tocotrienol--delta and Breast-Neoplasms

Neoadjuvant treatment of breast cancer is applied to an increasing extent, but treatment response varies and side effects pose a challenge. The vitamin E isoform delta-tocotrienol might enhance the efficacy of chemotherapy and reduce the risk of side effects. The aim of this study was to investigate the clinical effect of delta-tocotrienol combined with standard neoadjuvant treatment and the possible association between detectable circulating tumor DNA (ctDNA) during and after neoadjuvant treatment with pathological treatment response. This open-label, randomized phase II trial included 80 women with newly diagnosed, histologically verified breast cancer randomized to standard neoadjuvant treatment alone or in combination with delta-tocotrienol. There was no difference in the response rate or frequency of serious adverse events between the two arms. We developed a multiplex digital droplet polymerase chain reaction (ddPCR) assay for the detection of ctDNA in breast cancer patients that targets a combination of two methylations specific for breast tissue (LMX1B and ZNF296) and one cancer specific methylation (HOXA9). The sensitivity of the assay increased when the cancer specific marker was combined with the ones specific to breast tissue (p < 0.001). The results did not show any association between ctDNA status and pathological treatment response, neither at midterm nor before surgery.

Topics: Biological Assay; Breast Neoplasms; Drug-Related Side Effects and Adverse Reactions; Female; Humans; Neoadjuvant Therapy

In response to low oxygen supply, cancer cells elevate production of HIF-1α, a hypoxia-inducible transcription factor that subsequently acts to stimulate blood vessel formation and promote survival. Studies were conducted to determine the role of δ-tocotrienol and a semisynthetic δ-tocotrienol oxazine derivative, compound 44, on +SA mammary tumor cell hypoxic response. Treatment with 150 µM CoCl2 induced a hypoxic response in +SA mammary tumor cells as evidenced by a large increase in HIF-1α levels, and combined treatment with compound 44 attenuated this response. CoCl2-induced hypoxia was also associated with a large increase in Akt/mTOR signaling, activation of downstream targets p70S6K and eIF-4E1, and a significant increase in VEGF production, and combined treatment with compound 44 blocked this response. Additional in vivo studies showed that intralesional treatment with compound 44 in BALB/c mice bearing +SA mammary tumors significantly decreased the levels of HIF-1α, and this effect was associated with a corresponding decrease in Akt/mTOR signaling and activation of downstream targets p70S6 kinase and eIF-4E1. These findings demonstrate that treatment with the δ-tocotrienol oxazine derivative, compound 44, significantly attenuates +SA mammary tumor cell compensatory responses to hypoxia and suggests that this compound may provide benefit in the treatment of rapidly growing solid breast tumors.

Topics: Animals; Breast Neoplasms; Cell Hypoxia; Cell Line, Tumor; Cobalt; Female; Gene Expression Regulation, Neoplastic; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Mammary Neoplasms, Animal; Mice; Signal Transduction; Vitamin E

The vitamin E family members γ- and δ-tocotrienols (2 and 3, respectively) are known natural products with documented anticancer activities. Redox-silent structural modifications, such as esterification, etherification and carbamoylation, of 2 and 3 significantly enhanced their anticancer activities. However, hit-to-lead optimization of tocotrienols and their analogs was yet to be reported at the outset of the project described herein. Subjecting the chroman ring of 2 and 3 to the electrophilic substitution reactions, namely, Mannich and Lederer-Manasse procedures, afforded 42 new products. These included the 3,4-dihydro-1,3-oxazines 3-29 and 35-44, Mannich bases 30-31, and the hydroxymethyl analogs 32-34. Of these, the δ-tocotrienol analogs 8, 11, 18, 24, 25, 27, and 40 inhibited the proliferation of the highly metastatic +SA mammary epithelial cancer cell line, with IC(50) values in the nanomolar (nM) range. In NCI's 60 human tumor cell line panel, 8, 17, 38, and 40 showed antiproliferative activity, with nM GI(50) values. The δ-tocotrienol analogs 10 and 38 inhibited the migration of the highly metastatic human breast cancer cell line MDA-MB-231 with IC(50) values of 1.3 and 1.5 μM, respectively, in the wound-healing assay. A dose of 0.5 mg/day for 14 days of one of the active analogs, 30, significantly slowed the growth of +SA mammary tumors in the syngeneic BALB/c mouse model, compared to the vehicle- and the parent γ-tocotrienol-treated control groups. Electrophilic substitution reactions promoted tocotrienols to lead level and can enable their future use to control metastatic breast malignancies.

