• Istituto di Biologia e Patologia Molecolari


WELCOME TO THE IBPM NeuroNet CONSORTIUM: A network to investigate the molecular mechanisms of neuro-pathologies

“In the 19th century and at the beginning of the 20th, brain research belonged to many different areas that differed in methodology and targets: the morphological, the physiological and the psychological. The latter used to consider the brain as a black box where only the input and output were known but not at all the neuronal components and the way they interact with each other. At the beginning of the third millennium, due to prolonged ageing, neurodevelopmental disorders are growing and a much deeper knowledge of the brain is necessary.” Rita Levi Montalcini

The brain is the most fascinating as well as most complex organ of the human body, with over 200 billion neurons and several hundred trillion of contacts, the synapses. This huge network of cells receives, processes, and executes cognition and perception, controls motor activities, and stores memories over decades. Stimuli that disrupt the homeostasis of cellular processes, or alter the neuronal networks of the central nervous system (CNS), can originate neurodevelopmental, degenerative, or psychological disorders. 
Neurological disorders, affecting both the peripheral and central nervous systems, make up a large variety of diseases, the best known of which include:
• Alzheimer’s and related dementias, 
• Huntigton chorea, 
• Parkinson disease, 
• multiple sclerosis, 
• amyotrophic lateral sclerosis, 
• muscular dystrophies, 
• autism spectrum disease, 
• epilepsy, 
• neuroinfections, 
• brain tumors, 
• cerebrovascular diseases including stroke, 
• migraine and other headache disorders, 
• trauma- and malnutrition-related disorders. 
Concomitant with the longer human life span, neurological diseases are now ranging among the greatest challenges biomedical sciences are faced with. 

We have established the IBPM NeuroNet consortium to strengthen the interactions and synergies among IBPM scientists, with the understanding that diversified approaches are essential to clarify how the loss of control of these mechanisms drives neurological disorders.  
NeuroNet was launched at the Annual IBPM meeting 2018 “Health, disease and environmental research: biology, tools and applications” 
THE NeuroNet CONSORTIUM AbstractPF.pdf

Our background
Researchers of IBPM NeuroNet have diversified backgrounds: genetics and bioinformatics; crystallography and structural biology of disease-relevant protein and macromolecular complexes; transcriptional, epigenetics and post-transcriptional regulatory mechanisms; engineering of experimental systems for disease modelling and therapeutic testing; molecular mechanisms of cell differentiation, developmental and signalling pathways. 
The consortium integrates diversified competencies, and offers the use of diversified in vitro and in vivo experimental systems, including 
• ad-hoc engineered cell lines, 
• stem cell lines,
• differentiating systems, and 
• model organisms

Our goals
- identify macromolecules (DNA, RNA or proteins) that may represent novel potential diagnostic/prognostic biomarkers and targets in neuro-pathologies
- design and develop innovative therapeutic strategies aimed at preserving and restoring brain functions
- identify early stage biomarkers for CNS cancer and diseases
Our studies will also set a productive framework to expand our knowledge on two biological conditions, i.e. stress and inflammation, that are receiving increasing attention as major contributors to a number of diseases. 
Four min research areas
-  neurooncology, 
-  neuro-degenerative and neuromuscular diseases, 
-  neurodevelopment and its disorders, 
-  design of innovative diagnostic and therapeutic strategies. 
The IBPM Neuronet founding group
Neuronet Organization  - Contacts
c/o Dept. of Biology and Biotechnology, Sapienza University, Rome

RESEARCH AREA 1: Neuro-oncology  
Brain tumors are among the most difficult types of cancer to treat. Survival has barely increased over the last 40 years. There is an urgent need for improved diagnostic biomarkers and innovative therapeutic strategies for neurological tumors of both children and adults. 
IBPM researchers are exploring the structural and functional biology of relevant proteins, as well as RNA-based and epigenetic mechanisms regulating gene expression, to shed light on how deregulation of these molecules, either per se or as components of complex regulatory networks, may impact on brain tumors. 
The research focusses on medulloblastoma, a highly malignant paediatric tumor, and glioblastoma, the most prevalent and lethal brain cancer in adulthood. The common aim is to develop effective therapeutic strategies based on targeted inhibition of critical molecules or circuit components. 

