Delta-Tocotrienol Modulates Glutamine Dependence by Inhibiting ASCT2 and LAT1 Transporters in Non-Small Cell Lung Cancer (NSCLC) Cells: A Metabolomic Approach

Rajasinghe LD, Hutchings M, Gupta SV

Metabolites. 2019 Mar 13;9(3). pii: E50. doi: 10.3390/metabo9030050.

Abstract

The growth and development of non-small cell lung cancer (NSCLC) primarily depends on glutamine. Both glutamine and essential amino acids (EAAs) have been reported to upregulate mTOR in NSCLC, which is a bioenergetics sensor involved in the regulation of cell growth, cell survival, and protein synthesis. Seen as novel concepts in cancer development, ASCT2 and LAT transporters allow glutamine and EAAs to enter proliferating tumors as well as send a regulatory signal to mTOR. Blocking or downregulating these glutamine transporters in order to inhibit glutamine uptake would be an excellent therapeutic target for treatment of NSCLC. This study aimed to validate the metabolic dysregulation of glutamine and its derivatives in NSCLC using cellular 1H-NMR metabolomic approach while exploring the mechanism of delta-tocotrienol (δT) on glutamine transporters, and mTOR pathway. Cellular metabolomics analysis showed significant inhibition in the uptake of glutamine, its derivatives glutamate and glutathione, and some EAAs in both cell lines with δT treatment. Inhibition of glutamine transporters (ASCT2 and LAT1) and mTOR pathway proteins (P-mTOR and p-4EBP1) was evident in Western blot analysis in a dose-dependent manner. Our findings suggest that δT inhibits glutamine transporters, thus inhibiting glutamine uptake into proliferating cells, which results in the inhibition of cell proliferation and induction of apoptosis via downregulation of the mTOR pathway.

Ekicier Acar S, Sarıcaoğlu MS, Çolak A, Aktaş Z, Sepici Dinçel A

Eur J Ophthalmol. 2019 Mar 11:1120672119833271. doi: 10.1177/1120672119833271. [Epub ahead of print]

Abstract

PURPOSE::

We aimed to create mechanic optic nerve injury model in rats and investigate the neuroprotective effects of topical Coenzyme Q10 + Vitamin E (CoQ + Vit.E) molecules on retinal ganglion cells.

METHODS::

Mechanic optic nerve injury model was created in the right eyes of rats (n = 12). Rats were divided into two groups: glaucoma model with sham treatment and topical CoQ + Vit.E treatment. Treatment was applied for 4 weeks. Glial fibrillary acidic protein, Brn-3a antibody, and anti-Iba1 were examined by immunohistochemistry. Glial fibrillary acidic protein, Bax, Bcl-xL, and Tfam protein expression were measured by Western blot analysis.

RESULTS::

The number of Brn-3a-positive retinal ganglion cell was 15.0 ± 1.0 (min: 14, max: 16) in sham treatment group and 22.2 ± 4.8 (min: 18, max: 29) in topical CoQ10 + Vit.E treatment group. The protection of Brn-3a in CoQ10 + Vit.E was statistically significant (p < 0.05). Glial fibrillary acidic protein-positive astroglial counts were recorded as 11.7 ± 2.1 (min: 10, max: 14) in sham treatment and 2.5 ± 1.5 (min: 1, max: 4) in topical CoQ10 + Vit.E treatment group (p < 0.05). Topical CoQ10 + Vit.E treatment also decreased Iba1 expression in the retina of mechanic optic nerve injury groups. CoQ10 + Vit.E treatment prevented apoptotic cell death by increasing Bcl-xL protein expression. Also, CoQ10 + Vit.E preserved Tfam protein expression in the retina.

CONCLUSION::

This study has shown that in glaucoma treatment the neuron protecting effect of topical CoQ10 + Vit.E molecules can be valuable.

