Neutrophil-activating secretome characterizes palbociclib-induced senescence of breast cancer cells.

Senescent cells have a profound impact on the surrounding microenvironment through the secretion of numerous bioactive molecules and inflammatory factors. The induction of therapy-induced senescence by anticancer drugs is known, but how senescent tumor cells influence the tumor immune landscape, particularly neutrophil activity, is still unclear. In this study, we investigate the induction of cellular senescence in breast cancer cells and the subsequent immunomodulatory effects on neutrophils using the CDK4/6 inhibitor palbociclib, which is approved for the treatment of breast cancer and is under intense investigation for additional malignancies. Our research demonstrates that palbociclib induces a reversible form of senescence endowed with an inflammatory secretome capable of recruiting and activating neutrophils, in part through the action of interleukin-8 and acute-phase serum amyloid A1. The activation of neutrophils is accompanied by the release of neutrophil extracellular trap and the phagocytic removal of senescent tumor cells. These findings may be relevant for the success of cancer therapy as neutrophils, and neutrophil-driven inflammation can differently affect tumor progression. Our results reveal that neutrophils, as already demonstrated for macrophages and natural killer cells, can be recruited and engaged by senescent tumor cells to participate in their clearance. Understanding the interplay between senescent cells and neutrophils may lead to innovative strategies to cope with chronic or tumor-associated inflammation.

Comparative genomics provides insights into molecular adaptation to hypermetamorphosis and cantharidin metabolism in blister beetles (Coleoptera: Meloidae).

Blister beetles (Coleoptera: Meloidae) are currently subdivided into three subfamilies: Eleticinae (a basal group), Nemognathinae, and Meloinae. These are all characterized by the endogenous production of the defensive terpene cantharidin (CA), whereas the two most derived subfamilies show a hypermetamorphic larval development. Here, we provide novel draft genome assemblies of five species sampled across the three blister beetle subfamilies (Iselma pallidipennis, Stenodera caucasica, Zonitis immaculata, Lydus trimaculatus, and Mylabris variabilis) and performed a comparative analysis with other available Meloidae genomes and the closely-related canthariphilous species (Pyrochroa serraticornis) to disclose adaptations at a molecular level. Our results highlighted the expansion and selection of genes potentially responsible for CA production and metabolism, as well as its mobilization and vesicular compartmentalization. Furthermore, we observed adaptive selection patterns and gain of genes devoted to epigenetic regulation, development, and morphogenesis, possibly related to hypermetamorphosis. We hypothesize that most genetic adaptations occurred to support both CA biosynthesis and hypermetamorphosis, two crucial aspects of Meloidae biology that likely contributed to their evolutionary success.

Spermine Oxidase-Substrate Electrostatic Interactions: The Modulation of Enzyme Function by Neighboring Colloidal ɣ-Fe2O3.

Protein-nanoparticle hybridization can ideally lead to novel biological entities characterized by emerging properties that can sensibly differ from those of the parent components. Herein, the effect of ionic strength on the biological functions of recombinant His-tagged spermine oxidase (i.e., SMOX) was studied for the first time. Moreover, SMOX was integrated into colloidal surface active maghemite nanoparticles (SAMNs) via direct self-assembly, leading to a biologically active nano-enzyme (i.e., SAMN@SMOX). The hybrid was subjected to an in-depth chemical-physical characterization, highlighting the fact that the protein structure was perfectly preserved. The catalytic activity of the nanostructured hybrid (SAMN@SMOX) was assessed by extracting the kinetics parameters using spermine as a substrate and compared to the soluble enzyme as a function of ionic strength. The results revealed that the catalytic function was dominated by electrostatic interactions and that they were drastically modified upon hybridization with colloidal ɣ-Fe2O3. The fact that the affinity of SMOX toward spermine was significantly higher for the nanohybrid at low salinity is noteworthy. The present study supports the vision of using protein-nanoparticle conjugation as a means to modulate biological functions.

From Monoamine Oxidase Inhibition to Antiproliferative Activity: New Biological Perspectives for Polyamine Analogs.