Abstracts of research projects

1.1.1 GBM:Epigenetic regulators and small signaling molecules as therapeutic targets for GBM (Illi B. Salvatori L.-IBPM)

Glioblastoma multiforme (GBM), is still the most devastating and incurable brain tumour in the adult. The oncoprotein Myc is highly expressed in a wide variety of tumours, including GBM. In particular, it is required for the maintenance of the stem properties of Glioblastoma Stem Cells (GSCs), responsible for tumour recurrence and resistance to radio- and chemotherapy. The main focus of this research line is the identification of novel Myc post-translational modifications which may act as a switch to control Myc functions. Ongoing experiments show that Myc is both asymmetrically and symmetrically dimethylated by the protein arginine (R) methyltransferases PRMT1 and 5 respectively, being symmetric dimethylation typical of GSCs in their stem conditions, while asymmetrically dimethylated Myc is found in differentiating and less aggressive GSCs. Therefore, we believe that the modulation of differentially modified forms of Myc may control its deregulated activity during glioblastomagenesis and/or GBM progression. Moreover, the functional interaction between Myc, PRMT5 and PRMT1 will be investigated also at genomic level, in terms of chromatin co-occupancy and transcriptomic outcome.
Prof. Mai A. (Dept. of Chemical Pharmaceutics, Sapienza University Rome)
Dr. Nanni S. (Institute of Medical Pathology, Università Cattolica, Rome)

1.1.2 GBM: Effect of Nitric Oxide in glioblastoma stem cells (Salvatori L Illi B:-IBPM)

Glioblastoma (GBM) is the most common malignant brain tumor. It features local cancer stem cells (glioblastoma stem cells, GSCs) responsible for enhanced resistance to therapies and tumor recurrence. One potential approach to eradicate GSCs is to force these cells to undergo terminal differentiation. Therefore, the role of differentiation-inducing drugs is receiving increasing attention. Nitric oxide (NO) is an intercellular and intracellular signalling molecule in the brain and is involved in neural development. It plays also key roles in GBM pathophysiology as it is implicated in induction of apoptosis, radio- and chemosensitization. On these bases, we are currently exploring the effect of NO on patient-derived GSCs biological properties. In particular, we aim to understand the molecular mechanisms underlying NO influence on the maintenance of GSCs subpopulation and its pro-differentiation ability.

1.1.3 GBM: Neuroglobin a protein necessary for cell protection in hypoxia condition (Savino C.-IBPM)

Neuroglobin (Ngb) is expressed in brain and is involved in protection of neurons from hypoxia and is likely to be a sensor of hypoxia in cells to trigger neuroprotection, through involvement in anti-apoptotic or anti-oxidant pathways. Nevertheless, its functional role in the cell is not completely understood and it seems to be involved in several interactions with other proteins in different catalytic/signalling pathways. Moreover, Ngb is expressed in glioblastomas and is especially found in tumor regions adjacent to necrosis. Ngb structure is peculiar: it contains a large internal cavity and displays larger conformational transition upon ligand binding. Detailed analysis of the role of the structural elements determining its dynamics will provide a sounder basis to unravel its still unclear physiological function and mechanism of action. This goal will be achieved using the following techniques: X-ray crystallography, Time resolved X-ray crystallography (TR-XRD) and Time resolved Small and Wide angle X-ray Scattering (TR-SWAXS).

1.1.4 GBM: Sorcin a protein involved in MultiDrug Resistance (Ilari A., Colotti G.-IBPM)

Sorcin belongs to the penta EF-hand (PEF) protein family; as other members of this family, upon calcium binding, Sorcin becomes able to interact with binding partners as Ryanodine Receptors (RyRs). Sorcin is one of the most expressed calcium binding proteins in many brain cancers and is considered a histological marker for malignant glioma. Sorcin plays also an important role in multidrug resistance (MDR) in tumors, since its expression confers resistance to doxorubicin and to other chemotherapeutic drugs. In order to overcome the MDR in brain cancer, one of the possible strategy could be to inhibit the mechanisms of MDR induced by sorcin. For this purpose we aim at identifying peptide able to bind sorcin with high affinity thereby inhibiting the sorcin induced MDR.
Dr. Fazi F. (Sapienza University)
Dr. Calì T. (Padua University)

1.1.5 GBM: Mesenchymal stem/stromal cells as a possible anti-angiogenetic approach for glioblastoma treatment (Falchetti ML-IBCN)

Glioblastoma (GBM), the most aggressive astrocytic tumor of the adult, is characterized by a prominent angiogenesis. Mesenchymal stem/stromal cells (MSCs) are a very attractive therapeutic opportunity due to two main peculiarities: the innate tropism for tumor lesions and the ability to uptake and release huge amounts of drugs. Therefore, MSCs can be loaded with chemotherapeutic drugs and used as a Trojan horse for a specific drug delivery to tumors. Moreover, considerable attention to the effects of MSCs secretome on the tumor microenvironment has recently gained considerable attention. We previously demonstrated that MSCs of human origin, derived from bone marrow as well as from adipose tissue, exert an anti-tumorigenic effect on GBM growth in murine xenograft models of orthotopic tumors. MSCs were effective both on the tumor bulk, as we modelled by engrafting the U87MG tumor cell line, and on patient-derived cancer stem cells (CSCs), which represents the tumor cell subpopulation responsible for tumour recurrence and resistance to therapy. Paclitaxel loading didn’t significantly enhance the MSCs anti-tumor activity, suggesting that MSCs per se are responsible for the observed impairment of tumorigenicity. Now, we plan to address the effectiveness of MSCs delivery on neoangiogenesis. We will compare the effectiveness of adipose tissue, bone marrow and placental tissue-derived MSCs. Moreover, the tumor tropism and effectiveness on tumorigenicity of MSC-derived microvesicles will be addressed as well.