Emami MR, Safabakhsh M, Alizadeh S, Asbaghi O, Khosroshahi MZ

J Hum Hypertens. 2019 Mar 7. doi: 10.1038/s41371-019-0192-0. [Epub ahead of print]

Abstract

Although emerging evidence suggests that vitamin E may contribute to blood pressure improvement, the effects of vitamin E on systolic blood pressure (SBP), diastolic blood pressure (DBP), and mean arterial pressure (MAP) are still controversial. The aim was to evaluate the influence of vitamin E on SBP, DBP, and MAP through meta-analysis. We identified all studies that assessed the effect of vitamin Esupplementation on SBP, DBP, and MAP from PubMed/Medline, SCOPUS, and Google scholar up to March 2018. Weighted mean differences (WMD) and 95% confidence interval (CI) were expressed as effect size. Pre-specified subgroup analysis was conducted to evaluate potential sources of heterogeneity. Meta-regression analyses were performed to investigate association between blood pressure-lowering effects of vitamin E and duration of follow-up and dose of treatment. Eighteen trials, comprising 839 participants met the eligibility criteria. Results of this study showed that compared to placebo, SBP decreased significantly in vitamin E group (WMD = -3.4 mmHg, 95% CI = -6.7 to -0.11, P < 0.001), with a high heterogeneity across the studies (I² = 94.0%, P < 0.001). Overall, there were no significant effects on DBP and MAP. This meta-analysis suggested that vitamin E supplements decreased only SBP and had no favorable effect on DBP and MAP.

Tang KD, Liu J, Russell PJ, Clements JA, Ling MT

Int J Mol Sci. 2019 Mar 7;20(5). pii: E1164. doi: 10.3390/ijms20051164.

Abstract

Emerging evidence suggests that gamma-tocotrienol (γ-T3), a vitamin E isomer, has potent anti-cancer properties against a wide-range of cancers. γ-T3 not only inhibited the growth and survival of cancer cells in vitro, but also suppressed angiogenesis and tumour metastasis under in vivo conditions. Recently, γ-T3 was found to target cancer stem cells (CSCs), leading to suppression of tumour formation and chemosensitisation. Despite its promising anti-cancer potential, the exact mechanisms responsible for the effects of γ-T3 are still largely unknown. Here, we report the identification of Ang-1 (Angiopoietin-1)/Tie-2 as a novel γ-T3 downstream target. In prostate cancer cells, γ-T3 treatment leads to the suppression of Ang-1 at both the mRNA transcript and protein levels. Supplementing the cells with Ang-1 was found to protect them against the anti-CSC effect of γ-T3. Intriguingly, inactivation of Tie-2, a member receptor that mediates the effect of Ang-1, was found to significantly enhance the cytotoxic effect of γ-T3 through activation of AMP-activated protein kinase (AMPK) and subsequent interruption of autophagy. Our results highlighted the therapeutic potential of using γ-T3 in combination with a Tie-2 inhibitor to treat advanced prostate cancer.

Ghani SMA, Goon JA, Azman NHEN, Zakaria SNA, Hamid Z, Ngah WZW

Clinics (Sao Paulo). 2019 Mar 7;74:e688. doi: 10.6061/clinics/2019/e688.

Abstract

OBJECTIVES:

This study aims to compare the differential gene expression resulting from tocotrienol-rich fraction and α-tocopherol supplementation in healthy older adults.

METHODS:

A total of 71 eligible subjects aged 50 to 55 years from Gombak and Kuala Lumpur, Malaysia, were divided into three groups and supplemented with placebo (n=23), α-tocopherol (n=24) or tocotrienol-rich fraction (n=24). Blood samples were collected at baseline and at 3 and 6 months of supplementation for microarray analysis.

RESULTS:

The number of genes altered by α-tocopherol was higher after 6 months (1,410) than after 3 months (273) of supplementation. α-Tocopherol altered the expression of more genes in males (952) than in females (731). Similarly, tocotrienol-rich fraction modulated the expression of more genes after 6 months (1,084) than after 3 months (596) and affected more genes in males (899) than in females (781). α-Tocopherol supplementation modulated pathways involving the response to stress and stimuli, the immune response, the response to hypoxia and bacteria, the metabolism of toxins and xenobiotics, mitosis, and synaptic transmission as well as activated the mitogen-activated protein kinase and complement pathways after 6 months. However, tocotrienol-rich fraction supplementation affected pathways such as the signal transduction, apoptosis, nuclear factor kappa B kinase, cascade extracellular signal-regulated kinase-1 and extracellular signal-regulated kinase-2, immune response, response to drug, cell adhesion, multicellular organismal development and G protein signaling pathways.

CONCLUSION:

Supplementation with either α-tocopherol or tocotrienol-rich fraction affected the immune and drug response and the cell adhesion and signal transduction pathways but modulated other pathways differently after 6 months of supplementation, with sex-specific responses.