Topics: Animals; Breast Neoplasms; Cell Line, Tumor; Cell Movement; Cell Proliferation; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Humans; Inhibitory Concentration 50; Magnetic Resonance Spectroscopy; Mice; Mice, Inbred BALB C; Plant Oils; Tocotrienols

Tocotrienols and tocopherols are members of the vitamin E family, with similar structures; however, only tocotrienols have been reported to achieve potent anti-cancer effects. The study described here has evaluated anti-cancer activity of vitamin E to elucidate mechanisms of cell death, using human breast cancer cells.. Anti-cancer activity of a tocotrienol-rich fraction (TRF) and a tocotrienol-enriched fraction (TEF) isolated from palm oil, as well as pure vitamin E analogues (α-tocopherol, α-, δ- and γ-tocotrienols) were studied using highly aggressive triple negative MDA-MB-231 cells and oestrogen-dependent MCF-7 cells, both of human breast cancer cell lines. Cell population growth was evaluated using a Coulter particle counter. Cell death mechanism, poly(ADP-ribose) polymerase cleavage and levels of NF-κB were determined using commercial ELISA kits.. Tocotrienols exerted potent anti-proliferative effects on both types of cell by inducing apoptosis, the underlying mechanism of cell death being ascertained using respective IC50 concentrations of all test compounds. There was marked induction of apoptosis in both cell lines by tocotrienols compared to treatment with Paclitaxel, which was used as positive control. This activity was found to be associated with cleavage of poly(ADP-ribose) polymerase (a DNA repair protein), demonstrating involvement of the apoptotic cell death signalling pathway. Tocotrienols also inhibited expression of nuclear factor kappa-B (NF-κB), which in turn can increase sensitivity of cancer cells to apoptosis.. Tocotrienols induced anti-proliferative and apoptotic effects in association with DNA fragmentation, poly(ADP-ribose) polymerase cleavage and NF-κB inhibition in the two human breast cancer cell lines.

Topics: Antineoplastic Agents, Phytogenic; Apoptosis; Breast Neoplasms; Cell Count; Cell Proliferation; Chromans; DNA Fragmentation; Enzyme-Linked Immunosorbent Assay; Female; Humans; Inhibitory Concentration 50; MCF-7 Cells; NF-kappa B p50 Subunit; Paclitaxel; Palm Oil; Plant Oils; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerases; Proteolysis; Reagent Kits, Diagnostic; Signal Transduction; Tocotrienols; Vitamin E

Vitamin E (VE) is a generic term that represents a family of compounds composed of various tocopherol and tocotrienol isoforms. Tocotrienols display potent anti-angiogenic and antiproliferative activities. Redox-silent tocotrienol analogues also display potent anticancer activity. The ultimate objective of this study was to develop semisynthetically C-6-modified redox-silent tocotrienol analogues with enhanced antiproliferative and anti-invasive activities as compared to their parent compound. Examples of these are carbamate and ether analogues of alpha-, gamma-, and delta-tocotrienols (1-3). Various aliphatic, olefinic, and aromatic substituents were used. Steric limitation, electrostatic, hydrogen bond donor (HBD) and hydrogen bond acceptor (HBA) properties were varied at this position and the biological activities of these derivatives were tested. Three-dimensional quantitative structure-activity relationship (3D QSAR) studies were performed using Comparative Molecular Field (CoMFA) and Comparative Molecular Similarity Indices Analyses (CoMSIA) to better understand the structural basis for biological activity and guide the future design of more potent VE analogues.

