New paradigms in chronic intestinal inflammation and colon cancer: role of melatonin

Abstract

Abstract:  In intestinal bowel disease (IBD), immune-mediated conditions exert their effects through various cells and proinflammatory mediators. Recent data support a participation of the endoplasmic reticulum stress and mitochondrial dysfunctions in IBD. Moreover, it is evident that chronic degenerative pathologies, including IBD, share comparable disease mechanisms with alteration in the autophagy mechanisms. Chronic inflammation in IBD exposes these patients to a number of signals known to have tumorigenic effects. This circuitry of inflammation and cancer modifies apoptosis and autophagy, and promotes cellular cycle progression, invasion, and angiogenesis. Melatonin has been shown as a specific antioxidant reducing oxidative damage in both lipid and aqueous cell environments. However, several studies provide further insight into the molecular mechanisms of melatonin action in the colon. In this line, recent data suggest that melatonin modulates autophagy and sirtuin activity. An anti-autophagic property of melatonin has been demonstrated, and it could contribute to its anti-oncogenic activity. Nevertheless, there is no information about whether antitumoral effects of melatonin on colon cancer are dependent on autophagy. Sirtuins have pleiotropic effects on cancer development, being reported both as facilitator and as suppressor of colon cancer development. Sirtuins and melatonin are connected through the circadian clock machinery, and melatonin seems able to correct the alterations in sirtuin activity associated with several pathological conditions. Autophagy and sirtuin activities are linked through 5′AMP-activated protein kinase (AMPK) activation, which switches on autophagy and increases sirtuin. The effect of melatonin on AMPK and the impact of this effect on IBD and colon cancer remain an open question.

Molecular mechanisms in inflammatory bowel disease

The incidence of inflammatory bowel disease (IBD) continues to rise, including high-incidence areas (western countries), although both incidence and prevalence are also increasing in historically low-incidence areas such as Latin America, India, and Asia. Factors associated with this ‘westernization’ may be conditioning the expression of these pathologies, and the increase in the incidence among migrants from low- to high-incidence regions in just one generation suggests a strong environmental influence [1].

The hallmark of IBD, including ulcerative colitis (UC) and Crohn′s disease (CD), is chronic, uncontrolled inflammation of the intestinal mucus, which can affect any part of the gastrointestinal (GI) tract with the induction of structural alterations and superficial or transmural granulomatous infiltration [2]. IBD is associated with increased permeability of the epithelial lining of the intestine resulting in continuous stimulation of the mucosal immune system. Luminal bacteria appear to intensify the permeability defect further, establishing a self-sustaining cycle of mucosal inflammation. Intestinal epithelial cells have developed control mechanisms that organize the activation of the intestinal immune system. However, under pathological conditions, bacterial products cross the mucosal barrier and enter the mucus generating a classic immune response [2, 3].

The traditional paradigm for IBD pathogenesis was that cells from the adaptive immune system are the mediators of intestinal inflammation. However, now the participation of the innate immune system in IBD is accepted [4–6]. In this sense, the intestinal epithelium is believed to contribute to innate immunity and to the relative sterility of the mucosal surface, playing an active role in the maintenance of the mucosal immune homeostasis [7]. Likewise, the intestinal epithelium appears to act as a ‘gatekeeper’ that regulates the quality (type) and quantity (number) of leukocytes migrating from the intravascular compartment to the interstitial space (Fig. 1) [8]. This is a complex process mediated by cytokines, chemokines, and adhesion molecules. After exposure to abundant intestinal bacterial antigens or environmental factors, innate immune cells in the intestinal mucus are activated, leading to the overproduction of proinflammatory cytokines. Recently, the IL-23/IL-12 pathway has become the subject of intensive study, and the T-helper type 1 (Th1) cells, driven by IL-12, and interleukin (IL)-17-producing T-helper (Th17) cells, driven by IL-23, have been demonstrated to play an important role in IBD [9]. The Th17 pathway genes are shared between CD and UC, while others are IBD subtype-specific including autophagy genes, or epithelial barrier genes [10, 11] have demonstrated that the IL-23 receptor is vital for the maintenance of many types of CD-T cells that provide early adaptive immune responses to damage. These IL-17-producing effector T cells are crucial for protection against intercellular pathogens and for organ-specific inflammation; the therapeutic disruption of the IL-23 pathway suggests the control of auto-immune inflammation without impairing systemic immunity [12].

Figure 1.  Proposed inflammatory mechanisms in intestinal bowel disease (IBD). After exposure to the abundant intestinal bacterial antigens or environmental factors, innate immune cells are activated, leading to the overproduction of proinflammatory cytokines. The IL-23/IL-12 pathway has become the subject of intensive study and T-helper type 1 (Th1) cells, driven by IL-12, and interleukin (IL)-17-producing T-helper (Th17) cells, driven by IL-23, have been demonstrated to play an important role in IBD. Microbial imbalance, or dysbiosis, contributes to disease via Toll-like receptors (TLRs: TLR2 and TLR4) that have demonstrated their capacity to recognize a vast array of microbial components. NOD1-like receptors (NLRs), like TLRs, are also strongly linked to the development of chronic intestinal inflammation. NOD2 and TLRs stimulation induces autophagy in mononuclear phagocyte system (MPS), including dendritic cells. Paneth cells, at the base of the intestinal crypts, express the intracellular receptor NOD2, whose mutations alter the production of antimicrobial peptides and TNF-α, and induce autophagy. Nuclear factor kappa B (NFκB) is inside a complex network that regulates cellular pathways involved in the expression of a variety of genes that play critical roles in immune responses. The result is the augmentation of proinflammatory molecules (various cytokines, chemokines, reactive oxygen and nitrogen species, ROS and RNS respectively, and prostaglandins), which are primarily secreted by MPS cells and also epithelial cells, affecting innate and acquired immune response. In addition to NFκB activation, commensal bacteria dampen inflammation via peroxisome proliferator-activated receptor (PPAR-γ). The role of PPAR-γ in the immune response is through its ability to down-modulate the expression of inflammatory cytokines and to direct immune cell differentiation toward anti-inflammatory phenotypes.

Many receptors are implicated in the immunopathogenesis of IBD, and those located on the intestinal mucosal surface constitute an immunological barrier that is in continuous intimate contact with a variety of gut microflora commensals bacteria. Structurally distinct families of pattern recognition receptors (PRRs) are pivotal to the control of intestinal mucosal homeostasis [13]. Toll-like receptors (TLRs) and nucleotide-binding oligomerization domain-1 (NOD-1)-like receptors, both interconnected and coordinated through many signals pathways, provide an integrated system to recognize microbes and microbial molecules and to control antimicrobial effector pathways and adaptive immune responses.