Garg S, Sadhukhan R, Banerjee S, Savenka AV, Basnakian AG, McHargue V, Wang J, Pawar SA, Ghosh SP, Ware J, Hauer-Jensen M, Pathak R

Antioxidants (Basel). 2019 Mar 6;8(3). pii: E57. doi: 10.3390/antiox8030057.

Abstract

Natural antioxidant gamma-tocotrienol (GT3), a vitamin E family member, provides intestinal radiation protection. We seek to understand whether this protection is mediated via mucosal epithelial stem cells or sub-mucosal mesenchymal immune cells. Vehicle- or GT3-treated male CD2F1 mice were exposed to total body irradiation (TBI). Cell death was determined by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. Villus height and crypt depth were measured with computer-assisted software in tissue sections. Functional activity was determined with an intestinal permeability assay. Immune cell recovery was measured with immunohistochemistry and Western blot, and the regeneration of intestinal crypts was assessed with ex vivo organoid culture. A single dose of GT3 (200 mg/kg body weight (bwt)) administered 24 h before TBI suppressed cell death, prevented a decrease in villus height, increased crypt depth, attenuated intestinal permeability, and upregulated occludin level in the intestine compared to the vehicle treated group. GT3 accelerated mesenchymal immune cell recovery after irradiation, but it did not promote ex vivo organoid formation and failed to enhance the expression of stem cell markers. Finally, GT3 significantly upregulated protein kinase B or AKT phosphorylation after TBI. Pretreatment with GT3 attenuates TBI-induced structural and functional damage to the intestine, potentially by facilitating intestinal immune cell recovery. Thus, GT3 could be used as an intestinal radioprotector.

Pang KL, Chin KY

Molecules. 2019 Mar 6;24(5). pii: E923. doi: 10.3390/molecules24050923.

Abstract

Obesity is a major risk factor for diabetes, and these two metabolic conditions cause significant healthcare burden worldwide. Chronic inflammation and increased oxidative stress due to exposure of cells to excess nutrients in obesity may trigger insulin resistance and pancreatic β-cell dysfunction. Tocotrienol, as a functional food component with anti-inflammatory, antioxidant, and cell signaling-mediating effects, may be a potential agent to complement the current management of obesity and diabetes. The review aimed to summarize the current evidence on the anti-obesity and antidiabetic effects of tocotrienol. Previous studies showed that tocotrienol could suppress adipogenesis and, subsequently, reduce body weight and fat mass in animals. This was achieved by regulating pathways of lipid metabolism and fatty acid biosynthesis. It could also reduce the expression of transcription factors regulating adipogenesis and increase apoptosis of adipocytes. In diabetic models, tocotrienol was shown to improve glucose homeostasis. Activation of peroxisome proliferator-activated receptors was suggested to be responsible for these effects. Tocotrienol also prevented multiple systemic complications due to obesity and diabetes in animal models through suppression of inflammation and oxidative stress. Several clinical trials have been conducted to validate the antidiabetic of tocotrienol, but the results were heterogeneous. There is no evidence showing the anti-obesity effects of tocotrienol in humans. Considering the limitations of the current studies, tocotrienol has the potential to be a functional food component to aid in the management of patients with obesity and diabetes.

Wei CW, Yu YL, Chen YH, Hung YT, Yiang GT

Oncol Rep. 2019 Mar;41(3):2060-2066. doi: 10.3892/or.2019.6958. Epub 2019 Jan 9.

Abstract

Triple‑negative breast cancers (TNBCs) lack the estrogen receptor, progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2). Therefore, hormone or targeted therapies are not effective in the treatment of TNBC and thus the development of novel therapeutic strategies is crucial. Methotrexate (MTX), a folate antagonist, has been used in the treatment of various types of cancer; however, the anticancer effects of MTX treatment on breast cancer have thus far been ineffective. Vitamin E variants and derivatives have been applied for cancer therapy. Previous studies have indicated that vitamin E variants and derivatives exert distinct anticancer effects on different types of cancer. However, whether MTX plus vitamin E variants or its derivatives can inhibit TNBC remains unclear. The aim of the present study was to examine the anticancer effects and mechanisms of action of MTX in combination with vitamin E variants (α‑tocopherol) and derivatives (α‑tocopherol succinate) on TNBC. In the present study, MTT assay and western blot analysis were used to determine the cell survival rates and protein levels. The results demonstrated that combination treatment with MTX and α‑tocopherol suppressed TNBC cell proliferation. In addition, various concentrations of MTX exerted distinct cytotoxic effects on α‑tocopherol succinate‑treated cells. Furthermore, high‑dose MTX enhanced α‑tocopherol succinate‑induced anticancer activity; however, low‑dose MTX inhibited α‑tocopherolsuccinate‑induced anticancer activity. The present study also demonstrated that caspase‑3 activation and poly(adenosine diphosphate‑ribose) polymerase cleavage were observed in the α‑tocopherol succinate/MTX‑treated cells. In conclusion, the findings of the present study demonstrated that high‑dose MTX enhanced anticancer activity in α‑TOS‑treated TNBC, while low‑dose MTX reduced anticancer activity in α‑TOS‑treated TNBC.