Topics: Antineoplastic Agents; Breast Neoplasms; Cell Proliferation; Drug Design; Drug Screening Assays, Antitumor; Female; Humans; Models, Molecular; Molecular Structure; Neoplasm Invasiveness; Oxidation-Reduction; Quantitative Structure-Activity Relationship; Stereoisomerism; Tocotrienols

Tocotrienols are vitamin E members with potent antiproliferative activity against preneoplastic and neoplastic mammary epithelial cells with little or no effect on normal cell growth or functions. However, physicochemical and pharmacokinetic properties greatly limit their use as therapeutic agents. Tocotrienols' chemical instability, poor water solubility, NPC1L1-mediated transport, and rapid metabolism are examples of such obstacles which hinder the therapeutic use of these valuable natural products. Vitamin E esters like α-tocopheryl succinate were prepared to significantly improve chemical and metabolic stability, water solubility, and potency. Thus, 12 semisynthetic tocotrienol ester analogues 4-15 were prepared by direct esterification of natural tocotrienol isomers with various acid anhydrides or chlorides. Esters 4-15 were evaluated for their ability to inhibit the proliferation and migration of the mammary tumor cells +SA and MDA-MB-231, respectively. Esters 5, 9, and 11 effectively inhibited the proliferation of the highly metastatic +SA rodent mammary epithelial cells with IC(50) values of 0.62, 0.51, and 0.86μM, respectively, at doses that had no effect on immortalized normal mouse CL-S1 mammary epithelial cells. Esters 4, 6, 8-10, and 13 inhibited 50% of the migration of the human metastatic MDA-MB-231 breast cancer cells at a single 5μM dose in wound-healing assay. The most active ester 9 was 1000-fold more water-soluble and chemically stable versus its parent α-tocotrienol (1). These findings strongly suggest that redox-silent tocotrienol esters may provide superior therapeutic forms of tocotrienols for the control of metastatic breast cancer.

Topics: Animals; Antineoplastic Agents; Breast Neoplasms; Cell Line, Tumor; Cell Movement; Cell Proliferation; Epithelial Cells; Esters; Female; Humans; Mammary Glands, Animal; Mice; Oxidation-Reduction; Rats; Solubility; Tocotrienols

Breast cancer is the most common malignancy among women, with an age-specific incidence profile. During the last years much evidence has accumulated demonstrating the anticancer activity of tocotrienols (T3), a subfamily of natural vitamin E (VE). In this study, mouse and human breast cancer cells (with or without HER-2/neu oncogene overexpression) were used to investigate the anticancer effect of alpha-, gamma-, and delta-tocotrienols in comparison with alpha-tocopheryl succinate (alpha-TOS), a synthetic derivative with widely recognized anticancer properties.. Human and mouse breast cancer cell lines were used. The effect of VE compounds on cell viability was investigated using Alamar Blue assay. Apoptosis was assessed by propidium iodide and JC-1 staining. Expression of senescence-associated markers was evaluated by RT-PCR and Western blot analysis was used to examine the changes in the expression levels of HER-2/neu.. gamma- and delta-Tau3 reduced cell viability with IC(50) values of less than half those of alpha-T3 and alpha-TOS. gamma- and delta-Tau3, and alpha-TOS to a lesser extent, induced apoptosis possibly via the mitochondrial pathway, and the expression of senescent-like growth arrest markers as p53, p21, and p16. Both alpha-TOS and tocotrienols downregulated HER-2/neu in tumor cells overexpressing this oncogene, but this effect did not seem to be essential for the antitumor activity of these compounds.. We demonstrate that in HER-2/neu breast cancer cells, the non-alpha form of T3 shows stronger anticancer activity than the synthetic VE-derivative alpha-TOS and this effect occurs independently from the inhibition of HER-2/neu oncogene expression.

Topics: alpha-Tocopherol; Animals; Antineoplastic Agents; Apoptosis; Blotting, Western; Breast Neoplasms; Cell Line, Tumor; Cell Survival; Chromans; Drug Screening Assays, Antitumor; Female; Gene Expression Regulation, Neoplastic; Humans; Inhibitory Concentration 50; Mice; Receptor, ErbB-2; Tocotrienols; Vitamin E