In health, normal PRR signaling protects intestinal barrier integrity and confers commensal tolerance; in this line, different studies suggest that this kind of signaling exerts several important cytoprotective responses in the intestine epithelium, including barrier preservation, inhibition of apoptosis and inflammation, wound repair and regeneration, and autophagy control [13]. Autophagy, traditionally considered as simple as a degradation mechanism, is now believed to have numerous functions and to be able to play complex roles in human diseases [14] including IBD. The progress made by endeavors such as the human genome project and genotyping technologies has made the identification of susceptibility loci for IBD that are shared between UC and CD, but also a number of loci that are disease-specific [15]. Association between two genes, the autophagy-related 16-like 1 gene, ATG16L1 and IRGM, and CD was identified by a genome study and by a wide single-nucleotide polymorphism (SNP) study [16, 17].

Aberrant PRR signaling, when it occurs in disease, leads to deleterious tissue injury associated with chronic inflammatory and autoimmune responses. Several studies have reported that expression of TLR-4 is low in the normal colonic mucus and upregulated in UC [18], which suggests the possibility that abnormal bacterial sensing, microbial imbalance or dysbiosis contribute to disease pathogenesis. Along these lines, TLR-2, which belongs to the same TLR membrane surface receptors, recognizes a vast array of microbial components and is involved in different models of IBD [19]. However, TLRs do not discriminate between pathogenic and nonpathogenic microorganisms (commensal), which is important for understanding innate signaling [20]. Concerning NOD-1-like receptors (NLRs), they are expressed not only by DCs [21] but also by Paneth cells and macrophages [22]. NLRs, like TLRs, do not differentiate between pathogenic microorganisms and commensal flora [23]. The first innate receptor strongly linked to the development of chronic intestinal inflammation in a subset of patients with CD is the NOD2/CARD15 [24, 25]. NOD2 mutations are associated with an increased risk of CD, which suggests that the deregulated recognition of intestinal microbes leads to disease in a genetically predisposed individual. Recently, two independent groups have linked NOD2 and autophagy [26, 27]. NOD2 stimulation induces autophagy in dendritic cells (Fig. 1) and requires ATG5, ATG7, and ATG16L1. NOD2-mediated autophagy affects bacterial handling and antigen presentation in dendritic cells. Mutant NOD2 is retained in the cytosol and, therefore, fails to bring Atg16L1 to the plasma membrane, impairing autophagy targeting of bacteria. Autophagy is a key process in host resistence to bacterial infection, although little is known about the steps by which pathogens manipulate the cell to evade the autophagy pathway. The connection between autophagy and IBD development can exist on multiple levels, including intestinal homeostasis, bacterial clearance, cytokine production, and Paneth cell functions [15], whereas some genetic alterations in autophagy promote the development of IBD. In addition, other processes related to IBD pathology (response to pathogens, oxidative damage, and other stresses) would be presumably altered by malfunction of the autophagic process.

When contact is made between microbial components and both NOD2 and TLR, the nuclear factor kappa B (NFκB) signaling pathway stimulates the expression of multiple molecules relevant to the pathogenesis of various diseases including IBD and colorectal cancer [28, 29]. NFκB signaling pathway is a complex network that regulates a cellular pathway involved in the expression of a wide variety of genes that play critical roles in immune responses [30]. NFκB is regulated by the IκB family, with seven IκB members including IκBα, IκBβ, IκBγ, IκBε, Bcl-3, and the precursor proteins p100 and p105. Briefly, following stimulation with various inflammatory stimuli, such as certain members of the TNF-α cytokine family, IL-1, TLR ligands, and the p50 subunit mostly, translocates to the nucleus and activates the transcription of various target genes [31]. The result is the augmentation of proinflammatory molecule production during active IBD, including those encoding cytokines such as IL-1, IL-2, IL-6, IL-12, or TNF-α [32]. These cytokines are primarily secreted by monocytes and macrophages upon activation and induce intestinal macrophages, neutrophils, fibroblasts, and smooth muscle cells to elaborate prostanoids, proteases, and many other mediators of inflammatory tissue responses, and to promote the production of other chemotactic cytokines affecting innate as well as the acquired immune response at mucosal sites [33].

In addition to NFκB activation, commensal bacteria dampen inflammation via nucleocytoplasmic redistribution of peroxisome proliferator-activated receptor (PPAR)-γ, a member of the nuclear receptor group of transcription factors. PPAR-γ is highly expressed in the intestinal epithelium, immune cells, and adipocytes, and regulates a number of genes participating in metabolism, proliferation, signal transduction, and cellular motility. The role of PPAR-γ in the immune response is through its ability to down-modulate the expression of inflammatory cytokines and to direct immune cell differentiation toward anti-inflammatory phenotypes [34]. A recent animal study by Guri et al. [35] investigated the underlying mechanisms by which the deletion of PPAR-γ in intestinal epithelial cells modulates the severity of experimental IBD, immune cell distribution, and global gene expression. These authors observed that PPAR-γ expression is required for preventing colonic inflammatory lesions, upregulating lysosomal pathway genes and increasing the production of the anti-inflammatory cytokine IL-10. In a different experimental model of IBD, activation of PPAR-γ by different agonists suppresses gut inflammatory lesions, weight loss, and inflammatory mediator expression [36, 37]. Most notably, the PPAR-γ agonist rosiglitazone showed therapeutic efficacy in humans with UC, although this molecule and other drugs belonging to the thiazolidinedione class of antidiabetic drugs are unlikely to be adopted for the treatment of IBD because of their significant side effects [38].

Recent data support the participation of the endoplasmic reticulum (ER) stress in IBD. Moreover, it is now evident that chronic degenerative disorders, such as type 2 diabetes or IBD, share comparable disease mechanisms at cellular level, including ER and mitochondrial dysfunction, with inflammatory processes as a key disease-conditioning situation in different tissues [39, 40]. Stress in mitochondria, or the ER independently, causes cell death. However, it has been recently reported that ER stress causes mitochondrial dysfunction via p53-upregulated modulator of apoptosis (PUMA) and tumor necrosis factor receptor-associated protein 1 (TRAP1), located in the mitochondria and associated with the unfolded protein response (UPR) in the ER [41].