Thomsen CB, Andersen RF, Steffensen KD, Adimi P, Jakobsen A

Pharmacol Res. 2019 Mar;141:392-396. doi: 10.1016/j.phrs.2019.01.017. Epub 2019 Jan 9.

Abstract

Delta tocotrienol has anti-neoplastic activity as demonstrated in several in-vitro and in-vivo investigations. The effect relies on inhibition of different pathways. It also has antiangiogenic activity, and an additive effect to bevacizumab may be expected. The present study was a phase II trial of bevacizumab combined with tocotrienol in chemotherapy refractory ovarian cancer. The study also included analysis of circulating tumor specific HOXA9 methylated DNA (HOXA9 meth-ctDNA) during treatment. The study included 23 patients. The rate of disease stabilization was 70% with very low toxicity. The median PFS was 6.9 months and the median OS 10.9 months, which is rather high compared to the current literature. A division of the patients according to level of HOXA9 meth-ctDNA already after the first cycle of chemotherapy resulted in two groups of patients with different prognoses. Patients with an increasing level of HOXA9 meth-ctDNA had a median PFS and OS of 1.4 and 4.3 months, respectively, compared to 7.8 and 12 months in the group with stable or decreasing levels. The combination of bevacizumab and tocotrienol is potent in chemotherapy refractory ovarian cancer. The level of HOXA9 meth-ctDNA after one cycle of chemotherapy holds important prognostic information.

Rodriguez-Duarte J, Galliussi G, Dapueto R, Rossello J, Malacrida L, Kamaid A, Schopfer FJ, Escande C, López GV, Batthyány C

Br J Pharmacol. 2019 Mar;176(6):757-772. doi: 10.1111/bph.14561. Epub 2019 Feb 3.

Abstract

BACKGROUND AND PURPOSE:

Atherosclerosis is characterized by chronic low-grade inflammation with concomitant lipid accumulation in the arterial wall. Anti-inflammatory and anti-atherogenic properties have been described for a novel class of endogenous nitroalkenes (nitrated-unsaturated fatty acids), formed during inflammation and digestion/absorption processes. The lipid-associated antioxidant α-tocopherol is transported systemically by LDL particles including to the atheroma lesions. To capitalize on the overlapping and complementary salutary properties of endogenous nitroalkenes and α-tocopherol, we designed and synthesized a novel nitroalkene-α-tocopherol analogue (NATOH) to address chronic inflammation and atherosclerosis, particularly at the lesion sites.

EXPERIMENTAL APPROACH:

We synthesized NATOH, determined its electrophilicity and antioxidant capacity and studied its effects over pro-inflammatory and cytoprotective pathways in macrophages in vitro. Moreover, we demonstrated its incorporation into lipoproteins and tissue both in vitro and in vivo, and determined its effect on atherosclerosis and inflammatory responses in vivo using the Apo E knockout mice model.

KEY RESULTS:

NATOH exhibited similar antioxidant capacity to α-tocopherol and, due to the presence of the nitroalkenyl group, like endogenous nitroalkenes, it exerted electrophilic reactivity. NATOH was incorporated in vivo into the VLDL/LDL lipoproteins particles to reach the atheroma lesions. Furthermore, oral administration of NATOH down-regulated NF-κB-dependent expression of pro-inflammatory markers (including IL-1β and adhesion molecules) and ameliorated atherosclerosis in Apo E knockout mice.

CONCLUSIONS AND IMPLICATIONS:

In toto, the data demonstrate a novel pharmacological strategy for the prevention of atherosclerosis based on a creative, natural and safe drug delivery system of a non-conventional anti-inflammatory compound (NATOH) with significant potential for clinical application.