Vitamin E derivative, RRR-alpha-tocopheryl succinate (vitamin E succinate, VES), is a potent pro-apoptotic agent, inducing apoptosis by restoring both transforming growth factor-beta (TGF-beta) and Fas (CD95) apoptotic signaling pathways that contribute to the activation of c-Jun N-terminal kinase (JNK)-mediated apoptosis. Objectives of these studies were to characterize signaling events involved in the pro-apoptotic actions of a naturally occurring form of vitamin E, delta-tocotrienol, and a novel vitamin E analog, alpha-tocopherol ether acetic acid analog [alpha-TEA; 2,5,7,8-tetramethyl-2R-(4R,8R,12-trimethyltridecyl)chroman-6-yloxyacetic acid]. Like VES, alpha-TEA and delta-tocotrienol induced estrogen-nonresponsive MDA-MB-435 and estrogen-responsive MCF-7 human breast cancer cells to undergo high levels of apoptosis in a concentration- and time-dependent fashion. Like VES, the two compounds induced either no or lower levels of apoptosis in normal human mammary epithelial cells and immortalized but nontumorigenic human MCF-10A cells. The pro-apoptotic mechanisms triggered by the structurally distinct alpha-TEA and delta-tocotrienol were identical to those previously reported for VES, that is, alpha-TEA- and delta-tocotrienol-induced apoptosis involved up-regulation of TGF-beta receptor II expression and TGF-beta-, Fas- and JNK-signaling pathways. These data provide a better understanding of the anticancer actions of a dietary form of vitamin E (delta-tocotrienol) and a novel nonhydrolyzable vitamin E analog (alpha-TEA).

Topics: Anticarcinogenic Agents; Apoptosis; Blotting, Western; Breast Neoplasms; Cell Division; Cell Line, Tumor; Cell Survival; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; fas Receptor; Female; Gene Expression Regulation, Neoplastic; Humans; Receptors, Transforming Growth Factor beta; Signal Transduction; Tocopherols; Vitamin E

The apoptosis-inducing properties of RRR-alpha-, beta-, gamma-, and delta-tocopherols, alpha-, gamma-, and delta-tocotrienols, RRR-alpha-tocopheryl acetate (vitamin E acetate), and RRR-alpha-tocopheryl succinate (vitamin E succinate) were investigated in estrogen-responsive MCF7 and estrogen-nonresponsive MDA-MB-435 human breast cancer cell lines in culture. Apoptosis was characterized by two criteria: 1) morphology of 4,6-diamidino-2-phenylindole-stained cells and oligonucleosomal DNA laddering. Vitamin E succinate, a known inducer of apoptosis in several cell lines, including human breast cancer cells, served as a positive control. The estrogen-responsive MCF7 cells were more susceptible than the estrogen-nonresponsive MDA-MB-435 cells, with concentrations for half-maximal response for tocotrienols (alpha, gamma, and delta) and RRR-delta-tocopherol of 14, 15, 7, and 97 micrograms/ml, respectively. The tocotrienols (alpha, gamma, and delta) and RRR-delta-tocopherol induced MDA-MB-435 cells to undergo apoptosis, with concentrations for half-maximal response of 176, 28, 13, and 145 micrograms/ml, respectively. With the exception of RRR-delta-tocopherol, the tocopherols (alpha, beta, and gamma) and the acetate derivative of RRR-alpha-tocopherol (RRR-alpha-tocopheryl acetate) were ineffective in induction of apoptosis in both cell lines when tested within the range of their solubility, i.e., 10-200 micrograms/ml. In summary, these studies demonstrate that naturally occurring tocotrienols and RRR-delta-tocopherol are effective apoptotic inducers for human breast cancer cells.

Topics: Antioxidants; Apoptosis; Breast Neoplasms; Chromans; Chromatin; DNA, Neoplasm; Female; Humans; Neoplasms, Hormone-Dependent; Tocotrienols; Tumor Cells, Cultured; Vitamin E

