Koyamangalath Krishnan

Gamma (γ) tocopherol upregulates peroxisome proliferator activated receptor (PPAR) gamma (γ) expression in SW 480 human colon cancer cell lines

BackgroundTocopherols are lipid soluble antioxidants that exist as eight structurally different isoforms. The intake of γ-tocopherol is higher than α-tocopherol in the average US diet. The clinical results of the effects of vitamin E as a cancer preventive agent have been inconsistent. All published clinical trials with vitamin E have used α-tocopherol. Recent epidemiological, experimental and molecular studies suggest that γ-tocopherol may be a more potent chemopreventive form of vitamin E compared to the more-studied α-tocopherol. γ-Tocopherol exhibits differences in its ability to detoxify nitrogen dioxide, growth inhibitory effects on selected cancer cell lines, inhibition of neoplastic transformation in embryonic fibroblasts, and inhibition of cyclooxygenase-2 (COX-2) activity in macrophages and epithelial cells. Peroxisome proliferator activator receptor γ (PPARγ) is a promising molecular target for colon cancer prevention. Upregulation of PPARγ activity is anticarcinogenic through its effects on downstream genes that affect cellular proliferation and apoptosis. The thiazolidine class of drugs are powerful PPARγ ligands. Vitamin E has structural similarity to the thiazolidine, troglitazone. In this investigation, we tested the effects of both α and γ tocopherol on the expression of PPARγ mRNA and protein in SW 480 colon cancer cell lines. We also measured the intracellular concentrations of vitamin E in SW 480 colon cancer cell lines.ResultsWe have discovered that the α and γ isoforms of vitamin E upregulate PPARγ mRNA and protein expression in the SW480 colon cancer cell lines. γ-Tocopherol is a better modulator of PPARγ expression than α-tocopherol at the concentrations tested. Intracellular concentrations increased as the vitamin E concentration added to the media was increased. Further, γ-tocopherol-treated cells have higher intracellular tocopherol concentrations than those treated with the same concentrations of α-tocopherol.ConclusionOur data suggest that both α and γ tocopherol can upregulate the expression of PPARγ which is considered an important molecular target for colon cancer chemoprevention. We show that the expression of PPARγ mRNA and protein are increased and these effects are more pronounced with γ-tocopherol. γ-Tocopherols ability to upregulate PPARγ expression and achieve higher intracellular concentrations in the colonic tissue may be relevant to colon cancer prevention. We also show that the intracellular concentrations of γ-tocopherol are several fold higher than α-tocopherol. Further work on other colon cancer cell lines are required to quantitate differences in the ability of these forms of vitamin E to induce apoptosis, suppress cell proliferation and act as PPAR ligands as well as determine their effects in conjunction with other chemopreventive agents. Upregulation of PPARγ by the tocopherols and in particular by γ-tocopherol may have relevance not only to cancer prevention but also to the management of inflammatory and cardiovascular disorders.

Development of gamma (γ)-tocopherol as a colorectal cancer chemopreventive agent

Abstract Nutritional factors play an important role in the prevention and promotion of colorectal cancer. Vitamin E is a generic term that describes a group of lipid-soluble chain-breaking antioxidants that includes tocopherols and tocotrienols. Vitamin E occurs in nature as eight structurally related forms that include four tocopherols and four tocotrienols. Vitamin E is a potent membrane-soluble antioxidant. Antioxidants like vitamin E (tocopherols) may prevent colon cancer through several different cellular and molecular mechanisms. Vitamin E in the American diet is primarily available in plant-oil rich foods such as vegetable oils, seeds and nuts and these foods vary widely in their content of α-tocopherol and γ-tocopherol [1] . Vitamin E may help prevent colon cancer by decreasing the formation of mutagens arising from the oxidation of fecal lipids, by decreasing oxidative stress in the epithelial cells of the colon and by molecular mechanisms that influence cell death, cell cycle and transcriptional events. Most epidemiological, experimental and clinical studies have evaluated the α-isoform and not the γ-isoform of vitamin E. Recent epidemiological, experimental and mechanistic evidence suggests that γ-tocopherol may be a more potent cancer chemopreventive agent than α-tocopherol. The differences in chemical reactivity, metabolism and biological activity may contribute to these differences in the effects of γ-tocopherol when compared with α-tocopherol. The rationale supporting the development of γ-tocopherol as a colorectal cancer preventive agent is reviewed here.