1.1.6 GBM: Analysis of new molecular and cellular biomarkers and of natural tumor-selective cytotoxic drugs (Dell’Albani P. - IRIB)

Glioblastomas (GBM) are the most aggressive and deadliest brain tumours. Their infiltrative nature and the existence of cancer stem cells, able to relapse new glioma foci, highlight the need for the identification of new and cell-specific biomarkers that could be used as therapeutic targets also. The analysis of Notch receptors among the four grade glioma, both in freshly resected gliomas and in primary cultures, has highlighted a high expression of Notch-4 in GBM and low levels of Notch-1. Furthermore, the use of two Quercetin-derivatives has revealed cell-specific toxicity against glioma cells, while normal astrocytes and fibroblasts had very limited effects. Since GBMs are very heterogeneous neoplasms, future research should combine recent advances and try to target specific biomarkers, as key molecules, or disrupt their role in signalling pathways activated.
Department of Neurosciences, University of Catania, Catania, Italy;
Department of G.F. Ingrassia, Section of Anatomic Pathology, University of Catania, Catania, Italy;
Dep of Pharmaceutical Sciences, Section of Biochemistry, University of Catania, Italy.

1.2.1 MB: Noncoding RNA in brain tumors (Caffarelli E., Laneve P.-IBPM)

The main focus is on the role of the noncoding RNAs (microRNAs, long noncoding RNAs, circular RNAs) as crucial components of the regulatory networks orchestrating neuronal differentiation programs in mammals. The final aim is to understand how their deregulation may impact on brain tumors. In the last few years, we identified specific neuronal microRNAs (miRNAs) crucial for cell decision between proliferation and differentiation, and new molecular circuitries in which they function as potential onco-suppressors. According to their relevant role, their expression is significantly deregulated in primary brain tumors. Long noncoding RNAs (lncRNAs) are highly tissue-specific drivers of cancer phenotypes: we are studying novel lncRNAs involved in neuronal differentiation and specifically deregulated in medulloblastoma. One of them, functioning as a microRNA sponge, controls the expression of four specific Group 4 medulloblastoma driver genes, representing a novel biomarker and a potential therapeutic target for this enigmatic class of tumors.
Prof. Ferretti E. (Dept of Molecular Medicine Sapienza University, Rome)

1.2.2 Medulloblastoma: new therapy with the chemokine Cxcl3, targeting the migration of neoplastic precursor cells (Tirone F., Ceccarelli C., Micheli L., D’Andrea G. IBBC)

Our laboratory studies the development of the cerebellum, in normal and in the pathological conditions leading to medulloblastoma, the cerebellar tumor. Medulloblastoma is the most aggressive pediatric tumor and about 30% of medulloblastomas arise from cerebellar granule cells (GCPs) undergoing transformation, following activation of the Shh pathway. We have identified two genes, Tis21/Btg2 and Btg1, which regulate the proliferation and the migration of normal and neoplastic GCPs, and we demonstrated that both genes are medulloblastoma suppressors. We have observed that the chemokine Cxcl3, target of Tis21, promotes the migration of the GCPs outside the proliferating area of the cerebellum, inducing their differentiation and the arrest of tumorigenesis. We are studying this new therapy in Shh-type medulloblastoma models generated by us.
Dr. Angela Mastronuzzi, Hospital Bambino Gesù, Rome

1.3 The potential role of the secreted vesicles as biological regulator in the pathogenesis of Neurofibromatosis type I and II (Citrigno L. IRIB)

Neurofibromatosis (NF) type 1 is a condition characterized by changes in skin coloring (pigmentation) and the growth of tumors along nerves in the skin, brain, and other parts of the body. The signs and symptoms of this condition vary widely among affected people. Non‐coding RNA (ncRNA) species have emerged in as molecular fingerprints and regulators of tumor’s growth, pathogenesis and progression. Exosomes‐mediated vesicular export processes reduce intracellular levels of specific ncRNA in EV donor cells while creating a pool of EV‐associated ncRNA in the extracellular space and bio fluids that enables their uptake by other recipient cells; both aspects have functional consequences. Using a Next Generation Sequencing approach, we want to determine the presence and the differences of mRNAs, miRNAs, and lncRNAs within exosomes in patients affectedby different forms of NF
Hussman Institute for Human Genomics, Miami FL; Cedar Sinai Medical Center – Los Angeles, CA