Monoamine oxidases (MAOs) are well-known pharmacological targets in neurological and neurodegenerative diseases. However, recent studies have revealed a new role for MAOs in certain types of cancer such as glioblastoma and prostate cancer, in which they have been found overexpressed. This finding is opening new frontiers for MAO inhibitors as potential antiproliferative agents. In light of our previous studies demonstrating how a polyamine scaffold can act as MAO inhibitor, our aim was to search for novel analogs with greater inhibitory potency for human MAOs and possibly with antiproliferative activity. A small in-house library of polyamine analogs (2-7) was selected to investigate the effect of constrained linkers between the inner amine functions of a polyamine backbone on the inhibitory potency. Compounds 4 and 5, characterized by a dianiline (4) or dianilide (5) moiety, emerged as the most potent, reversible, and mainly competitive MAO inhibitors (Ki < 1 µM). Additionally, they exhibited a high antiproliferative activity in the LN-229 human glioblastoma cell line (GI50 < 1 µM). The scaffold of compound 5 could represent a potential starting point for future development of anticancer agents endowed with MAO inhibitory activity.

Spermine oxidase induces DNA damage and sensitizes fusion negative rhabdomyosarcoma cells to irradiation.

Rhabdomyosarcoma (RMS) is a pediatric myogenic soft tissue sarcoma that includes fusion-positive (FP) and fusion-negative (FN) molecular subtypes. FP-RMS expresses PAX3-FOXO1 fusion protein and often shows dismal prognosis. FN-RMS shows cytogenetic abnormalities and frequently harbors RAS pathway mutations. Despite the multimodal heavy chemo and radiation therapeutic regimens, high risk metastatic/recurrent FN-RMS shows a 5-year survival less than 30% due to poor sensitivity to chemo-radiotherapy. Therefore, the identification of novel targets is needed. Polyamines (PAs) such as putrescine (PUT), spermidine (SPD) and spermine (SPM) are low-molecular-mass highly charged molecules whose intracellular levels are strictly modulated by specific enzymes. Among the latter, spermine oxidase (SMOX) regulates polyamine catabolism oxidizing SPM to SPD, which impacts cellular processes such as apoptosis and DNA damage response. Here we report that low SMOX levels are associated with a worse outcome in FN-RMS, but not in FP-RMS, patients. Consistently, SMOX expression is downregulated in FN-RMS cell lines as compared to normal myoblasts. Moreover, SMOX transcript levels are reduced FN-RMS cells differentiation, being indirectly downregulated by the muscle transcription factor MYOD. Noteworthy, forced expression of SMOX in two cell lines derived from high-risk FN-RMS: 1) reduces SPM and upregulates SPD levels; 2) induces G0/G1 cell cycle arrest followed by apoptosis; 3) impairs anchorage-independent and tumor spheroids growth; 4) inhibits cell migration; 5) increases γH2AX levels and foci formation indicative of DNA damage. In addition, forced expression of SMOX and irradiation synergize at activating ATM and DNA-PKCs, and at inducing γH2AX expression and foci formation, which suggests an enhancement in DNA damage response. Irradiated SMOX-overexpressing FN-RMS cells also show significant decrease in both colony formation capacity and spheroids growth with respect to single approaches. Thus, our results unveil a role for SMOX as inhibitor of tumorigenicity of FN-RMS cells in vitro. In conclusion, our in vitro results suggest that SMOX induction could be a potential combinatorial approach to sensitize FN-RMS to ionizing radiation and deserve further in-depth studies.

The Impact of Spermidine on C2C12 Myoblasts Proliferation, Redox Status and Polyamines Metabolism under H2O2 Exposure.

