Vitamin E succinate induces apoptosis in human prostate cancer cells: role for Fas in vitamin E succinate-triggered apoptosis

Published on Thursday, 18 July 2013

Abstract

The apoptosis-triggering properties of vitamin E succinate (VES, RRR-alpha-tocopheryl succinate) for human LNCaP and PC-3 prostate carcinoma cells and normal PrEC human prostate epithelial cells were investigated.

LNCaP and PC-3 cells were sensitive to VES-induced apoptosis, with 100% and 60% of cells undergoing apoptosis after three days of treatment with 10 micrograms of VES/ml, respectively. PrEC cells were resistant to VES-induced apoptosis.

Treatment of prostate cells with agonistic anti-Fas antibody triggered apoptosis in approximately 50% of PC-3 cells within 48 hours, whereas LNCaP and PrEC cells were resistant. Prostate cells simultaneously treated with VES and agonistic anti-Fas antibodies revealed 1) no effect on PrEC cells, 2) an additive effect on Fas-sensitive PC-3 cells, and 3) a synergistic effect on LNCaP cells.

VES treatment of LNCaP cells caused depletion of cytosolic 43-kDa Fas, enhanced membrane levels of 43-kDa Fas, and induced Fas sensitivity. PC-3 cells expressed high levels of membrane 43-kDa Fas that were enhanced by VES treatments. Fas ligand expression by LNCaP cells was enhanced by VES treatments. In summary, VES triggers apoptosis in human prostate carcinoma cells but not normal prostate cells in vitro, and VES modulates Fas signaling.

INTRODUCTION

Vitamin E succinate (VES, RRR-alpha-tocopheryl succinate), a derivative of vitamin E, is currently being characterized for its chemopreventive and chemotherapeutic potential. VES has been shown to inhibit the proliferation of several transformed cell types, including human prostate cancer cells. Previous studies have shown VES to inhibit tumor cell growth by a variety of mechanisms, including DNA synthesis arrest, induction of apoptosis, G1 cell cycle blockage, induced cellular differentiation, induced secretion and activation of transforming growth factor (TGF)-, enhanced expression of TGF- type II receptors, and enhanced cell surface expression of Fas (CD95).

A considerable amount of discussion regarding a hypothesis that vitamin E can protect against cancer, especially prostate cancer, has been generated by the recent alpha-Tocopherol, -Carotene Cancer Prevention Study of 29,133 male smokers aged 50-69 years in Finland. When the data were evaluated for the effect of daily supplements with 50 mg of synthetic dl-alpha-tocopheryl acetate for five to eight years on lung cancer incidence and mortality, vitamin E was reported to have no effect on the occurrence of lung cancer and no apparent effect on total mortality.

When the data were evaluated for effects of synthetic dl- -tocopheryl acetate on prostate cancer, it was reported that vitamin E-supplemented subjects had a 32% decrease in incidence of clinical prostate cancer but not in latent cancer. Furthermore, it was reported that prostate cancer mortality was 41% lower among the vitamin E-supplemented men. In view of these findings, it is important to point out that the studies reported here involve a different form of vitamin E, a succinate derivative of natural vitamin E (RRR-alpha-tocopheryl), RRR-alpha-tocopheryl succinate or VES. It is also important to point out that succinic acid and acetate esters of vitamin E [the naturally occurring single isomer form (RRR-alpha) or the synthetic mixture of 8 stereoisomers (dl-alpha)] are not active antioxidants unless the esterification is hydrolyzed and the free tocopherol is regenerated (reviewed in Reference 22). Because the acetate and succinate forms of RRR-alpha-Tocopherol and dl-alpha-Tocopherol are available over-the-counter as dietary supplements, it is of interest to ascertain potential mechanisms of action of these vitamin E compounds in human prostate cancer cells in cell culture and in animal models.

One interesting and consistent effect of VES on cancer cells in culture is its ability to induce apoptosis (9,15,17- 19,23,24). Apoptosis or programmed cell death is a well-established phenomenon in tissue development and homeostasis. This physiological process has been described as having independent triggering and transducing pathways, depending not only on the initiating agent itself, but on the environmental context of the cell or tissue. However, it appears that all cells undergoing the final execution phase of apoptosis exhibit similar end-stage morphological characteristics, such as cell membrane blebbing, nuclear condensation, and DNA fragmentation, caused by activation of common death-inducing proteases and endonucleases.

Defects in key components of death signaling pathways provide a survival advantage to cells and have been implicated as factors contributing to malignancy. Because induction of apoptosis by antineoplastic drugs is vital to the treatment of most cancers, alterations in the cell death signaling pathway may result in drug resistance. Defective signaling through the Fas-Fas ligand pathway has been demonstrated in certain cancer cell types. It is of interest that studies show that sensitivity to Fas-Fas ligand pathway(s) can be restored in certain tumor cells. Recent studies by this laboratory have demonstrated that Fas-resistant human breast cancer cells become responsive to agonistic anti-Fas antibody-triggered apoptosis after pretreatment or cotreatment with VES.

Fas (known as CD95 or APO-1) is a member of the tumor necrosis factor receptor and nerve growth factor receptor superfamily (reviewed in Reference 37). Fas contains a cytoplasmic death domain and, when trimerizered at the cell surface via Fas ligand or agonistic anti-Fas antibody, can initiate an apoptotic cascade that results in rapid cell death. Although Fas-induced apoptosis has been most extensively studied in T lymphocytes, recent studies show Fas expression in a wide range of cell types, including normal prostate cells. Downstream effects of Fas receptor activation are under investigation in an effort to identify the cellular components critical to its role in cell death, inasmuch as the presence of immunoreactive Fas is not itself sufficient to guarantee functionality of the receptor. Furthermore, the presence of altered Fas signaling has been demonstrated in prostate, breast, colon, and central nervous system cancer cells.

Here we report on the apoptosis-inducing abilities of VES on PrEC, LNCaP, and PC-3 cells. We show that 1) VES is an effective inducer of apoptosis in PC-3 and LNCaP human prostate carcinoma cells, but not in normal human prostate epithelial cells, PrEC, in cell culture, 2) VES-induced apoptosis can be augmented in an additive fashion in PC-3 cells and, in a synergistic fashion, in LNCaP cells by cotreatment with agonistic anti-Fas antibody, 3) cotreatment of cells with VES + agonistic Fas antibody induces apoptosis in LNCaP cells, which involves, at least in part, Fas signaling, 4) VES treatment enhances cell surface expression of Fas (CD95) by human prostate carcinoma cells, and 5) VES treatment enhances cell surface expression of Fas ligand.

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