Genetic and environmental factors can affect ER stress in the intestinal epithelium and consequently inflammation [42]; genetic factors include either primary or secondary ER stress and environmental factors include bacteria, diet, or drugs. Evidence supports the hypothesis that the ER and the mitochondrial share common mechanisms in triggering the unfolded or misfolded protein response (UPR). Moreover, a protective signaling pathway from the ER to the nucleus controls cell stress response caused by unfolded and/or misfolded proteins. Accumulation of UPR or aggregated proteins in the endoplasmic reticulum results in increased chaperone expression, translational arrest, and induction of autophagy (Fig. 1). UPR signaling is mainly driven by inositol-requiring endoplasmic reticulum-to-nucleus signaling protein 1α (IRE1α), X-box-binding-1 (XBP1) pathway (IRE-XBP1 pathway) [43]. IREα is a transmembrane kinase/endoribonuclease, which initiates the nonconventional splicing of the messenger RNA encoding a key transcription activator XBP1. XBP1 is a key component of the ER stress response and is required for the differentiation and function of certain secretory epithelial cells [44, 45]. IREα is ubiquitous, whereas IRE1β is specifically expressed in the intestinal epithelium. IRE1α exhibits both endoribonuclease, with XBP1 being the only known substrate, and kinase activities that engage both JNK and classical NF-kB pathways. XBP1 deletion causes ER stress in the epithelium, enteritis, increased susceptibility to DSS colitis, lacks Paneth cells in the intestinal epithelia, and decreases crypt bactericidal function, among others.

The full understanding of the different immunological mechanisms implicated in the development and perpetuation of the disease is very important as therapeutic interventions are subject to these mechanisms. In that respect, IBD treatment must be customized for each specific group or subgroup of patients.

Intestinal cancer as a consequence of a chronic inflammatory disorder

Morphometric alterations

Because Hinton [46] reported in 1966 that patients with ulcerous colitis were at greater risk of developing colorectal cancer (CC), many papers have reported that the length of the disease, its extent and association with other inflammatory diseases such as sclerosant colangitis, and anti-inflammatory treatment are factors that concede inflammation an advantageous role, at least, in carcinogenesis [47, 48]. Intestinal neoplasias, which originate under inflammatory conditions, have been the object of numerous clinical, anatomopathological, genetic, and molecular studies in both humans and animals. The association between UC and elevated risk for colorectal cancer is clear; however, there has been some debate about whether CD possesses a similar risk [49]. Earlier studies did not find a significant increase in the risk, although several other studies support the association between CD and cancer. Therefore, although most of the knowledge about colon cancer from chronic intestinal conditions comes from UC evidence, the data suggest that both of these conditions of the intestine confer increased intestinal cancer risk.

Experimental models in animals have been tested, but only a few are applicable in the study of the inflammation-dysplasia-cancer sequence [50, 51]. The histological changes observed in patients with IBD who develop neoplasm correspond to the inflammation-dysplasia-cancer sequence. In this progression, all works base the histological classification on the data established by Riddle et al. [52] and Pascal [53]: (i) undefined for dysplasia/probably negative, (ii) undefined for dysplasia/probably positive, (iii) low-grade dysplasia, (iv) high-grade dysplasia, and (v) carcinoma. However, the identification of dysplasia in intestinal inflammatory diseases represents a huge challenge for both clinicians and pathologists, so a clear diagnosis of dysplasia in IBD is not always possible. Potential markers, such as p53 and alpha-methylacyl coenzyme, have been used. The combination of these two markers is positive in 75.8% for cancers and 30.3% for undefined biopsies for dysplasia, while only in 0.6% for non-neoplastic epithelium.

The suspect lesions can be visible macroscopically or only microscopically. Bird and Good [54], in 1987, described focal points of aberrant crypts (AC) such as preneoplastic lesions in rodents treated with a carcinogen. Now, it has been proven that these AC can be high or flat lesions. ‘High AC’ are characterized by being microscopically large and being raised above the surrounding epithelium with round, elongated, open lights. ‘Flat AC’ are characterized by lesions not vising above the epithelium, showing a brilliant methylene blue and being small or slightly enlarged as well as compressed open lights. Their detection is the result of the combination of methylene blue staining and transillumination. β-catenin and cycline D1 expression has been studied in both high and flat ACs; while β-catenin is found in the cytoplasm in flat lesions, it moves to the cytoplasm and nucleus in polypoidal lesions. Thus, inflammation seems to play an important role in the dysplasia-cancer sequence both in flat lesions and in mass, while the translocation of β-catenin to the cytoplasm and nucleus is an early event that occurs in polypoidal lesions (Fig. 2) [55].

Figure 2.  Inflammation and progression of colon cancer. The histological changes observed in intestinal bowel disease (IBD) that develops neoplasm correspond to the inflammation-dysplasia-cancer sequence. Chronic inflammation exposes to a number of alterations known to have procarcinogenic effects. Carcinogenesis, divided into three phases (initiation, promotion, and progression), is influenced by nuclear factors and inflammatory regulators: activation of nuclear factor kappa B (NFκB) signaling pathway, decrease in peroxisome proliferator-activated receptor (PPAR-γ) response, production of reactive oxygen species (ROS), and cyclooxygenase-2 protein expression (COX-2), and prostaglandin E₂ production (PGE₂). The stimulation of NFκB signaling pathway induces the expression of a number of genes: the angiogenic factor named vascular endothelial growth factor (VEGF), the anti-apototic factor Bcl-2, or and the proliferation factor cyclin D1. Inflammatory regulators also act also in a feedback loop to modify regulative systems, including AMP-activated protein kinase (AMPK) and sirtuin deacetylase enzyme (SIRT1)-1, and to weaken p53 activity. Finally, the vicious circuitry of inflammation and cancer modifies apoptosis and autophagy and promotes cell cycle progression, invasion, and angiogenesis. Endoplasmic reticulum (ER) stress and mitochondrial dysfunctions in epithelial cells are also disease-conditioning situations in IBD. The UPR/IRE1α/XBP1 pathway results in induction of autophagy.

Molecular pathways to connect inflammation and colorectal cancer

At present, reasons for the elevated cancer risk in patients with IBD are not clear. Genetic predisposition and data from familial intestinal cancers could indicate their connections, although it is not clear which heritable genetic factors contribute to elevated colorectal cancer risk in IBD.

Several lines of evidence implicate chronic intestinal inflammation as a key predisposing factor in the modulation of tumorigenesis. The relationship and mechanisms through which infection and inflammation increase cancer risk and promote tumor development have been recently the object of extensive discussion [49, 56]. Chronic inflammation and repeated events of inflammatory relapse in IBD expose a number of alterations known to have procarcinogenic effects (Fig. 2). Carcinogenesis, divided into three phases (initiation, promotion, and progression), is influenced by different inflammatory mediators.