A central feature of the skeletal muscle is its ability to regenerate through the activation, by environmental signals, of satellite cells. Once activated, these cells proliferate as myoblasts, and defects in this process profoundly affect the subsequent process of regeneration. High levels of reactive oxygen species such as hydrogen peroxide (H2O2) with the consequent formation of oxidized macromolecules increase myoblasts' cell death and strongly contribute to the loss of myoblast function. Recently, particular interest has turned towards the beneficial effects on muscle of the naturally occurring polyamine spermidine (Spd). In this work, we tested the hypothesis that Spd, upon oxidative challenge, would restore the compromised myoblasts' viability and redox status. The effects of Spd in combination with aminoguanidine (Spd-AG), an inhibitor of bovine serum amine oxidase, on murine C2C12 myoblasts treated with a mild dose of H2O2 were evaluated by analyzing: (i) myoblast viability and recovery from wound scratch; (ii) redox status and (iii) polyamine (PAs) metabolism. The treatment of C2C12 myoblasts with Spd-AG increased cell number and accelerated scratch wound closure, while H2O2 exposure caused redox status imbalance and cell death. The combined treatment with Spd-AG showed an antioxidant effect on C2C12 myoblasts, partially restoring cellular total antioxidant capacity, reducing the oxidized glutathione (GSH/GSSG) ratio and increasing cell viability through a reduction in cell death. Moreover, Spd-AG administration counteracted the induction of polyamine catabolic genes and PA content decreased due to H2O2 challenges. In conclusion, our data suggest that Spd treatment has a protective role in skeletal muscle cells by restoring redox balance and promoting recovery from wound scratches, thus making myoblasts able to better cope with an oxidative insult.

The Involvement of Polyamines Catabolism in the Crosstalk between Neurons and Astrocytes in Neurodegeneration.

In mammalian cells, the content of polyamines is tightly regulated. Polyamines, including spermine, spermidine and putrescine, are involved in many cellular processes. Spermine oxidase specifically oxidizes spermine, and its deregulated activity has been reported to be linked to brain pathologies involving neuron damage. Spermine is a neuromodulator of a number of ionotropic glutamate receptors and types of ion channels. In this respect, the Dach-SMOX mouse model overexpressing spermine oxidase in the neocortex neurons was revealed to be a model of chronic oxidative stress, excitotoxicity and neuronal damage. Reactive astrocytosis, chronic oxidative and excitotoxic stress, neuron loss and the susceptibility to seizure in the Dach-SMOX are discussed here. This genetic model would help researchers understand the linkage between polyamine dysregulation and neurodegeneration and unveil the roles of polyamines in the crosstalk between astrocytes and neurons in neuroprotection or neurodegeneration.

Molecular Characterization of Kunitz-Type Protease Inhibitors from Blister Beetles (Coleoptera, Meloidae).

Protease inhibitors are widely studied since the unrestricted activity of proteases can cause extensive organ lesions. In particular, elastase activity is involved in the pathophysiology of acute lung injury, for example during SARS-CoV-2 infection, while serine proteases and thrombin-like proteases are involved in the development and/or pathology of the nervous system. Natural protease inhibitors have the advantage to be reversible and with few side effects and thus are increasingly considered as new drugs. Kunitz-type protease inhibitors (KTPIs), reported in the venom of various organisms, such as wasps, spiders, scorpions, and snakes, have been studied for their potent anticoagulant activity and widespread protease inhibitor activity. Putative KTPI anticoagulants have been identified in transcriptomic resources obtained for two blister beetle species, Lydus trimaculatus and Mylabris variabilis. The KTPIs of L. trimaculatus and M. variabilis were characterized by combined transcriptomic and bioinformatics methodologies. The full-length mRNA sequences were divided on the base of the sequence of the active sites of the putative proteins. In silico protein structure analyses of each group of translational products show the biochemical features of the active sites and the potential protease targets. Validation of these genes is the first step for considering these molecules as new drugs for use in medicine.

MET Inhibition Sensitizes Rhabdomyosarcoma Cells to NOTCH Signaling Suppression.

Rhabdomyosarcoma (RMS) is a pediatric myogenic soft tissue sarcoma. The Fusion-Positive (FP) subtype expresses the chimeric protein PAX3-FOXO1 (P3F) while the Fusion-Negative (FN) is devoid of any gene translocation. FP-RMS and metastatic FN-RMS are often unresponsive to conventional therapy. Therefore, novel therapeutic approaches are needed to halt tumor progression. NOTCH signaling has oncogenic functions in RMS and its pharmacologic inhibition through γ-secretase inhibitors blocks tumor growth in vitro and in vivo. Here, we show that NOTCH signaling blockade resulted in the up-regulation and phosphorylation of the MET oncogene in both RH30 (FP-RMS) and RD (FN-RMS) cell lines. Pharmacologic inhibition of either NOTCH or MET signaling slowed proliferation and restrained cell survival compared to control cells partly by increasing Annexin V and CASP3/7 activation. Co-treatment with NOTCH and MET inhibitors significantly amplified these effects and enhanced PARP1 cleavage in both cell lines. Moreover, it severely hampered cell migration, colony formation, and anchorage-independent growth compared to single-agent treatments in both cell lines and significantly prevented the growth of FN-RMS cells grown as spheroids. Collectively, our results unveil the overexpression of the MET oncogene by NOTCH signaling targeting in RMS cells and show that MET pathway blockade sensitizes them to NOTCH inhibition.

