte colony-stimulating factor (G-CSF). The radioprotective efficacy of at least two tocols has been abrogated using a neutralizing antibody of G-CSF. Based on encouraging results of radioprotective efficacy, laboratory testing of gamma-tocotrienol has moved from a small rodent model to a large nonhuman primate model for preclinical evaluation, In this brief review we identify and discuss selected tocols and their derivatives currently under development as radiation countermeasures, and attempt to describe in some detail their in vivo efficacy.
Novel drug to ameliorate gastrointestinal normal tissue radiation toxicity in clinical practice: Berbee, M., et.al (2012). Curr Opin Support Palliat Care.	To give an overview of promising novel agents under development for the prevention and reduction of gastrointestinal radiation injury. Gastrointestinal radiation injury is the most important dose-limiting factor during radiotherapy of the abdomen or pelvis. It may severely affect the quality of life both during radiotherapy treatment and in cancer survivors. To date, there are no agents that can prevent or reduce intestinal radiation injury. Hence, there is an urgent need for the development of novel drugs to ameliorate intestinal toxicity during and after radiotherapy. This review summarizes the several agents that have been shown to reduce intestinal radiation injury in animals. Further research is needed to investigate their safety and efficacy in patients receiving radiotherapy for abdominal or pelvic tumours.

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In-vivo / Animal Studies

In vitro Studies

Review Article