RESEARCH AREA 2: Neurodegenerative and neuro-muscular diseases
Neurodegeneration in the central nervous system involves the loss of neuronal structure and function. Neurodegeneration is observed after viral insults, mostly in the so called ‘neurodegenerative diseases’ generally occurring with ageing, such as Alzheimer's and Parkinson's disease, but also multiple sclerosis and amyotrophic lateral sclerosis. All negatively affect mental and physical functioning. 
The common goal of researchers is to understand how alterations in specific signalling pathways are responsible for neuronal death, a common feature of these diseases. 
Studies are carried out using biochemistry, genetics and cell biology approaches as well as high-throughput transcriptomics and proteomics strategies.
Abstracts of research projects

2.1.1 AD: Impairment of protein homeostasis (Foppoli C.-IBPM)

Disturbance of components of the proteostasis network, that provides a critical protective role against stress conditions, may trigger neuronal death. In Alzheimer’s Disease (AD), because of increased oxidative stress, the occurrence of oxidation/dysfunction of specific members of the protein quality control, regulating protein folding, surveillance and degradation has been shown. We aim at identifying the specific pathways involved in the proteostasis network, whose alteration could contribute to the AD development. To this purpose, proteomic techniques will allow to identify specific proteins with altered expression levels or post-translational modifications in AD brain, that could be related to pathways associated with neurodegeneration.
Perluigi M., Di Domenico F. (Department of Biochemical Sciences, Sapienza University Rome).

2.1.2 AD and MCI: Genetic factors in neurodegenerative diseases (Scarabino D.-IBPM)

The main interest is to investigate susceptibility genes for age-related complex neurodegenerative diseases, in particular Alzheimer's disease (AD) and Mild Cognitive Impairment (MCI), which prevent the achievement of advanced age. Alzheimer's disease (AD) is a neurodegenerative disease associated with a drastic decline in life expectancy. Along with the AD, an additional "clinical phenotype", namely Mild Cognitive Impairment (MCI), has been identified, characterized by memory alterations in subjects with normal global cognitive function. Sporadic AD is a complex disease whose onset depends on the interaction of environmental factors and numerous susceptibility genes. Our recent data have shown that some polymorphisms of the telomerase genes TERC and TERT are involved in susceptibility to AD, confirming the relationship between the onset of AD / telomerase / telomere length. We investigate also the role of APOE gene on AD susceptibility, the subjects were genotyped for apoliprotein E, and plasma ApoE was assayed. The aim of the project was compared the distribution of genetic and biochemical markers observed in subjects with worsening of cognitive status and progression to AD, with subjects with persistence over time of MCI, in order to highlight predicting markers for AD.
Corbo R.M. (Department of Biology and Biotechnology, Sapienza University)
Gambina G., Brogio E. (Alzheimer's Disease Center, Department of Neuroscience, University and Hospital of Verona)
Businaro R. (Department of Medico-Surgical Sciences and Biotechnologies, Sapienza University)
Maida C., Gaudio M.R. (S. Giovanni –Addolorata Hospital, Rome)
Mantuano E., Veneziano L. (CNR Institute of Translational Pharmacology (IFT))

2.1.3 AD: miR-101 as a regulator of APP and other genes associated to AD in hippocampal neurons. (Ruberti F. Barbato C-IBCN)

Our group is studying the function of specific microRNAs engaged in the regulation of gene targets and pathways in neuronal cells using lentiviral vectors expressing miRNAs or miRNA sponge inhibitors. Amyloid precursor protein (APP) and its metabolites are associated to Alzheimer’s Disease (AD). MicroRNAs are also involved in AD pathogenesis. We have identified miR-101 as a regulator of APP and other genes associated to AD in hippocampal neurons. By using primary hippocampal neurons and murine models, we are currently studying miRNA-regulated networks implicated in learning and memory impairment, and in neurodegeneration

2.1.4 AD: metabolic disorders, nutraceutical (Di Carlo M.,Nuzzo D.,Picone P.,Galizzi G.(IRIB)

Studies on the effect of diet-induced obesity as risk factor for the onset of neurodegenerative diseases (ND), including Alzheimer’s disease (AD). The impact of nutrition and dietary constituent on ND are investigated by using a mouse model (C57BL/6) fed with high fat diet (HFD). The systemic and central metabolic conditions such as insulin resistance, are evaluated. Diet-induced stress and inflammation conditions associated to neurodegeneration are assessed by using specific assays and biomarkers. Expression of proteins involved in AD are also investigated. The effects of the addition of food with high content of bioactive components, including polyphenols, are explored as potential nutraceutical approach for preventing and/or reducing both metabolic and neurogenerative disorders.
Prof. Flavia Mulè and Prof. Antonella Amato Department of Biological, Chemical and Pharmaceutical Sciences and Technologies (STEBICEF), Palermo University

2.2.1 Investigating ncRNA roles in ALS pathogenesis (Caffarelli E., Laneve P.-IBPM)

The main interest is on the ALS-linked RNA binding proteins TDP-43 and FUS. They widely control RNA metabolism and make ALS an RNA disorder. Transcriptome analysis, carried out on FUS-ALS murine motoneurons, unveiled different classes of RNAs (microRNAs, long noncoding RNAs and circular RNAs) affected by FUS-ALS mutations and allowed the construction of regulatory networks relevant for the pathology.
Prof. Bozzoni I. (Department of Biology and Biotechnology, Sapienza University Rome)