Potential antiproliferative effects of tocotrienols, the major vitamin E component in palm oil, were investigated on the growth of both estrogen-responsive (ER+) MCF7 human breast cancer cells and estrogen-unresponsive (ER-) MDA-MB-231 human breast cancer cells, and effects were compared with those of alpha-tocopherol (alphaT). The tocotrienol-rich fraction (TRF) of palm oil inhibited growth of MCF7 cells in both the presence and absence of estradiol with a nonlinear dose-response but such that complete suppression of growth was achieved at 8 microg/mL. MDA-MB-231 cells were also inhibited by TRF but with a linear dose-response such that 20 microg/mL TRF was needed for complete growth suppression. Separation of the TRF into individual tocotrienols revealed that all fractions could inhibit growth of both ER+ and ER- cells and of ER+ cells in both the presence and absence of estradiol. However, the gamma- and delta-fractions were the most inhibitory. Complete inhibition of MCF7 cell growth was achieved at 6 microg/mL of gamma-tocotrienol/delta-tocotrienol (gammaT3/deltaT3) in the absence of estradiol and 10 microg/mL of deltaT3 in the presence of estradiol, whereas complete suppression of MDA-MB-231 cell growth was not achieved even at concentrations of 10 microg/mL of deltaT3. By contrast to these inhibitory effects of tocotrienols, alphaT had no inhibitory effect on MCF7 cell growth in either the presence or the absence of estradiol, nor on MDA-MB-231 cell growth. These results confirm studies using other sublines of human breast cancer cells and demonstrate that tocotrienols can exert direct inhibitory effects on the growth of breast cancer cells. In searching for the mechanism of inhibition, studies of the effects of TRF on estrogen-regulated pS2 gene expression in MCF7 cells showed that tocotrienols do not act via an estrogen receptor-mediated pathway and must therefore act differently from estrogen antagonists. Furthermore, tocotrienols did not increase levels of growth-inhibitory insulin-like growth factor binding proteins (IGFBP) in MCF7 cells, implying also a different mechanism from that proposed for retinoic acid inhibition of estrogen-responsive breast cancer cell growth. Inhibition of the growth of breast cancer cells by tocotrienols could have important clinical implications not only because tocotrienols are able to inhibit the growth of both ER+ and ER- phenotypes but also because ER+ cells could be growth-inhibited in the presence as well as

Topics: Breast Neoplasms; Cell Division; Estradiol; Female; Humans; Insulin-Like Growth Factor Binding Proteins; Receptors, Estrogen; Tocotrienols; Tumor Cells, Cultured; Vitamin E

Tocotrienols are a form of vitamin E, having an unsaturated isoprenoid side-chain rather than the saturated side-chain of tocopherols. The tocotrienol-rich fraction (TRF) from palm oil contains alpha-tocopherol and a mixture of alpha-, gamma- and delta-tocotrienols. Earlier studies have shown that tocotrienols display anticancer activity. We previously reported that TRF, alpha-, gamma- and delta-tocotrienols inhibited proliferation of estrogen receptor-negative MDA-MB-435 human breast cancer cells with 50% inhibitory concentrations (IC50) of 180, 90, 30 and 90 microg/mL, respectively, whereas alpha-tocopherol had no effect at concentrations up to 500 microg/mL. Further experiments with estrogen receptor-positive MCF-7 cells showed that tocotrienols also inhibited their proliferation, as measured by [3H] thymidine incorporation. The IC50s for TRF, alpha-tocopherol, alpha-, gamma- and delta-tocotrienols were 4, 125, 6, 2 and 2 microg/mL, respectively. Tamoxifen, a widely used synthetic antiestrogen inhibits the growth of MCF-7 cells with an IC50 of 0.04 microg/mL. We tested 1:1 combinations of TRF, alpha-tocopherol and the individual tocotrienols with tamoxifen in both cell lines. In the MDA-MB-435 cells, all of the combinations were found to be synergistic. In the MCF-7 cells, only 1:1 combinations of gamma- or delta-tocotrienol with tamoxifen showed a synergistic inhibitory effect on the proliferative rate and growth of the cells. The inhibition by tocotrienols was not overcome by addition of excess estradiol to the medium. These results suggest that tocotrienols are effective inhibitors of both estrogen receptor-negative and -positive cells and that combinations with tamoxifen should be considered as a possible improvement in breast cancer therapy.

Topics: Antineoplastic Agents, Hormonal; Antioxidants; Breast Neoplasms; Cell Division; Cell Survival; Chromans; Dietary Fats, Unsaturated; Drug Interactions; Estrogen Antagonists; Female; Humans; Palm Oil; Plant Oils; Receptors, Estrogen; Tamoxifen; Tocotrienols; Tumor Cells, Cultured; Vitamin E