Tocopherols and the treatment of colon cancer

Abstract: Colorectal cancer is the second most common cause of cancer deaths in the United States. Vitamin E (VE) and other antioxidants may help prevent colon cancer by decreasing the formation of mutagens arising from the free radical oxidation of fecal lipids or by “non‐antioxidant” mechanisms. VE is not a single molecule, but refers to at least eight different molecules, that is, four tocopherols and four tocotrienols. Methods: Both animal models and human colon cancer cell lines were used to evaluate the chemopreventive potential of different forms of VE. Rats were fed diets deficient in tocopherols or supplemented with either α‐tocopherol or γ‐tocopherol. Half the rats in each of these groups received normal levels of dietary Fe and the other half Fe at eight times the normal level. In our cell experiments, we looked at the role of γ‐tocopherol in upregulating peroxisome proliferator‐activated receptor‐γ (PPAR‐γ) in the SW 480 human cell line. Results: Rats fed the diets supplemented with α‐tocopherol had higher levels of VE in feces, colonocytes, plasma, and liver than did rats fed diets supplemented with γ‐tocopherol. Dietary Fe levels did not influence tocopherol levels in plasma, liver, or feces. For colonocytes, high dietary Fe decreased tocopherol levels. Rats fed the γ‐tocopherol‐supplemented diets had lower levels of fecal lipid hydroperoxides than rats fed the α‐tocopherol‐supplemented diets. Ras‐p21 levels were significantly lower in rats fed the γ‐tocopherol‐supplemented diets compared with rats fed the α‐tocopherol‐supplemented diets. High levels of dietary Fe were found to promote oxidative stress in feces and colonocytes. Our data with the SW480 cells suggest that both α‐ and γ‐tocopherol upregulate PPAR‐γ mRNA and protein expression. γ‐tocopherol was, however, found to be a better enhancer of PPAR‐γ expression than α‐tocopherol at the concentrations tested.

Tocotrienols inhibit AKT and ERK activation and suppress pancreatic cancer cell proliferation by suppressing the ErbB2 pathway.

Tocotrienols are members of the vitamin E family but, unlike tocopherols, possess an unsaturated isoprenoid side chain that confers superior anti-cancer properties. The ability of tocotrienols to selectively inhibit the HMG-CoA reductase pathway through posttranslational degradation and to suppress the activity of transcription factor NF-κB could be the basis for some of these properties. Our studies indicate that γ- and δ-tocotrienols have potent antiproliferative activity in pancreatic cancer cells (Panc-28, MIA PaCa-2, Panc-1, and BxPC-3). Indeed both tocotrienols induced cell death (>50%) by the MTT cell viability assay in all four pancreatic cancer cell lines. We also examined the effects of the tocotrienols on the AKT and the Ras/Raf/MEK/ERK signaling pathways by Western blotting analysis. γ- and δ-tocotrienol treatment of cells reduced the activation of ERK MAP kinase and that of its downstream mediator RSK (ribosomal protein S6 kinase) in addition to suppressing the activation of protein kinase AKT. Suppression of activation of AKT by γ-tocotrienol led to downregulation of p-GSK-3β and upregulation accompanied by nuclear translocation of Foxo3. These effects were mediated by the downregulation of Her2/ErbB2 at the messenger level. Tocotrienols but not tocopherols were able to induce the observed effects. Our results suggest that the tocotrienol isoforms of vitamin E can induce apoptosis in pancreatic cancer cells through the suppression of vital cell survival and proliferative signaling pathways such as those mediated by the PI3-kinase/AKT and ERK/MAP kinases via downregulation of Her2/ErbB2 expression. The molecular components for this mechanism are not completely elucidated and need further investigation.