The contribution of the inducible isoform of cyclooxygenase-2 (COX-2) to carcinogenesis is well established. COX-2 can activate procarcinogens and indirectly increase free-radical production. The relationship between ROS and COX-2 expression has been addressed in a recent study which demonstrated that the chemopreventive sulindac and nitric oxide-donating aspirin (NO-ASA) generates ROS to induce COX-2 expression, which can be abolished by two antioxidants and an inhibitor of NADPH oxidase [57]. Recent data implicate COX-2 production down stream of TLR4 activation in the development of inflammation-associated colorectal neoplasia [49, 58]. Activation of TLR4 initiates a signaling cascade that culminates in NF-kB activation and subsequent transcription of a number of proinflammatory mediators, including COX-2. Whether NF-kB is required for ROS-induced COX-2 expression, however, needs further elucidation. NSAID may inhibit the development of gastrointestinal cancers through inhibition of COX-2. Nevertheless, COX-2-independent mechanisms also contribute significantly to the protective effects of NSAIDs. In any case, the available data reinforce the role of COX-2 in chronic inflammation and the development of CC [49].

There is growing evidence of a connection between inflammation, NF-kB and tumor development. Viral oncogenes, hepatitis B and C proteins, and human papillomavirus infection have been implicated in NF-kB activation. Besides, some chemical and physical carcinogens, especially nicotine and carcinogens in tobacco, promote cell proliferation, survival, and inflammation via NF-kB activation [59]. The role of NF-kB in promoting carcinogenesis is evidenced by numerous studies which indicate that this factor blocks apoptosis by regulating anti-apoptotic proteins, including IAPs, or by inhibiting JNK activation and the accumulation of ROS [60]. In chronic inflammation, the cytokines and chemokines produced by inflammatory cells activate NF-kB, which translocates into the nucleus, inducing the expression of certain tumorigenic, adhesion proteins, chemokines, and inhibitors of apoptosis that promote cell survival. Therefore, NF-kB may contribute to the development of colitis-associated CC by sustaining the ongoing inflammatory process in the gut mucosa. NF-kB is also connected to the regulation of many genes differently expressed in invasion and metastasis: cyclin D1 and cMyc oncogenes, and VEGF and IL-8 are directly or indirectly enhanced by NF-kB activation [61]. Several products have been suggested to inhibit NF-kB activation, including curcumin, ginseng extract, resveratrol, green tea extract, among others, and are known by their antiproliferative properties [62]. At present, there is significant enthusiasm for in vitro and animal studies for the use of specific NF-kB inhibitors as new anticancer therapy [63, 64]. However, it is important to introduce new drugs that inhibit NF-kB activation without any apparent effects on other signaling pathways or immunological effects [32].

In colon tumor tissue, expression of PPAR-γ has also been detected in several studies using clinical samples [65]. Activation of PPAR-γ leads to cell differentiation and apoptosis. In addition, PPAR-γ ligands have been shown to be potent inhibitors of angiogenesis, a process necessary for tumor growth and metastasis, and to protect against cellular transformation. A recent study on colon carcinogenesis induced by dimethylhydrazine reported a decrease in PPAR-γ expression after tumor induction and that the diclofenac chemopreventive action may be through PPAR-γ, regulation of COX-2, and the subsequent initiation of apoptosis [66]. There are interesting data from the study conducted in the laboratory of Bassaganya and Hontecillas [67] who have recently observed the beneficial effects of dietary n-3 polyunsaturated fatty acids (PUFAs) in experimemtal IBD and inflammation-induced CC through, at least in part, PPARgamma-dependent mechanism. Further studies are needed to fully elucidate the antiproliferative and prodifferentiation mechanisms of PPAR-γ activators and their expedient evaluation in the clinical management of CC.

A leading theory is that oxidative stress accompanying chronic inflammation contributes to neoplastic transformation. ROS/RNS, including the superoxide anion, the nitric oxide radical (NO˙), and its metabolite peroxynitrite anion, can interact with DNA in proliferating epithelium resulting in permanent genomic alterations. In addition, in colitis-associated colon carcinogenesis, ROS/RNS may contribute to the p53 mutations and can functionally impair the protein components of the DNA mismatch repair system [14, 68]. iNOS expression is induced during inflammation and catalyzes the production of nitric oxide (NO). Moreover, depending on the concentration, genetic background, and NO enzyme involved, NO may induce protective effects [69]. Clinical data show that iNOS levels are elevated in actively inflamed mucosa from IBD; however, there is controversy about its role in intestinal carcinogenesis [51, 70].

Autophagy and colorectal cancer

Autophagy plays a dual role in tumorigenesis: the promotion of cell death as a tumor suppressor and the prevention of cell death as an oncogenic mechanism [14]. Mitochondrial DNA is more susceptible to damage because of the lack of repair systems and histone protein protection, and it is thought that autophagy removes damaged mitochondria, thus alleviating oxidative stress to prevent carcinogenesis. However, autophagy is also a cytoprotective mechanism that prevents cells from death under starvation or stress conditions. Studies have shown that ROS can induce autophagy, which instead of causing cell death, protects cells from apoptosis or necrosis and suggests that autophagy plays both promotion and suppression roles in tumorigenesis [71].

The coordinated regulation of autophagy and apoptosis is essential for cells to make a dynamic choice between death and survival when under stress. These processes are not only regulated by common factors, such as p53, but they also share some key autophagy genes including Beclin-1 and Atg5 [72]. Beclin-1 is inhibited by the anti-apoptotic Bcl-2 family and plays a key role in convergence between autophagy and apoptosis [73]. Enforced expression of Atg5 sensitizes cells to apoptosis, and a calpain-mediated cleavage of Atg5 switches autophagy to apoptosis [74]. The tumor-suppressor protein p53 is altered in more than 50% of human cancers, and mutation of the p53 gene is one of the most common genetic changes in the development of human colorectal cancer. Recently, this protein has been shown to regulate autophagy in a dual fashion [75, 76]. Nuclear p53 stimulates autophagy and sustains the challenge of cells to deal with stress. Meanwhile, cytoplasmic p53 inhibit autophagy by managing the AMP-activated protein kinase (AMPK), a positive regulator of autophagy, and activates mTOR, the main negative regulator of autophagy [77]. In HCT-116 human colorectal cancer cells, loss of p53 impairs autophagic flux upon starvation, culminating in apoptosis [78]. This could explain why some cancer cells retain p53. Although the interplay between autophagy and p53 is complex, the understanding of it would provide new strategies to deal with cancer.

Experimental evidence suggests that autophagy is activated by tumor cells as a prosurvival mechanism against cytotoxic agents. Thus, inhibition of autophagy can be used as a tumor cell sensitizing strategy. Several colorectal cancer cells or cell lines (HT-29, HTC-116, DLD-1, SW480, WiDr, LoVo, SW620), treated with pharmacological inhibitors of autophagy or subjected to siRNA-mediated downregulation, show increased sensibility to cyclooxygenase inhibition [79], TRAIL-induced cell death [80], amino acid and glucose deprivation [81], sulphoraphane-induced apoptosis [82], and 5-fluorouracil chemotherapy [83].