2.2.2 ALS Drosophila models (Cestra G.-IBPM)

Drosophila melanogaster has emerged as a valuable in vivo system in the study of ALS. Flies carrying mutations in genes essential for motor neuron survival show reduced life span and defective locomotive behaviour. We are currently involved in different research projects on ALS in fly:
1) the identification of novel interactors of FUS and analysis of their in vivo role in motor neuron function;
2) to study the in vivo effect of different chemical chaperons, which exhibit reverse aggregation of misfolded proteins, on different Drosophila models of ALS-mediated proteinopathy;
3) to collaborate in the identification and characterization of the mRNAs with altered nuclear-cytoplasm distribution in Drosophila models of C9orf72-mediated ALS.

2.2.3 ALS: Aberrant H2S metabolism (Giuffrè A.-IBPM)

Hydrogen sulfide (H2S), the third gaseous signaling molecule along with nitric oxide (NO) and carbon monoxide (CO), exerts neuroprotective antioxidant, anti-inflammatory and anti-apoptotic effects at physiological concentrations, while leading to toxic impairment of energy metabolism at higher levels. Aberrant H2S metabolism was interestingly found to be implicated in the pathogenesis of several neurodegenerative diseases. Namely, H2S levels proved to be pathologically low in Alzheimer’s and Parkinson disease, and abnormally high in the central nervous system of ALS patients. The project aims to gain molecular insights into H2S metabolism and its regulation by investigating the human enzymes responsible of endogenous H2S production and catabolism, studied both as isolated proteins and in cellular models of ALS. The ultimate goal of this research is to develop innovative therapeutic interventions based on the pharmacological modulation of H2S levels by natural or synthetic modulators of H2S biosynthesis/catabolism.
Vicente J.B. (ITQB, New University of Lisbon, Portugal)
Forte E. (Department of Biochemical Sciences, Sapienza University Rome)
Saso L. (Department of Physiology and Pharmacology, Sapienza University of Rome)

2.2.4 Role of endothelin 1 in motor neuron degeneration: a possible therapeutic target for amyotrophic lateral sclerosis (D’Antoni S., Catania M.V. IRIB)

Evidence indicates that factors released by activated astrocytes may contribute to motor neurons (MNs) degeneration. We have found that the expression of endothelin-1 (ET-1), a vasopeptide produced in the CNS by astrocytes and microglia, is increased in reactive astrocytes in the spinal cord of SOD1-G93A mice and ALS patients. Importantly, ET-1 is toxic for MNs in an in vitro model of mixed spinal cord cultures enriched with reactive astrocytes. The toxic effect of ET-1 on cultured MNs is associated with a reduced activation of the PI3K pathway and requires NO synthesis. Our study suggests that the modulation of ET-1 signaling could be a therapeutic strategy to slow down MN degeneration in ALS.
Our mixed spinal cord cultures can be used to validate other potential targets for ALS.
Dr. Eleonora Aronica, Dept of (Neuro) Pathology, Academic Medical Center, Amsterdam, The Netherlands; Dr. Patrizia Longone and Dr. Alida Spalloni, Molecular Neurobiology Unit, Experimental Neurology, Fondazione Santa Lucia, Rome

2.2.5 Mechanisms of neuroinflammation in ALS pathogenesis and the role of microRNAs (Parisi C., IBBC)

Neuroinflammation is a feature of ALS and RNA dysregulation a mechanisms related to the pathology. We study how microRNA-mediated post-transcriptional regulation affect neuroinflammation in ALS focusing on microglia in the SOD1-G93A mouse. We revealed that termination of A20 function by miR-125b prolongs the activation of NF-κB in ALS. Given the known NF-κB involvement in ALS we are extending our analysis to other pathway components and to human ALS models.
Dr. Roberto Rizzi ITB, CNR/INGM Milan, IT
Dr. Claudia Bearzi IBBC, CNR/INGM Milan, IT
Dr. Sara Marinelli IBBC, CNR Rome, IT
Dr. Tommaso Mazza, CSS Mendel Rome, IT

2.2.6 Effects of voluntary exercise in the SOD1G93A low copy mouse model of ALS: sex differences, metabolism, inflammation (Mandillo S.,Golini E., IBBC)