Chemoprevention for colorectal cancer.

3. Experimental models for colorectal cancer chemoprevention studies . . . . . . . . . . . . . . . 201 3.1. Animal models of chemical carcinogenesis . . . . . . . . . . . . . . . . . . . . . . . . . . . 201 3.2. Aberrant crypt foci (ACF) assay . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 202 3.3. Colon cancer cell lines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 202 3.4. Transfection models . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 202 3.5. Genetic and knock-out models . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 202

Differential effects of pravastatin and simvastatin on the growth of tumor cells from different organ sites.

3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR) inhibitors, commonly known as statins, may possess cancer preventive and therapeutic properties. Statins are effective suppressors of cholesterol synthesis with a well-established risk-benefit ratio in cardiovascular disease prevention. Mechanistically, targeting HMGCR activity primarily influences cholesterol biosynthesis and prenylation of signaling proteins. Pravastatin is a hydrophilic statin that is selectively taken up by a sodium-independent organic anion transporter protein-1B1 (OATP1B1) exclusively expressed in liver. Simvastatin is a hydrophobic statin that enters cells by other mechanisms. Poorly-differentiated and well-differentiated cancer cell lines were selected from various tissues and examined for their response to these two statins. Simvastatin inhibited the growth of most tumor cell lines more effectively than pravastatin in a dose dependent manner. Poorly-differentiated cancer cells were generally more responsive to simvastatin than well-differentiated cancer cells, and the levels of HMGCR expression did not consistently correlate with response to statin treatment. Pravastatin had a significant effect on normal hepatocytes due to facilitated uptake and a lesser effect on prostate PC3 and colon Caco-2 cancer cells since the OATP1B1 mRNA and protein were only found in the normal liver and hepatocytes. The inhibition of cell growth was accompanied by distinct alterations in mitochondrial networks and dramatic changes in cellular morphology related to cofilin regulation and loss of p-caveolin. Both statins, hydrophilic pravastatin and hypdrophobic simvastatin caused redistribution of OATP1B1 and HMGCR to perinuclear sites. In conclusion, the specific chemical properties of different classes of statins dictate mechanistic properties which may be relevant when evaluating biological responses to statins.

Colon cancer chemoprevention: Clinical development of aspirin as a chemopreventive agent

We have studied aspirin as a potential chemopreventive for colorectal cancer, completing Phase I studies on aspirin pharmacology and potential biomarker assays (prostaglandins, PGE2 and PGF2α and cyclooxygenase modulation) in normal human subjects. These studies have determined the optimal dose of aspirin for future Phase IIa and IIb chemopreventive trials in high‐risk cohorts of patients for colon cancer. Aspirins effects on rectal prostaglandins are prolonged, detectable even after aspirin and its metabolite are removed from the plasma. Aspirin‐mediated inhibition of prostaglandin production in the human rectal epithelium may be related to direct suppression of cyclooxygenase transcription and not to enzyme inactivation by acetylation. A systematic method to monitor adherence (self‐report, telephone contact, pill count, and microelectronic monitoring) has been established for future trials. Strategies to improve recruitment of high‐risk cohorts have been developed. Phase IIa non‐randomized studies with aspirin at 81 mg in high‐risk cohorts (resected Dukes A colon cancer, Dukes C colon cancer treated with adjuvant therapy and disease‐free at 5 years, history of colon adenomas > 1 cm, two or more first‐degree relatives with colon cancer, and familial adenomatous polyposis and hereditary non‐polyposis colorectal cancer syndromes) are currently being conducted for surrogate end‐point biomarker (prostaglandins, cyclooxygenase, cellular mucins, and proliferation) modulation. J. Cell. Biochem. Suppls. 28/29:148–158. Published 1998 Wiley‐Liss, Inc. This article is a US Government work and, as such, is in the public domain in the United States of America.