Sirtuins, inflammatory bowel disease, and colorectal cancer

Sirtuins are a group of highly phylogenetically conserved proteins that occur in organisms from bacteria to human beings, and which catalyze the deacetylation of target proteins. The deacetylation reaction spends NAD⁺, a key molecule in energy metabolism, thus linking protein regulatory control to metabolic conditions [84, 85]. SIRT1 deacetylation of p53 causes a weakening of its apoptotic effect. As a consequence, SIRT1 might be considered a facilitator for cancer development. Nevertheless, although pro-oncogenic effects of SIRT1 have been reported in a number of studies, there are also reports showing a tumor-suppressor role for this protein as well [86]. Additionally, sirtuins have been implicated in circadian rhythms by deacetilating proteins in the clock mechanism of circadian control [87]. As discussed in the next paragraph, this establishes a link between sirtuins and melatonin, and between sirtuins and cancer [88]. In this respect, sleep disruption has been associated with IBD [89].

Although information about the role of sirtuins in IBD is limited, there are several reports that show an anti-inflammatory effect for these molecules. Resveratrol, the best-known SIRT1 activator, reverses colitis-associated decrease in SIRT-1 gene expression, activation of NF-ΚB, increase in COX-2 expression, and other changes, in a dextran sulfate sodium-induced colitis, and in a spontaneous IL-10^−/− mouse model of colitis [90, 91]. The same authors show that resveratrol suppressed colon cancer associated with colitis [92]. In addition, SIRT1 is a negative regulator of NFkB activity through the deacetylation of the p65 lysine 310 [93].

With respect to colorectal cancer, several studies support the notion that SIRT1 could be involved in carcinogenesis [88, 94], and SIRT1 has been found to be upregulated in various human cancers, including colon cancer [95]. SIRT1 expression is associated with microsatellite instability and CpG island methylator phenotype in human colorectal cancer [96]. Conversely, there are also studies that indicate that SIRT1 can act as tumor suppressor. SIRT1 suppresses intestinal tumorigenesis and colon cancer growth in a β-catenin-driven mouse model of colon cancer [97]. Abnormal levels of β-catenin may contribute to neoplastic transformation in colon cells [98], and the stability of this protein increases by acetylation [99]. SIRT1 deacetylates β-catenin and promotes cytoplasmic localization of the otherwise oncogenic form of β-catenin [97]. On the other hand, defects in the Wnt signaling, which is upstream of β-catenin, have also been associated with human cancers [100]. SIRT-1 has been shown to regulate Wnt signaling in four colon cancers cell lines (HT-29, HCT116, RKO, and DLD-1) [101]; in this study, SIRT1 promotes constitutive Wnt signaling and Wnt-induced cell migration, therefore having more of a protumor action than an antitumor effect. In another study, SIRT1 inhibited proliferation of the colon cancer cell line HCT116, and SIRT1 inhibition promoted colon tumor formation in a tumor xenograft assay [102]. These results show that sirtuins have pleiotropic effects on cancer development. Thus, there is a wide therapeutic potential for both activators and inhibitors of sirtuins.

Caloric restriction (CR) is the only factor extends maximum life span in diverse species [103], including primates [104]. Studies on yeast, worms, flies, and mice point to a role for nutrient-responsive molecules, such as SIRT1 and mTOR, in aging and CR [105]. SIRT1 is proposed to mediate the health benefits of CR, and it has shown its capacity to ameliorate degenerative diseases associated with aging [106]. Recently, Firestein et al. [97] demonstrated that CR induces an increase in SIRT1 expression in the intestine of rodents. Moreover, this study showed that the ectopic SIRT1 induction in a mouse model of colon cancer significantly reduced tumor formation proliferation and animal morbidity in the absence of CR. An autophagy relationship with sirtuins has been demonstrated and includes AMPK, which senses the intracellular AMP/ATP ratio and inhibits mTOR-dependent signaling [107] while augmenting cellular NAD⁺ levels [108]. Consequently, AMPK activation switches on autophagy and increases sirtuin activity. In addition, SIRT1 regulates autophagy by deacetylation of several components of the autophagic cascade [109], and the life span prolonging the effect of sirtuins has been proposed to be mediated by autophagy [110]. To date, there are no studies connecting autophagy–sirtuin activity and colon cancer.

The identification of these signals has led to a greater mechanistic understanding of IBD pathogenesis and its evolution to colon cancer, and points to potentially new therapeutic targets.

Responses associated with melatonin in IBD and colon cancer

Melatonin is an indoleamine produced, among other tissues, in the gastrointestinal tract where it plays an important physiological regulatory role [111]. Previous data from in vitro studies, animal experiments, and limited studies in humans suggest that melatonin, because of its antioxidative [112–114] and anti-inflammatory [115–117] properties, may be useful to prevent or treat pathological conditions such as esophageal and gastric ulcers, pancreatitis, colitis, irritable bowel disease, and some types of cancer [118, 119]. However, other studies regarding the utility of this hormone in the treatment of these immune-active pathologies are either ambiguous or negatives (Fig. 3) [120].

Figure 3.  Proposed mechanisms implied in the anti-inflammatory and anti-oxidative effects of melatonin. The anti-inflammatory actions of melatonin have been attributed to the attenuation of nuclear factor kappa B (NF-kB) activation, inhibition of cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS) expression and the subsequent production of prostaglandin E2 (PGE₂) and nitric oxide (NO), suppression of proinflammatory cytokines and adhesion molecules, and reduction in matrix metalloproteinase-2 and -9 (MMP-2 and MMP-9) activity and expression. In addition, melatonin exerts its antioxidant properties through both direct and indirect effects: the direct actions include scavenging of free radicals, such as superoxide anion () and peroxynitrite (ONOO⁻), and while the indirect properties include the stimulation of the activity of antioxidant enzymes, superoxide dismutase (SOD), and glutathione (GSH).

In vitro and animal IBD studies

Melatonin is a powerful antioxidant because of its amphiphilic features that allow it to cross physiological barriers, thereby reducing oxidative damage in both lipid and aqueous cell environments [121, 122]. It protects membrane lipids from peroxidation by scavenging peroxyl and hydroxyl radicals, superoxide anion, and peroxynitrite. Thus, this molecule is thought to play a protective role in the initial and advanced stages of diseases whose pathogenesis involves damage by reactive oxygen metabolites, including IBD [114, 123–125]. Melatonin’s action as a scavenger of free radicals has been implicated in the ameliorative effect of experimentally induced colitis and ulcers [126, 127]. Pentney and Bubenik [126] showed for the first time that melatonin reduced the severity of DSS-induced colitis in mice because of an improved microcirculation, stimulation of intestinal epithelium, and its capability as an antioxidant and scavenger of free radicals. Along the same lines, instillation of melatonin in animals with colitis decreased lipid peroxidation products, confirming that this molecule reduces colon mucosal oxidative injury [128–133]. Cuzzocrea et al. [128] demonstrated that melatonin reduced the appearance of nitrotyrosine, a specific marker of nitrosative stress, in colonic mucosa from rats subjected to experimental colitis by dinitrobenzene sulfonic acid (DNBS). These authors, based on previous in vitro studies [133], reported that this effect was likely related to a direct scavenging effect of melatonin on peroxynitrite (Fig. 3).