Male Accessory Glands of Blister Beetles and Cantharidin Release: A Comparative Ultrastructural Analysis.

Members of the family Meloidae are known to produce cantharidin, a highly toxic monoterpene found in their hemolymph and exuded as droplets capable of deterring many predators. As a nuptial gift, males transfer large amounts of cantharidin to females via a spermatophore, which is formed by specific accessory glands containing high concentrations of this terpene. Using light, electron and ion beam microscopy, the ultrastructural features of the three pairs of male accessory glands as well as the glandular part of the vasa deferentia were comparatively investigated in seven species of blister beetles belonging to five different tribes and two subfamilies. All gland pairs examined share common features such as mesodermal derivation, the presence of muscle sheath, a developed rough endoplasmic reticulum, abundant mitochondria, secretory vesicles, and microvillated apical membranes. Within the same species, glands exhibit distinctive features, suggesting that each pair is responsible for the formation of a specific substance. The vasa deferentia, while showing many similarities within the family, often exhibit features unique to each of the individual species investigated, whereas the accessory glands of the first and second pairs display the highest degree of ultrastructural variability. A comparison across the species shows an interesting constancy limited to ultrastructural features in the third pair of accessory glands. The similarities and differences among the species are discussed in the light of the available literature and in relation to the potential role that blister beetles' male accessory glands could play in the storage and management of cantharidin.

Transgenic Mouse Overexpressing Spermine Oxidase in Cerebrocortical Neurons: Astrocyte Dysfunction and Susceptibility to Epileptic Seizures.

Polyamines are organic polycations ubiquitously present in living cells. Polyamines are involved in many cellular processes, and their content in mammalian cells is tightly controlled. Among their function, these molecules modulate the activity of several ion channels. Spermine oxidase, specifically oxidized spermine, is a neuromodulator of several types of ion channel and ionotropic glutamate receptors, and its deregulated activity has been linked to several brain pathologies, including epilepsy. The Dach-SMOX mouse line was generated using a Cre/loxP-based recombination approach to study the complex and critical functions carried out by spermine oxidase and spermine in the mammalian brain. This mouse genetic model overexpresses spermine oxidase in the neocortex and is a chronic model of excitotoxic/oxidative injury and neuron vulnerability to oxidative stress and excitotoxic, since its phenotype revealed to be more susceptible to different acute oxidative insults. In this review, the molecular mechanisms underlined the Dach-SMOX phenotype, linked to reactive astrocytosis, neuron loss, chronic oxidative and excitotoxic stress, and susceptibility to seizures have been discussed in detail. The Dach-SMOX mouse model overexpressing SMOX may help in shedding lights on the susceptibility to epileptic seizures, possibly helping to understand the mechanisms underlying epileptogenesis in vulnerable individuals and contributing to provide new molecular mechanism targets to search for novel antiepileptic drugs.

Unraveling the role of male reproductive tract and haemolymph in cantharidin-exuding Lydus trimaculatus and Mylabris variabilis (Coleoptera: Meloidae): a comparative transcriptomics approach.