CHEMOPREVENTION OF COLORECTAL CANCER

This review summarizes the principles of cancer chemoprevention and discusses the evidence from epidemiologic and experimental studies and preclinical and clinical trials of potential colorectal chemopreventive agents. The putative mechanisms of action of the drugs in chemoprevention and their potential to reduce the incidence and mortality rate of colorectal neoplasms are discussed. The future of colorectal chemoprevention will depend on important new insights into molecular carcinogenesis of colorectal cancer, application of molecular markers as surrogate endpoints, and ultimately on therapeutic targets of prevention in clinical trials.

Anti-Neoplastic Activity of Two Flavone Isomers Derived from Gnaphalium elegans and Achyrocline bogotensis

Over 4000 flavonoids have been identified so far and among these, many are known to have antitumor activities. The basis of the relationships between chemical structures, type and position of substituent groups and the effects these compounds exert specifically on cancer cells are not completely elucidated. Here we report the differential cytotoxic effects of two flavone isomers on human cancer cells from breast (MCF7, SK-BR-3), colon (Caco-2, HCT116), pancreas (MIA PaCa, Panc 28), and prostate (PC3, LNCaP) that vary in differentiation status and tumorigenic potential. These flavones are derived from plants of the family Asteraceae, genera Gnaphalium and Achyrocline reputed to have anti-cancer properties. Our studies indicate that 5,7-dihydroxy-3,6,8-trimethoxy-2-phenyl-4H-chromen-4-one (5,7-dihydroxy-3,6,8-trimethoxy flavone) displays potent activity against more differentiated carcinomas of the colon (Caco-2), and pancreas (Panc28), whereas 3,5-dihydroxy-6,7,8-trimethoxy-2-phenyl-4H-chromen-4-one (3,5-dihydroxy-6,7,8-trimethoxy flavone) cytototoxic action is observed on poorly differentiated carcinomas of the colon (HCT116), pancreas (Mia PaCa), and breast (SK-BR3). Both flavones induced cell death (>50%) as proven by MTT cell viability assay in these cancer cell lines, all of which are regarded as highly tumorigenic. At the concentrations studied (5–80 µM), neither flavone demonstrated activity against the less tumorigenic cell lines, breast cancer MCF-7 cells, androgen-responsive LNCaP human prostate cancer line, and androgen-unresponsive PC3 prostate cancer cells. 5,7-dihydroxy-3,6,8-trimethoxy-2-phenyl-4H-chromen-4-one (5,7-dihydroxy-3,6,8-trimethoxy flavone) displays activity against more differentiated carcinomas of the colon and pancreas, but minimal cytotoxicity on poorly differentiated carcinomas of these organs. On the contrary, 3,5-dihydroxy-6,7,8-trimethoxy-2-phenyl-4H-chromen-4-one (3,5-dihydroxy-6,7,8-trimethoxy flavone) is highly cytotoxic to poorly differentiated carcinomas of the colon, pancreas, and breast with minimal activity against more differentiated carcinomas of the same organs. These differential effects suggest activation of distinct apoptotic pathways. In conclusion, the specific chemical properties of these two flavone isomers dictate mechanistic properties which may be relevant when evaluating biological responses to flavones.

PROSTAGLANDIN INHIBITORS AND THE CHEMOPREVENTION OF NONCOLONIC MALIGNANCY

Much has been learned about the role of NSAIDs as cancer preventives through epidemiologic and experimental studies. The pathways of carcinogenesis in the gastrointestinal tract are initiated by many different genetic, environmental, infective, and lifestyle factors. It is possible that the final common pathway of all these malignancies may have some common features. It is conceivable that head and neck, esophageal, gastric, and colorectal epithelial carcinogenesis all are influenced by or require COX-2 up-regulation as a step toward transformation. Intuitively, it is possible that selective COX-2 inhibitors may have a preventive role in all these epithelial malignancies. Todays challenge is to translate this information into clinical trials to define what role, if any, COX inhibition might play in the prevention of these malignancies.