Besides its direct free-radical-scavenging actions, melatonin has shown to have indirect antioxidant properties by regulating the activity of antioxidant enzymes, including superoxide dismutase (SOD) and glutathione peroxidase (GPx) [134–136], which protect against free-radical damage (Fig. 3.). Several papers have reported a reduction in the extent of colonic damage induced by TNBS or acetic acid after melatonin administration, due in part to the preservation of the endogenous antioxidant reserve by the indoleamine [130, 132, 137].

In addition to its direct and indirect antioxidant properties, melatonin regulates the extensive intestine immune system, showing important general anti-inflammatory and immunomodulatory effects in colitis models. Thus, previous papers had shown that administration of melatonin ameliorated the experimental colitis through the downregulation of the inducible enzymes iNOS and COX-2 in the colonic mucus [128, 138], as well as a decrease in colonic NO and PGE2 content [138]. Mei et al. [129] investigated the change of NO in colitis and its inhibition by melatonin in vivo and in vitro. For their in vitro study, they constructed a coculture model of the inflamed colon mucosal tissue and found that melatonin significantly reduced LPS-induced NO production. These results were confirmed after intracolonic melatonin administration in rats with TNBS-mediated colitis. Other published reports have suggested that the anti-inflammatory effect of melatonin is related to the inhibition of the production or expression of proinflammatory cytokines, including TNF-α and IL-1 [131, 139–141] or the reduction in the adhesion molecule expression, i.e., ICAM-1 or P-selectin [128, 140]. Moreover, Tahan et al. [132] have recently noted a decrease in the proinflammatory cytokines IL-1β, IL-6, and TNF-α in UC mucosal tissue by the indoleamine. Also, Esposito et al. [131] reported the beneficial properties of this molecule in the experimental colitis in rats through downregulation of the matrix metalloproteinases MMP-9 and MMP-2 activity and expression, which was most likely attributed to the inhibitory effect on the TNF-α production. Collectively, these findings may help to explain the reduced neutrophil infiltration into the inflamed tissue detected in all of these experimental models of colitis (Fig. 3). However, a previous publication from our group, which investigates in depth the role of melatonin in TNBS-induced intestinal damage, found divergent data because the short-term administration of melatonin caused a reduction in the damage score and myeloperoxidase (MPO) activity, accompanied by attenuation in the TNF-α production, while chronic treatment negatively influenced the development of colitis [142].

Several interesting studies providing further insight into the molecular mechanisms of melatonin action in the colon reported that the anti-inflammatory effects of this molecule may be related to the known inhibitory effect on the activation of NF-kB [143, 144]. In accordance with these findings, Li et al. [140] demonstrated that intrarectal administration of melatonin reduced colonic inflammatory injury induced by TNBS and improved colitis symptoms through the downregulation of proinflammatory molecules, mediated by NF-κB inhibition and blockade of IκBα degradation. Mazzon et al. [141] obtained similar results after the intraperitoneal injection of melatonin in rats with DNBS-induced colitis. Moreover, these authors found that this molecule reduced the expression of proapoptotic Bax and prevented the loss of antiapoptotic Bcl-2 proteins as well as the presence of apoptotic cells caused by DNBS, suggesting that the reduction in colon damage was also associated with a reduction in apoptosis because of melatonin. Along the same lines, treatment with melatonin significantly decreased caspase-3 activity compared with that in TNBS-treated rats [130]. These results support the idea that a reduction in mucosal damage is a consequence of the anti-inflammatory and anti-apoptotic effects of melatonin.

IBD clinical studies

In addition to experimental in vivo and in vitro studies, preliminary data from human trials provide evidence that melatonin supplementation may have positive effects on UC. Thus, Mann [145] reported a case of a patient with UC who intermittently self-administered melatonin to ameliorate jet lag, each time resulting in the disappearance of his symptoms of colitis. However, there are a few cases, in which this indoleamine seemed to aggravate the symptoms of colitis. Calvo et al. [146] published the history of a woman with controlled CD who started to take melatonin capsules (3 mg) before going to sleep. Four days later, the patient experienced the symptoms of active CD, including diarrhea and abdominal cramps. She then stopped taking melatonin, and 24 hr later, there was a complete remission of symptoms. More recently, these authors reported another case of a man whose UC symptoms reappeared 2 months after starting to take daily melatonin (3 mg) for sleep promotion that subsided shortly after discontinuing melatonin [147]. The reasons for these conflicting observations are unclear and may be attributed to the immunostimulatory role of melatonin. More clinical studies are required to clarify the utility of this molecule in the treatment of IBD.

In summary, the anti-inflammatory effects of melatonin have been attributed to a variety of mechanisms, including inhibition of COX-2 and iNOS expression and the subsequent production of PGE2 and NO, suppression of specific inflammation-related cytokines and cell adhesion molecules, attenuation of NF-kB activation, reduction in matrix metalloproteinase-2 and metalloproteinase-9 activity, and modulation of apoptosis [148]. Further studies are needed to enhance the understanding of the role of melatonin in the pathophysiology of IBD and to determine how this indoleamine combines with established drugs such as sulfasalazine and corticosteroids in a way that is compatible with the quality of life of individual patients.

Role of melatonin in colon cancer

A series of studies have shown the oncostatic activity of melatonin both in vitro and in vivo for different types of tumors [149–153]. A growing body of evidence implicates melatonin’s antioxidant/free-radical-scavenging actions in the inhibition of cancer development and growth [154]. Other mechanisms involved in the anticarcinogenic effect of the indoleamine are related to the modulation of the endocrine system [155], its immunoenhancing properties [156], and its direct effect on cancer cell proliferation [157]. Interest in its possible role in colorectal cancer increased following the identification of melatonin-binding sites in human colon tissue from patients with carcinoma of the rectum or colon [158]. Melatonin secretion has been shown to be impaired in patients suffering from colorectal cancer [159]. Moreover, a higher risk of developing this kind of cancer has been observed in nurses and other night-shift workers, suggesting a possible link between diminished secretion of melatonin and increased exposure to light during nighttime [160, 161].