ALS is a fatal disease in which the role of environmental factors and their interactions with affecting genes are not well understood. Effective treatments are lacking and the need of new therapies and early diagnostic tools is urgent. In our projects we use the classical SOD1G3A high-copy transgenic mouse model and the less studied SOD1G3A low-copy line. The latter gives the opportunity to examine the very early stages of pathology and allows a longer therapeutic window to test new interventions (Mandillo et al., 2014; Garbugino et al., 2018).
We focus on the behavioral aspects of ALS to elucidate its neurobiological mechanisms in reference to:
1) Influence of environmental factors (physical activity) on onset, progression and survival
2) Potential pharmacological treatments to alleviate symptoms : Preclinical validation of candidate drug target for ALS
3) Detection of early behavioral pathological signs via automated monitoring systems : Home cage monitoring using the DVC Tecniplast system: validation in ALS mouse models
Sara Marinelli, IBBC
Prof. Antonio Musarò, Università Sapienza Roma
Sebastiano Cavallaro, IRIB-Catania
Fabio Mammano, IBBC Associate
Marcello Raspa, Ferdinando Scavizzi, IBBC
Fabio Ianniello, Tecniplast, Varese

2.3 Juvenile Hungtinton Disease: Rhes, a key protein in motor and cognitive impairments (Ilari A., Morea V., Colotti G.-IBPM)

Human Rhes (Ras-homolog enriched in striatum) has SUMO-E3 ligase function and among its targets include mutated huntingtin (mHtt) which upon sumoylation becomes more soluble and hence more cytotoxic. mHtt is the well-known cause of Huntington disease (HD), an inherited neurodegenerative disease. The mutation consists in a >35 CAG repeats in exon 1 of the Htt gene. The expansion of the CAG repeats stretch makes mHtt very unstable and able to aggregate with itself and/or other proteins interfering with different metabolic pathways. In addition to its catalytic activity, Rhes exerts a role in autophagy by activating autophagy inhibitor mTOR and by directly interacting with autophagy activator Beclin-1. Rhes bidirectional regulation of autophagy induction, together with the SUMO-E3 ligase activity on mHTT and specific expression in corpus striatum, make Rhes an attractive target for HD treatment. We aim at determining Rhes three-dimensional structure and at identifying its interactors in human cells.

2.4 Hereditary Spastic Paraplegia HSP: Mechanisms underlying spastin regulation (Rinaldo C.-IBPM)

HSP neurodegenerative disease: the most common type is due to heterozygous mutations in spastin gene. Precise levels of spastin appear crucial for its biological functions. A gene-dosage rescue of neurite defects in HSP patients’ neurons has been recently reported, stressing the relevance of studying the mechanisms that regulate spastin to develop therapeutic approaches. The aims of this research project is investigate the molecular mechanism underlying spastin regulation mediated by the kinase HIPK2 and evaluate whether manipulating this pathway is possible to restore spastin physiological levels and rescue pathological phenotype in neurons from HSP patients

2.5 Motor Neuron Disorders MNDs: The role of RNA-binding proteins in synaptic plasticity (Onori A., Passananti C., Pisani C., Corbi N.-IBPM)

Transcription, mRNA localization and translation are key events at the synaptic region, determining proper neuron/neuron and muscle/neuron communication. In this context, one of our research interest is to understand how RNA-related proteins, such as the survival motor neuron protein SMN, the apoptosis antagonizing transcription factor AATF/Che-1 and the master gene NF-κB operate within cellular networks implicated in the RNA processing, trafficking and in local translation. We are also studying the potential involvement of AATF in memory-related events in which NF-κB is known to play a fundamental role. Defects in RNA metabolism have been linked to a variety of neuropathological conditions including intellectual disabilities and motor neuron diseases (MNDs), while memory deficits correlate with severe neurodegenerative disease, such as Alzheimer’s disease.These studies will provide new insight into molecular and cellular mechanisms underlying neuronal development, synaptic plasticity and cognitive processes, as well as to develop therapeutic strategies for severe neurological and neuromuscular diseases.
Di Certo M.G., Gabanella F. Institute of Cellular Biology and Neurobiology (IBCN) CNR, Rome.
Prof. Romano A., Dr. Freudenthal R. (University of Buenos Aires UBA/CONICET)

2.6.1 Glucocorticoids as regulatory signals in neuronal functions in DMD (Fragapane P.IBPM)

Stressful events induce activation of the autonomic nervous system and the hypothalamus-pituitary-adrenal axis, with increase in glucocorticoid (GC) synthesis and release. The ubiquitous expression of GC receptors (Gr) confer to this system an essential role in the response to stress and restoration of homeostasis. These stress-induced changes are mediated by modifications in gene expression and protein synthesis. Precise regulation of the HPA (Hypothalamic-Pituitary-Adrenal) axis activity is very important for the organism; indeed, chronic exposure to GCs results in various adverse side effects, such as osteoporosis, diabetes, hypertension and neurodegeneration. The goal of this research project is to uncover the effect that GCs exert in brain regions susceptible to stressful stimuli as hippocampus, through gene expression analysis of the Gr and his target genes by administration of different glucocorticoid hormones. The study will be performed in vitro neuron cultures from wild-type and in mdx mice (a Duchenne Muscular Dystrophy animal model) and neuroblastoma cell lines administered with Dexamethasone and Cortisone. RNAs and proteins factors involved in this regulation will be the subject of this study.
De Stefano E., Camilloni G., Bozzoni I. (Department of Biology and Biotechnology, Sapienza University Rome)

