M. D. Mediavilla

Clinical uses of melatonin: evaluation of human trials.

During the last 20 years, numerous clinical trials have examined the therapeutic usefulness of melatonin in different fields of medicine. The objective of this article is to review, in depth, the science regarding clinical trials performed to date. The efficacy of melatonin has been assessed as a treatment of ocular diseases, blood diseases, gastrointestinal tract diseases, cardiovascular diseases, diabetes, rheumatoid arthritis, fibromyalgia, chronic fatigue syndrome, infectious diseases, neurological diseases, sleep disturbances, aging and depression. Melatonin has been also used as a complementary treatment in anaesthesia, hemodialysis, in vitro fertilization and neonatal care. The conclusion of the current review is that the use of melatonin as an adjuvant therapy seems to be well funded for macular degeneration, glaucoma, protection of the gastric mucosa, irritable bowel syndrome, arterial hypertension, diabetes, side effects of chemotherapy and radiation in cancer patients or hemodialysis in patients with renal insufficiency and, especially, for sleep disorders of circadian etiology (jet lag, delayed sleep phase syndrome, sleep deterioration associated with aging, etc.) as well as in those related with neurological degenerative diseases (Alzheimer, etc.,) or Smith-Magenis syndrome. The utility of melatonin in anesthetic procedures has been also confirmed. More clinical studies are required to clarify whether, as the preliminary data suggest, melatonin is useful for treatment of fibromyalgia, chronic fatigue syndrome, infectious diseases, neoplasias or neonatal care. Preliminary data regarding the utility of melatonin in the treatment of ulcerative colitis, Crohns disease, rheumatoid arthritis are either ambiguous or negative. Although in a few cases melatonin seems to aggravate some conditions, the vast majority of studies document the very low toxicity of melatonin over a wide range of doses.

Melatonin increases p53 and p21WAFl expression in MCF-7 human breast cancer cells in vitro

The aim of the present work was to study whether melatonin, at physiological concentrations, exerts its antiproliferative effects on MCF-7 human breast cancer cells by inducing the expression of some of the proteins involved in the control of the cell cycle. MCF-7 cells were cultured for 48 h in DMEM media containing either melatonin (1 nM) or the diluent (0.001% ethanol). At this concentration, after 48 hours of incubation, melatonin reduced the number of viable cells in relation to controls. The decreased cell proliferation was coincident with a significant increase in the expression of p53 as well as p21WAF1 proteins. These results demonstrate that melatonin inhibits MCF-7 cell proliferation by inducing an arrest of cell cycle dependent on an increased expression of p21WAF1 protein, which is mediated by the p53 pathway.

Basic mechanisms involved in the anti-cancer effects of melatonin.

It is commonly accepted that melatonin (N-acetyl-5-methoxytryptamine), the most relevant pineal secretory product, has oncostatic properties in a wide variety of tumors and, especially, in those identified as being hormonedependent. The objective of the present article is to offer a global and integrative view of the mechanisms involved in the oncostatic actions of this indoleamine. Due to the wide spectrum of melatonins actions, the mechanisms that may be involved in its ability to counteract tumor growth are varied. These include: a) antioxidant effects; b) regulation of the estrogen receptor expression and transactivation; c) modulation of the enzymes involved in the local synthesis of estrogens; d) modulation of cell cycle and induction of apoptosis; e) inhibition of telomerase activity; f) inhibition of metastasis; g) prevention of circadian disruption; h) antiangiogenesis; i) epigenetic effects; j) stimulation of cell differentiation; and k) activation of the immune system. The data supporting each of these oncostatic actions of melatonin are summarized in this review. Moreover, the list of actions described may not be exhaustive in terms of how melatonin modulates tumor growth.

Melatonin modulates aromatase activity in MCF-7 human breast cancer cells

Abstract:  Most of the current knowledge about the mechanisms by which melatonin inhibits the growth of breast cancer cells point to an interaction of melatonin with estrogen‐responsive pathways, thus behaving as an antiestrogenic hormone. However, a possible effect of melatonin on the local synthesis of estrogens had not been examined. The objective of this work was to study whether melatonin may modify the aromatase activity in MCF‐7 breast cancer cells thus modulating the local estrogen biosynthesis. In MCF‐7 cells cultured with testosterone in estradiol‐free media, melatonin (1 nm) counteracts the testosterone‐induced cell proliferation dependent on the local biosynthesis of estrogens from testosterone by the aromatase activity of the cells. We found that melatonin reduces the aromatase activity (measured by the tritiated water release assay) of MCF‐7 cells both at basal conditions and when aromatase activity was stimulated by cAMP or cortisol. The greatest inhibition of the aromatase activity was obtained with 1 nm melatonin, the same concentration that gives the highest antiproliferative and anti‐invasive effects of MCF‐7 cells. Finally, by RT‐PCR, we found that melatonin downregulates aromatase expression at the transcriptional level in the MCF‐7 cells. We conclude that melatonin, at physiological concentrations, decreases aromatase activity and expression in MCF‐7 cells. This aromatase inhibitory effect of melatonin, together with its already known antiestrogenic properties interacting with the estrogen‐receptor, makes this indoleamine an interesting tool to be considered in the prevention and treatment of hormone‐dependent mammary neoplasias.