The ability of melatonin to inhibit the growth of several tumor cell lines, including colon cancer cells, has been repeatedly reported. Thus, Karasek et al. [162] found that melatonin exerted an anti-proliferative effect on murine colon 38 cancer cells both in vitro and in vivo. In another interesting study, this research group evaluated the role of the nuclear melatonin receptor, which is considered identical to the so-called nuclear orphan receptor RZR/ROR, on control of cell growth and differentiation. These authors compared the effects of melatonin with those from CGP 52608, an exogenous ligand for RZR/ROR receptors, and demonstrated that both compounds exerted similar inhibitory effects on the proliferation of neoplastic cells in mouse colonic adenocarcinoma [163]. A recent study further supports the nuclear effect of melatonin in the inhibition of HT-29 cell proliferation. This suggests that the antioxidative and anti-inflammatory actions of melatonin, by counteracting the oxidative status and reducing the production of NO by HT-29 cells, are directly involved in the oncostatic properties of melatonin [164].

As estradiol receptors are involved in the antiproliferative properties of melatonin in breast tumor cells [165], Farriol et al. [166] evaluated the presence of these receptors in the murine colon carcinoma-derived cell line CT-26 and the effect of melatonin on suppression of cell growth. Data from this study showed that although melatonin had no effect on cell growth at low doses, a statistically significant and progressive inhibition of DNA synthesis was found as the dose od melatonin increased. Inasmuch as no measurable estradiol receptors were found in the cell line selected, it was concluded that melatonin exerted its antiproliferative action through a nonhormonal-dependent mechanism.

Apart from the inhibition of tumor cell proliferation, another mechanism by which the oncostatic agents may exert their antitumoral effects is via the induction of apoptosis [167]. In neurons and immune cells, melatonin protects cells from apoptosis [168, 169], but in various cancer cells, it was found to induce apoptosis [150, 167, 170]. Melen-Mucha et al. [171] evaluated the effect of this indoleamine on the apoptosis of colon cancer cells using a xenograft model. In this study, mice were implanted a suspension of Colon-38 cells subcutaneously, and after 6 days, the animals were subcutaneously injected melatonin. The data presented showed, for the first time, that this molecule enhanced apoptosis in murine colonic cancer. Subsequently, this group reported the role of the nuclear RZR/ROR receptor in the proapoptotic action of melatonin, by using either a ligand of RZR/ROR, which mimicked the effect of melatonin [172] or an antagonist, which diminished the oncostatic effect of melatonin on murine colon [173]. Melatonin has also been shown to promote apoptosis when initiated by flavone in HT-29 human colon cancer cells by enhancing the level of oxidizable substrates that can be incorporated into mitochondria in the presence of this flavonoid [174]. Along the same lines, Gonzalez-Puga et al. [175] reported that the indoleamine significantly inhibited cell proliferation and increased cancer cell death. Interestingly enough, melatonin acted synergistically with several CCK-A antagonists, mainly devazepide, to further reduce HT-29 cell growth. These findings suggest that combined therapy with both molecules may be a good choice for colon cancer treatment.

Experimental in vivo studies have also documented the protective effects of exogenous melatonin on colon carcinogenesis. The first of these studies was carried out by Anisimov et al. in 1997 [176]; they reported an inhibitory effect of the indoleamine on intestinal carcinogenesis induced by 1,2-dimethylhydrazine (DMH) in rats. This effect was manifested by a reduction in the incidence and multiplicity of bowel tumors, mainly of the colon, and by a reduced invasion rate and dimensions of colon carcinomas. A similar study by these authors showed that the antineoplastic effect of melatonin in rats exposed to DMH was correlated with a significant inhibitory effect of indoleamine on mitotic index and a stimulatory effect on the relative number of apoptotic cell in colon tumors [177]. In a subsequent report, it was demonstrated that the anticarcinogenic effects of melatonin were related to activation of several elements of the host’s lymphatic system [178]. Tanaka et al. [179] have recently demonstrated that melatonin administration in drinking water effectively suppressed AOM/DSS-induced rat colitis-related colonic oncogenesis. The cancer chemopreventive ability by melatonin was related to the suppression of several colon carcinogenesis biomarkers, including NF-κB, TNF-α, IL-1β, and STAT3, as well as the reduction in the mitotic and apoptotic indices in the colonic adenocarcinomas.

Collectively, the findings indicate a beneficial effect of melatonin on colon carcinogenesis and suggest its potential application for inhibiting colorectal cancer development. However, there are few definitive data establishing a beneficial function of melatonin in chronic intestinal inflammation and its evolution to colon cancer. It would be of interest to examine these mechanisms and the role played by melatonin in them.

New paradigms in inflammatory intestinal disease and colorectal cancer: role of melatonin

Both autophagy [72, 180] and sirtuins [88, 181] have a relevant role in disease, and remarkably in the control of cancer. As melatonin directly or indirectly modulates both processes, some of the effects of melatonin on colon cancer could be related to sirtuin regulation. This section is dedicated to analyzing the connections between melatonin, sirtuins and autophagy, and the relationship of these processes to inflammatory intestinal disease and colorectal cancer.

Melatonin and autophagy

Melatonin increases autophagy in a few experimental systems, such as senescence-accelerated prone mice 8 [182] and ischemia/reperfusion-injured neurons [183]. However, a number of data supports an inhibitory role of melatonin on autophagy [184–190].

Oxidative stress can switch on autophagy; for this reason, the antioxidant activity of melatonin could account for its inhibitory effects on autophagy. If so, melatonin would be acting on mechanisms capable of stimulating autophagy and not on the process itself. In most of the studies, melatonin reduced the autophagic process that were triggered by circumstances that caused ROS formation and oxidative stress [184–189]. Melatonin protected neuronal cell from death in ischemic brain injury in rats by preventing the injury-induced decrease in the phosphorylation of mTOR [187]. Similar negative effects on mTOR inactivation were reported in hepatoma cells [186] and SK-N-SH cells [188]. In this latter experimental system, the effects of melatonin were also related to Bcl-2/Beclin1 pathway [189]. Thus, melatonin seems to inhibit autophagy triggered either by mTOR inactivation or by Beclin 1 activation.

Cyclosporine A is known to induce autophagy via endoplasmic reticulum stress [84]. Melatonin suppresses cyclosporine-induced autophagy in rat pituitary GH3 cells [190]. This effect is related to the MAPK/ERK pathway and consequently, totally or in part, to the antioxidant properties of melatonin.

In summary, anti-autophagic properties of melatonin could contribute to its anti-oncogenic activity. As discussed in an earlier section, the inhibition of autophagy has excellent potential in cancer treatment.