BACKGROUND: Meloidae (blister beetles) are known to synthetize cantharidin (CA), a toxic and defensive terpene mainly stored in male accessory glands (MAG) and emitted outward through reflex-bleeding. Recent progresses in understanding CA biosynthesis and production organ(s) in Meloidae have been made, but the way in which self-protection is achieved from the hazardous accumulation and release of CA in blister beetles has been experimentally neglected. To provide hints on this pending question, a comparative de novo assembly transcriptomic approach was performed by targeting two tissues where CA is largely accumulated and regularly circulates in Meloidae: the male reproductive tract (MRT) and the haemolymph. Differential gene expression profiles in these tissues were examined in two blister beetle species, Lydus trimaculatus (Fabricius, 1775) (tribe Lyttini) and Mylabris variabilis (Pallas, 1781) (tribe Mylabrini). Upregulated transcripts were compared between the two species to identify conserved genes possibly involved in CA detoxification and transport. RESULTS: Based on our results, we hypothesize that, to avoid auto-intoxication, ABC, MFS or other solute transporters might sequester purported glycosylated CA precursors into MAG, and lipocalins could bind CA and mitigate its reactivity when released into the haemolymph during the autohaemorrhaging response. We also found an over-representation in haemolymph of protein-domains related to coagulation and integument repairing mechanisms that likely reflects the need to limit fluid loss during reflex-bleeding. CONCLUSIONS: The de novo assembled transcriptomes of L. trimaculatus and M. variabilis here provided represent valuable genetic resources to further explore the mechanisms employed to cope with toxicity of CA in blister beetle tissues. These, if revealed, might help conceiving safe and effective drug-delivery approaches to enhance the use of CA in medicine.

Effect of Tadalafil Administration on Redox Homeostasis and Polyamine Levels in Healthy Men with High Level of Physical Activity.

Background: The phosphodiesterase type 5 inhibitor (PDE5I) tadalafil, in addition to its therapeutic role, has shown antioxidant effects in different in vivo models. Supplementation with antioxidants has received interest as a suitable tool for preventing or reducing exercise-related oxidative stress, possibly leading to the improvement of sport performance in athletes. However, the use/abuse of these substances must be evaluated not only within the context of amateur sport, but especially in competitions where elite athletes are more exposed to stressful physical practice. To date, very few human studies have addressed the influence of the administration of PDE5Is on redox balance in subjects with a fitness level comparable to elite athletes; therefore, the aim of this study was to investigate for the first time whether acute ingestion of tadalafil could affect plasma markers related to cellular damage, redox homeostasis, and blood polyamines levels in healthy subjects with an elevated cardiorespiratory fitness level. Methods: Healthy male volunteers (n = 12), with a VO2max range of 40.1-56.0 mL/(kg × min), were administered with a single dose of tadalafil (20 mg). Plasma molecules related to muscle damage and redox-homeostasis, such as creatine kinase (CK), lactate dehydrogenase (LDH), total antioxidant capacity (TAC), reduced/oxidized glutathione ratio (GSH/GSSG), free thiols (FTH), antioxidant enzyme activities (superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx)), as well as thiobarbituric acid reactive substances (TBARs), protein carbonyls (PrCAR), and polyamine levels (spermine (Spm) and spermidine (Spd)) were evaluated immediately before and 2, 6 and 24 hours after the acute tadalafil administration. Results: A single tadalafil administration induced an increase in CK and LDH plasma levels 24 after consumption. No effects were observed on redox homeostasis or antioxidant enzyme activities, and neither were they observed on the oxidation target molecules or polyamines levels. Conclusion: Our results show that in subjects with an elevated fitness level, a single administration of tadalafil induced a significant increase in muscle damage target without affecting plasma antioxidant status.

Reactive Astrocytosis in a Mouse Model of Chronic Polyamine Catabolism Activation.

BACKGROUND: In the brain, polyamines are mainly synthesized in neurons, but preferentially accumulated in astrocytes, and are proposed to be involved in neurodegenerative/neuroinflammatory disorders and neuron injury. A transgenic mouse overexpressing spermine oxidase (SMOX, which specifically oxidizes spermine) in the neocortex neurons (Dach-SMOX mouse) was proved to be a model of increased susceptibility to excitotoxic injury. METHODS: To investigate possible alterations in synapse functioning in Dach-SMOX mouse, both cerebrocortical nerve terminals (synaptosomes) and astrocytic processes (gliosomes) were analysed by assessing polyamine levels, ezrin and vimentin content, glutamate AMPA receptor activation, calcium influx, and catalase activity. RESULTS: The main findings are as follows: (i) the presence of functional calcium-permeable AMPA receptors in synaptosomes from both control and Dach-SMOX mice, and in gliosomes from Dach-SMOX mice only; (ii) reduced content of spermine in gliosomes from Dach-SMOX mice; and (iii) down-regulation and up-regulation of catalase activity in synaptosomes and gliosomes, respectively, from Dach-SMOX mice. CONCLUSIONS: Chronic activation of SMOX in neurons leads to major changes in the astrocyte processes including reduced spermine levels, increased calcium influx through calcium-permeable AMPA receptors, and stimulation of catalase activity. Astrocytosis and the astrocyte process alterations, depending on chronic activation of polyamine catabolism, result in synapse dysregulation and neuronal suffering.