Melatonin inhibits the growth of DMBA-induced mammary tumors by decreasing the local biosynthesis of estrogens through the modulation of aromatase activity

Melatonin inhibits the growth of breast cancer cells by interacting with estrogen‐responsive pathways, thus behaving as an antiestrogenic hormone. Recently, we described that melatonin reduces aromatase expression and activity in MCF‐7 human breast cancer cells, thus modulating the local estrogen biosynthesis. To investigate the in vivo aromatase‐inhibitory properties of melatonin in our current study, this indoleamine was administered to rats bearing DMBA‐induced mammary tumors, ovariectomized (ovx) and treated with testosterone. In these castrated animals, the growth of the estrogen‐sensitive mammary tumors depends on the local aromatization of testosterone to estrogens. Ovariectomy significantly reduced the size of the tumors while the administration of testosterone to ovx animals stimulated tumor growth, an effect that was suppressed by administration of melatonin or the aromatase inhibitor aminoglutethimide. Uterine weight of ovx rats, which depends on the local synthesis of estrogens, was increased by testosterone, except in those animals that were also treated with melatonin or aminoglutethimide. The growth‐stimulatory effects of testosterone on the uterus and tumors depend exclusively on locally formed estrogens, since no changes in serum estradiol were appreciated in testosterone‐treated rats. Tumors from animals treated with melatonin had lower microsomal aromatase activity than tumors of animals from other groups, and incubation with melatonin decreased the aromatase activity of microsomal fractions of tumors. Animals treated with melatonin had the same survival probability as the castrated animals and significantly higher survival probability than the uncastrated. We conclude that melatonin could exert its antitumoral effects on hormone‐dependent mammary tumors by inhibiting the aromatase activity of the tumoral tissue.

Does melatonin induce apoptosis in MCF-7 human breast cancer cells in vitro?

Melatonin inhibits proliferation of the estrogen‐responsive MCF‐7 human breast cancer cells. The objective of this work was to assess whether melatonin not only regulates MCF‐7 cell proliferation but also induces apoptosis. In this experiment we used 1,25‐dihydroxycholecalciferol (D3) as a positive control because it inhibits MCF‐7 cell proliferation and induces apoptosis. MCF‐7 cells were cultured with either 1 nM melatonin, 100 nM D3 or its diluent to determine their effects on cell proliferation, cell viability, cell‐cycle phase distribution, population of apoptotic cells, and expression of p53, p21WAF1, bcl‐2, bcl‐XL and bax proteins. After 24 or 48 hr of incubation, both melatonin and D3‐treatment significantly decreased the number of viable cells in relation to the controls, although no differences in cell viability were observed between the treatments. The incidence of apoptosis, measured as the population of cells falling in the sub‐G1 region of the DNA histogram, or by the TUNEL reaction, was similar in melatonin‐treated and control cells whereas, as expected, apoptosis was higher among cells treated with D3 than in controls. The expression of p53 and p21WAF1 proteins significantly increased after 24 or 48 hr of incubation with either melatonin or D3. No significant changes in bcl‐2, bcl‐XL and bax mRNAs were detected after treatment with melatonin whereas in D3‐treated cells, a significant drop in bcl‐XL was observed. These data support the hypothesis that melatonin reduces MCF‐7 cell proliferation by modulating cell‐cycle length through the control of the p53–p21 pathway, but without clearly inducing apoptosis.

Melatonin inhibits both ERα activation and breast cancer cell proliferation induced by a metalloestrogen, cadmium

Abstract:  Cadmium (Cd) is a heavy metal affecting human health both through environmental and occupational exposure. There is evidence that Cd accumulates in several organs and is carcinogenic to humans. In vivo, Cd mimics the effect of estrogens in the uterus and mammary gland. In estrogen‐responsive breast cancer cell lines, Cd stimulates proliferation and can also activate the estrogen receptor independent of estradiol. The ability of this metalloestrogen to increase gene expression in MCF7 cells is blocked by anti‐estrogens suggesting that the activity of these compounds is mediated by ERα. The aims of this work were to test whether melatonin inhibits Cd‐induced proliferation in MCF7 cells, and also to study whether melatonin specifically inhibits Cd‐induced ERα transactivation. We show that melatonin prevents the Cd‐induced growth of synchronized MCF7 breast cancer cells. In transient transfection experiments, we prove that both ERα‐ and ERβ‐mediated transcription are stimulated by Cd. Melatonin is a specific inhibitor of Cd‐induced ERα‐mediated transcription in both estrogen response elements (ERE)‐ and AP1‐containing promoters, whereas ERβ‐mediated transcription is not inhibited by the pineal indole. Moreover, the mutant ERα‐(K302G, K303G), unable to bind calmodulin, is activated by Cd but becomes insensitive to melatonin treatment. These results proved that melatonin inhibits MCF7 cell growth induced by Cd and abolishes the stimulatory effect of the heavy metal in cells expressing ERα at both ERE‐luc and AP1‐luc sites. We can infer from these experiments that melatonin regulates Cd‐induced transcription in both ERE‐ and AP1 pathways. These results also reinforce the hypothesis of the anti‐estrogenic properties of melatonin as a valuable tool in breast cancer therapies.