2.6.2 The role of histone methyltransferases in the epigenetic regulation of neuronal and muscle stem cells in DMD (Mozzetta C.-IBPM)

Epigenetic regulation of gene transcription is crucial to shape the gene expression programs and cell fate choices in stem cells. Therefore, understanding how these processes are regulated both at the physiological and pathological level is paramount to devise strategies aimed to improve the regenerative capacity of organs affected by degenerative diseases, such as dystrophic muscles. Histone lysine methyltransferases (KMTs) are crucial chromatin-modifying enzymes that have been implicated in the maintenance of cell-specific transcriptional homeostasis in a variety of cell types, including neuronal and muscle progenitors. Among this class of enzymes, we are interested in studying the role of H3K9 KMTs in shaping higher ordered chromatin structures (i.e. nuclear-lamina associated domains) both during muscle and neuronal differentiation. More specifically, we aim to understand the specific transcriptional programs regulated by H3K9 KMTs in the different population of muscle progenitor cells involved in skeletal muscle regeneration and degeneration, with the final goal to elucidate the epigenetic control of their phenotypical plasticity in relation to the environmental changes imposed by DMD progression. Moreover, since H3K9 KMTs have been firmly implicated in the maintenance of neuronal transcriptional homeostasis, we aim also to understand how these enzymes influences the different fates of neuronal progenitor cells. Given the availability of small molecules inhibiting specifically the action of the different KMTs, our study will help to identify possible new pharmacological targets to be exploited to revert pathological epigenetic states associated with DMD.
Prof. Sigmar Stricker (Freie Universitat, Berlin)
Dr. Giovanna Peruzzi (IIT, Rome)
Prof. Stefano Biagioni (Department of Biology and Biotechnology, Sapienza University Rome)

2.6.3 Role of cell cycle regulators in the control of skeletal muscle metabolism and regeneration in DMD (Caruso M., Agnese Bonato A., IBBC)

In addition to their well-known function in cell proliferation, there is increasing evidence that cell cycle regulators play important roles in metabolic control. Skeletal muscles are composed of heterogeneous myofibers that differ in their contractile response to motor nerve action (slow or fast) and metabolism (oxidative or glycolytic). Recently, we observed that cyclin D3-/- mice display an increased number of oxidative myofibers and increased basal metabolism and muscle performance. Interestingly, oxidative fibers in patients with DMD are more resistant to the dystrophic pathology in comparison with glycolytic fibers.
Our objective is to evaluate whether inactivation of cyclin D3 in the mdx mouse model of DMD can attenuate the dystrophic pathology by promoting a slower, more oxidative muscle phenotype.

2.7 Possible role of p65 iso5, a new isoform of the NF-kB complex, in neurodegenerative diseases (Francesco Di Blasi IRIB)

Nuclear Factor kB (NF-kB) consist of a family of transcription factors that regulates the expression of genes involved in many processes as inflammation. Among the various NF-kB complex, the most characterized is the heterodimer p65/p50. NF-kB is ubiquitously expresses in neurons and when constitutively activated is associated with neuronal information. A dysregulation of NF-kB activity is related to neurodegenerative diseases. Synthetic glucocorticoids (Gcs) are the most frequently prescribed anti-inflammatory drugs, associated with a reduction in the AD risk. In our lab we have founded a new spliced form of p65, named p65 iso5. This protein is able to bind Dexamethasone and doubling glucocorticoid response GR-mediated. We hypothesize that this natural protein, could act synergistically with synthetic glucocorticoids, thus reducing the dosage and time therapy and consequently minimize synthetic glucocorticoids side effects.

RESEARCH AREA 3: Neurodevelopmental disorders
Neurodevelopmental disorders result in severe cognitive, neurological and/or psychiatric dysfunctions. They can be due to environmental or genetic causes. The latter involve mutations in genes involved in early neurogenesis, including cell migration and synaptogenesis, or in metabolic pathways. 
Adult neurogenesis, albeit limited to specific brain regions, consists in the capacity to generate new neurons throughout life and is associated with brain plasticity. This process, while being severely impaired in neurodegenerative diseases, where the capacity of the adult brain for cell renewal is compromised, represents an event of neuro-repair relevant in brain injury. 
Projects in this area address these aspects to understand the molecular and cellular mechanisms compromised in several pathologies. 
Abstracts of research projects

3.1. Congenital disorder of Glycosylation (CDG type II): A Drosophila model to study associated neurological defects (Frappaolo A., Sechi S., Giansanti M.G.-IBPM)