Melatonin and sirtuins

Epidemiological and genetic studies show that the alterations in circadian rhythms may lead to an increased risk for some cancers [191]; for example, Per2-mutant mice, which have a shortened circadian period, develop colonic polyps and have increased β-catenin and cyclin D protein levels in small intestinal mucosa [192]. Several recent studies have linked SIRT1 to the circadian rhythm machinery through direct deacetylation activity as well as through the nicotinamide adenine dinucleotide (NAD⁺) salvage pathway [88, 193]. The regulation by SIRT1 of several circadian clock-associated genes would transduce signals from cellular metabolism to the circadian clock. Also, the intracellular levels of NAD⁺, which positively regulate SIRT1 activity, show circadian oscillations dependent on NAMPT (nicotinamide ribosyltransferase) circadian expression. Alteration in the activity of clock regulators, such as SIRT1, could contribute to cancer by causing higher proliferation and defects in metabolic pathways.

Melatonin is synthesized and released in a circadian rhythm, and a reduction in melatonin levels has been linked to an increase in cancer risk [194, 195], including colon cancer [196]. In addition, several studies suggest that melatonin regulates clock gene expression [197–200], and decreased melatonin levels have been linked to disruption of circadian rhythms. Thus, it is possible to establish a link between sirtuin, melatonin, and cancer through the regulation of circadian rhythms [88]. In this model, SIRT1 and melatonin would operate in opposite ways, SIRT1 counteracting the activity of the core components and melatonin re-synchronizing the clock. Increased levels of SIRT1 have been reported in some colon cancers [95]; this would cause alterations of circadian rhythms. Melatonin, counteracting the effects of SIRT1, would prevent molecular damage resulting from SIRT1 changes. In this respect, melatonin inhibits SIRT1 protein expression and activity in human prostate cancer cell line and in transgenic adenocarcinoma of mice prostate cancer [201].

While the ‘circadian connection’ model shows melatonin-reducing SIRT1 activity, there is also experimental evidence pointing to the opposite. Melatonin treatment effectively preserved the relative protein levels of SIRT1 in hippocampus of adult male Wistar rats. In this animal model, sleep deprivation caused a drastic reduction of SIRT1, while exogenous melatonin administration preserved SIRT1 expression [202]. However, the effect of melatonin on SIRT1 levels in control rats was not examined. Similarly, the level of SIRT1 expression was lower in senescence-accelerated mice (SAMP8) than in senescence-accelerated resistant mice (SAMR1), and this reduction was prevented by melatonin [203]. Again, there was no information regarding the effects of melatonin in sirtuin levels of control mice. An in vitro murine model of neuronal aging was used to test whether melatonin acts as a sirtuin inducer; it was observed that melatonin enhanced the level of SIRT1 and the degree of deacetylation of several substrates of SIRT1 (p53, PGC-1α, FoxO1, ADAM10, and NFκB) [204]. In this latter model, melatonin exhibited a neuroprotective effect mediated by its positive regulation of SIRT1 activity.

These apparent contradictory results (melatonin both inhibiting and increasing sirtuin activity) could be reconciled if it is assumed that melatonin corrects the alterations in sirtuin activity associated with several pathological conditions, inhibiting SIRT1 in cancer cells that have abnormally high levels of SIRT1 activity, and preserving it in situations that cause its decrease. The vast information being generated in recent years will hopefully help to clarify these issues.

Concluding remarks and open questions

Current data show that the inhibition of autophagy is a good approach to arrest cancer progression. Furthermore, this treatment sensitizes cancer cells to cytotoxic agents. In most of the available studies, melatonin inhibited autophagy, and this effect was commonly related to its antioxidant activity. Does melatonin regulate the autophagic process by other means? Pharmacological studies with cancer cell lines would likely provide information if melatonin membrane and nuclear receptors, or calmodulin-mediated action, were implicated in its effects in these experimental systems. In addition, there is no information about whether the antitumor effects of melatonin on colon cancer are dependent or independent of autophagy. This should be evaluated.

Genetic alterations in autophagy promote IBD and colorectal cancer. As melatonin has well-known anti-inflammatory properties, it would be interesting to test whether it can prevent the progression from IBD to colon cancer in these genetically altered individuals. Fig. 4 summarizes autophagy-related effects of melatonin on colon cancer development.

Figure 4.  Melatonin and autophagy in intestinal bowel disease and colorectal cancer. Genetic alteration in autophagy (risk alleles for ATG 16L1, IRGM, NOD2, and XBP1) promotes the progression from chronic inflammation to cancer development. Its proinflammatory effects are counteracted by melatonin, although whether melatonin is able to antagonize genetic alteration in autophagy at further steps is not known. Most of experimental data support the inhibition of autophagy by melatonin, probably due to its antioxidant activity. As autophagy promotes cancer development, the inhibition of autophagy could contribute to the antioncogenic effects of melatonin.

Sirtuins have pleiotropic effects on cancer, being depicted both as tumor promoter and as tumor-suppressor agents. It is hypothesized that the important issue is an imbalance of sirtuinas levels, more than whether their levels are high or low. Through the ‘circadian clock connection’, melatonin could balance the distorted levels of sirtuins and promote anti-oncogenic effects. Melatonin could also prevent or correct damage derived from imbalanced levels of sirtuins.

There is no information related to the direct effects of melatonin on sirtuin activity. We have not found an effect of melatonin on SIRT1, SIRT2, or SIRT3 activity with the ‘Fluor-de-Lys’ fluorimetric assay, so further investigation is necessary to clarify this point. In addition, the majority of available data concerns SIRT1. It would be desirable to evaluate the levels of the several human sirtuins in colorectal cancers and to determine whether melatonin alters these levels. Fig. 5 summarizes sirtuin-related effects of melatonin on colon cancer development.

Figure 5.  Melatonin and sirtuins in intestinal bowel disease and colorectal cancer. Perturbations of circadian rhythms sustain chronic inflammation and further development of colon cancer. Both melatonin and SIRT1 activity regulate, and are regulated by, the circadian machinery. Through the circadian clock connection, melatonin could counteract the pro-oncogenic effects of altered levels of sirtuins. In addition, independently from the circadian clock, melatonin could restore distorted sirtuin activity, preserving its anti-inflammatory action, and preventing the pro-oncogenic effects of too high level of sirtuins.

Longevity and cancer are the two faces of the same coin. Resveratrol and rapamycin, the best-known activators of SIRT1 and autophagy, respectively, have been named ‘anti-aging drugs’ [205]. Both molecules modulate AMPK, which seems to be a central regulator of cell metabolism and to control the delicate equilibrium between prosurvival and prodeath pathways. The effect of melatonin, another molecule with known anti-aging properties via AMPK, as well as the impact of this action on IBD and colorectal cancer remains to be investigated.