Emerging Role for Linear and Circular Spermine Oxidase RNAs in Skeletal Muscle Physiopathology.

Skeletal muscle atrophy is a pathological condition so far without effective treatment and poorly understood at a molecular level. Emerging evidence suggest a key role for circular RNAs (circRNA) during myogenesis and their deregulation has been reported to be associated with muscle diseases. Spermine oxidase (SMOX), a polyamine catabolic enzyme plays a critical role in muscle differentiation and the existence of a circRNA arising from SMOX gene has been recently identified. In this study, we evaluated the expression profile of circular and linear SMOX in both C2C12 differentiation and dexamethasone-induced myotubes atrophy. To validate our findings in vivo their expression levels were also tested in two murine models of amyotrophic lateral sclerosis: SOD1G93A and hFUS+/+, characterized by progressive muscle atrophy. During C2C12 differentiation, linear and circular SMOX show the same trend of expression. Interestingly, in atrophy circSMOX levels significantly increased compared to the physiological state, in both in vitro and in vivo models. Our study demonstrates that SMOX represents a new player in muscle physiopathology and provides a scientific basis for further investigation on circSMOX RNA as a possible new therapeutic target for the treatment of muscle atrophy.

The male reproductive accessory glands of the blister beetle Meloe proscarabaeus Linnaeus, 1758 (Coleoptera: Meloidae): Anatomy and ultrastructure of the cantharidin-storing organs.

Blister beetles owe their name to their ability to release cantharidin, a blistering terpene, the highest concentration of which is retained in male accessory glands. The anatomy and ultrastructure of the three pairs of male reproductive accessory glands and the glandular region of the two vasa deferentia of Meloe proscarabaeus were investigated using light, electron and ion beam microscopy. All of the mesodermal glands here analysed share a common structural organization with an outer muscular layer and an inner glandular epithelium facing a broad lumen in which the secretory products are released. Developed rough endoplasmic reticulum, Golgi systems, abundant mitochondria, numerous secretory vesicles and a microvillated apical membrane are commonly found in the cells of different glandular epithelia, suggesting that all accessory gland pairs as well as the vasa deferentia are involved in an active synthesis. Nevertheless, each pair of glands appears specialized in the production of a specific set of substances, as suggested by the peculiarities in cellular ultrastructure and by the different aspect of the secretions stored in their glandular lumen. The above cited features of male accessory glands of M. proscarabaeus are compared with those of other beetles and some hints on their potential role in producing and/or concentrating cantharidin are provided.

Epileptic seizures and oxidative stress in a mouse model over-expressing spermine oxidase.

Several studies have demonstrated high polyamine levels in brain diseases such as epilepsy. Epilepsy is the fourth most common neurological disorder and affects people of all ages. Excitotoxic stress has been associated with epilepsy and it is considered one of the main causes of neuronal degeneration and death. The transgenic mouse line Dach-SMOX, with CD1 background, specifically overexpressing spermine oxidase in brain cortex, has been proven to be highly susceptible to epileptic seizures and excitotoxic stress induced by kainic acid. In this study, we analysed the effect of spermine oxidase over-expression in a different epileptic model, pentylenetetrazole. Behavioural evaluations of transgenic mice compared to controls showed a higher susceptibility towards pentylentetrazole. High-performance liquid chromatography analysis of transgenic brain from treated mice revealed altered polyamine content. Immunoistochemical analysis indicated a rise of 8-oxo-7,8-dihydro-2'-deoxyguanosine, demonstrating an increase in oxidative damage, and an augmentation of system xc- as a defence mechanism. This cascade of events can be initially linked to an increase in protein kinase C alpha, as shown by Western blot. This research points out the role of spermine oxidase, as a hydrogen peroxide producer, in the oxidative stress during epilepsy. Moreover, Dach-SMOX susceptibility demonstrated by two different epileptic models strongly indicates this transgenic mouse line as a potential animal model to study epilepsy.