Melatonin inhibits aromatase promoter expression by regulating cyclooxygenases expression and activity in breast cancer cells

Background:Melatonin reduces the development of breast cancer interfering with oestrogen-signalling pathways, and also inhibits aromatase activity and expression. Our objective was to study the promoters through which melatonin modifies aromatase expression, evaluate the ability of melatonin to regulate cyclooxygenases and assess whether the effects of melatonin are related to its effects on intracellular cAMP, in MCF-7 cells.Methods:Total aromatase mRNA, aromatase mRNA promoter regions and cyclooxygenases mRNA expression were determined by real-time RT–PCR. PGE2 and cAMP were measured by kits.Results:Melatonin downregulated the gene expression of the two major specific aromatase promoter regions, pII and pI.3, and also that of the aromatase promoter region pI.4. Melatonin 1 nM was able to counteract the stimulatory effect of tetradecanoyl phorbol acetate on PGE2 production and inhibit COX-2 and COX-1 mRNA expression. Melatonin 1 nM elicited a parallel time-dependent decrease in both cyclic AMP formation and aromatase mRNA expression.Conclusions:This study shows that melatonin inhibits aromatase activity and expression by regulating the gene expression of specific aromatase promoter regions. A possible mechanism for these effects would be the regulation by melatonin of intracellular cAMP levels, mediated by an inhibition of cyclooxygenase activity and expression.

Melatonin as a Selective Estrogen Enzyme Modulator

Melatonin exerts oncostatic effects on different kinds of tumors, especially on hormone-dependent breast cancer. The general conclusion is that melatonin, in vivo, reduces the incidence and growth of chemically-induced mammary tumors in rodents, and, in vitro, inhibits the proliferation and invasiveness of human breast cancer cells. Both studies support the hypothesis that melatonin inhibits the growth of breast cancer by interacting with estrogen-signaling pathways through three different mechanisms: (a) the indirect neuroendocrine mechanism which includes the melatonin down-regulation of the hypothalamic-pituitary-reproductive axis and the consequent reduction of circulating levels of gonadal estrogens, (b) direct melatonin actions at tumor cell level by interacting with the activation of the estrogen receptor, thus behaving as a selective estrogen receptor modulator (SERM), and (c) the regulation of the enzymes involved in the biosynthesis of estrogens in peripheral tissues, thus behaving as a selective estrogen enzyme modulator (SEEM). As melatonin reduces the activity and expression of aromatase, sulfatase and 17beta-hydroxysteroid dehydrogenase and increases the activity and expression of estrogen sulfotransferase, it may protect mammary tissue from excessive estrogenic effects. Thus, a single molecule has both SERM and SEEM properties, one of the main objectives desired for the breast antitumoral drugs. Since the inhibition of enzymes involved in the biosynthesis of estrogens is currently one of the first therapeutic strategies used against the growth of breast cancer, melatonin modulation of different enzymes involved in the synthesis of steroid hormones makes, collectively, this indolamine an interesting anticancer drug in the prevention and treatment of estrogen-dependent mammary tumors.

Inhibitory effects of pharmacological doses of melatonin on aromatase activity and expression in rat glioma cells

Melatonin exerts oncostatic effects on different kinds of neoplasias, especially on oestrogen-dependent tumours. Recently, it has been described that melatonin, on the basis of its antioxidant properties, inhibits the growth of glioma cells. Glioma cells express oestrogen receptors and have the ability to synthesise oestrogens from androgens. In the present study, we demonstrate that pharmacological concentrations of melatonin decreases the growth of C6 glioma cells and reduces the local biosynthesis of oestrogens, through the inhibition of aromatase, the enzyme that catalyses the conversion of androgens into oestrogens. These results are supported by three types of evidence. Firstly, melatonin counteracts the growth stimulatory effects of testosterone on glioma cells, which is dependent on the local synthesis of oestrogens from testosterone. Secondly, we found that melatonin reduces the aromatase activity of C6 cells, measured by the tritiated water release assay. Finally, by (RT)–PCR, we found that melatonin downregulates aromatase mRNA steady-state levels in these glioma cells. We conclude that melatonin inhibits the local production of oestrogens decreasing aromatase activity and expression. By analogy to the implications of aromatase in other forms of oestrogen-sensitive tumours, it is conceivable that the modulation of the aromatase by pharmacological melatonin may play a role in the growth of glioblastomas.