Polyamines: The possible missing link between mental disorders and epilepsy (Review).

Polyamines are small positively charged alkylamines that are essential in a number of crucial eukaryotic processes, like normal cell growth and development. In normal physiological conditions, intracellular polyamine content is tightly regulated through a fine regulated network of biosynthetic and catabolic enzymes and a transport system. The dysregulation of this network is frequently associated to different tumors, where high levels of polyamines has been detected. Polyamines also modulate ion channels and ionotropic glutamate receptors and altered levels of polyamines have been observed in different brain diseases, including mental disorders and epilepsy. The goal of this article is to review the role of polyamines in mental disorders and epilepsy within a frame of the possible link between these two brain pathologies. The high comorbidity between these two neurological illnesses is strongly suggestive that they share a common background in the central nervous system. This review proposes an additional association between the noradrenalin/serotonin and glutamatergic neuronal circuits with polyamines. Polyamines can be considered supplementary defensive shielding molecules, important to protect the brain from the development of epilepsy and mental illnesses that are caused by different types of neurons. In this contest, the modulation of polyamine metabolism may be a novel important target for the prevention and therapeutic treatment of these diseases that have a high impact on the costs of public health and considerably affect quality of life.

Link between spermine oxidase and apoptosis antagonizing transcription factor: A new pathway in neuroblastoma.

Neuroblastoma (NB) is a heterogeneous extra­cranial childhood type of cancer, responsible for approximately 15% of all paediatric cancer­related deaths. Although several critical genetic aberrations have been related to NB, only a few established molecular markers have been associated with prognosis [V­myc avian myelocytomatosis viral oncogene (MYCN) locus amplification, deletions of part of chromosome 1p, 11q23 and gain of 17q]. Regrettably, direct evidence of NB­related tumour suppressors or oncogenes has not been currently identified at these chromosomal regions. MYCN locus amplification is present in approximately 20­30% of cases and is associated with a poor clinical outcome, representing the most important genetic prognostic marker. The functional guidelines for the prognosis of NB identify high­risk patients (<40% survival probabilities), but fail to identify patients at low and intermediate stages of the disease, which remains an issue to be resolved in NB. It has been shown that in NB cell lines and in a total­spermine oxidase (SMOX) transgenic mouse model, SMOX overexpression induces cellular stress via reactive oxygen species (ROS) imbalance. In this study, we demonstrated that the high expression level of the cytoprotective gene, apoptosis-antagonizing transcription factor (AATF), was driven by SMOX gene overexpression in both NB cells and Total­SMOX mice. The anti­apoptotic effect of AATF was supported by analysing the inhibition of the expression of the pro­apoptotic genes, BAX, BAK and PUMA, which were decreased, in both the in vitro and in vivo SMOX overexpressing model systems investigated. On the whole, this study supports the hypothesis that the SMOX gene can be considered as a novel anti­apoptotic marker in NB.

Exploring the activity of polyamine analogues on polyamine and spermine oxidase: methoctramine, a potent and selective inhibitor of polyamine oxidase.

Fourteen polyamine analogues, asymmetric or symmetric substituted spermine (1-9) or methoctramine (10-14) analogues, were evaluated as potential inhibitors or substrates of two enzymes of the polyamine catabolic pathway, spermine oxidase (SMOX) and acetylpolyamine oxidase (PAOX). Compound 2 turned out to be the best substrate for PAOX, having the highest affinity and catalytic efficiency with respect to its physiological substrates. Methoctramine (10), a well-known muscarinic M2 receptor antagonist, emerged as the most potent competitive PAOX inhibitor known so far (Ki = 10 nM), endowed with very good selectivity compared with SMOX (Ki=1.2 µM vs SMOX). The efficacy of methoctramine in inhibiting PAOX activity was confirmed in the HT22 cell line. Methoctramine is a very promising tool in the design of drugs targeting the polyamine catabolism pathway, both to understand the physio-pathological role of PAOX vs SMOX and for pharmacological applications, being the polyamine pathway involved in various pathologies.