Congenital Disorders of Glycosylation (CDG) comprise a family of human diseases caused by mutations in genes required for the synthesis of glycoconjugates. More than 80% of these diseases display severe neurological impairment. The Conserved Oligomeric Golgi (COG) complex mediates tethering of vesicles carrying glycosylation enzymes across the Golgi cisternae. Mutations affecting human COG1, COG2, COG4-COG8 cause monogenic forms of inherited, autosomal recessive CDGs. Typical clinical manifestations of COG-CDGs include psychomotor delay, epileptic seizures and hypothonia. Yet, it is unknown how the glycosylation defects cause the neurological aspects of CDGs.
We have generated a Drosophila COG7-CDG model, which closely parallels the pathological characteristics of COG7-CDG patients, including pronounced neuromotor defects and altered N-glycome profiles. We have demonstrated that the COG complex cooperates with Rab1 and Golgi phosphoprotein 3, to regulate Golgi trafficking and that overexpression of Rab1 can rescue the locomotor defects associated with loss of Cog7. Our results suggest that the Drosophila COG7-CDG model can be used to test novel potential therapeutic strategies by modulating trafficking pathways.
Tiemeyer M. (Complex Carbohydrate Research Center, The University of Georgia, Athens, USA)
Fraschini R. (Università degli studi di Milano Bicocca)

3.2.1 Role of regulators on synapse formation and activity (Cestra G.-IBPM)

Synapse is a cell junction specialization responsible of directional communication between neurons. In the chemical synapse, the pre-synaptic neuron releases neurotransmitters into the cleft that, by binding to membrane receptors, elicits post-synaptic currents. Activity and dynamics of neurotransmitter receptors it tightly controlled by the elaborate network of adapter proteins of the postsynaptic density. The actin cytoskeleton extends throughout the post-synapse and controls its morphology and activity.
In our laboratory we are currently studying the role of the adapter protein named neuronal Arg Binding Protein 2 (nArgBP2), which interacts and is phosphorylated by the Arg/Abl kinase protein family. nArgBP2 is implicated in the regulation of actin dynamics at the post-synaptic specialization and it is involved in the formation of glutamatergic synapses. Since preliminary observations have suggests a possible interaction of nArgBP2 with proteins involved in ASD, such as FMRP, Shank3 and SAPAP3, we are currently investigating its possible implication in these pathways.

3.2.2 Role of noncoding RNA in ASD altered plasticity mechanisms (Mannironi C.-IBPM)

Presutti C., Rinaldi A., Mele A. (Department of Biology and Biotechnology, Sapienza University Rome)
Scattoni A.M., Ricceri L. (Higher Health Institute, Rome)

3.3 Molecular basis of neonatal epileptic encephalopathy (NEE): a severe neurological disease caused by defects in the vitamin B6 salvage pathway enzyme pyridoxine 5’-phosphate oxidase. (Tramonti A. and Nogués I. – IBPM-IBAF)

Vitamin B6 plays important roles in the metabolism of neurotransmitters such as glutamate, dopamine, serotonin, epinephrine, and gamma-aminobutyric acid. The vitamin derivative pyridoxal 5’-phosphate (PLP) is the cofactor of a plethora of enzymes, whose activity is of pivotal importance for central metabolism and is essential for the correct functioning of the central nervous system. The activity of these enzymes relies on an appropriate availability of PLP in the neuronal cells. Mammals are not able to synthesize PLP but they recycle it through a salvage pathway (in which the enzymes pyridoxal kinase and pyridoxine 5’-phosphate oxidase are key components) from the different B6 vitamers contained in food and from protein turnover. Once made available, PLP is somehow targeted to apo-PLP-enzymes. Deficiency of vitamin B6 has been implicated in pathologies such as autism, schizophrenia, Alzheimer, Parkinson, epilepsy and Down's syndrome.
The focus of our project is a severe, newly recognized and rare neurological disease known as neonatal epileptic encephalopathy (NEE), determined by autosomal recessive mutations in the gene encoding pyridoxine 5'-phosphate oxidase (PNPO), that result in inadequate levels of PLP. The main feature of the disease is the onset of severe seizures, often within hours from birth, which respond to PLP administration and, in some cases, also to pyridoxine. If not promptly treated, neonates will present a drug resistant epileptic status that might determine death. To date, 18 pathogenic mutations of the PNPO gene and about 40 patients have been reported in the literature. Recently, we have studied the functional effects of the c.347G>A (p.R116Q) mutation of the human PNPO gene, and discussed its pathogenic role in epileptic encephalopathy. Novel missense mutations have been found in children affected by NEE, but have yet not been characterized at a molecular level. We believe that a detailed structural and functional characterization of these mutant forms is very important for a full understanding of the disease and for the rational devise of treatment strategies.

3.4 The role of p21-lacking adult Neural Stem Cells in the neuro-regenerative responses following traumatic brain injury (Farioli Vecchioli S., IBCN).

Adult neural stem cells (aNSCs) play a pivotal role in maintaining a high rate of the adult neurogenesis in the neurogenic niches throughout life. In physiological conditions most of aNSCs are in the reversible G0 phase (also called quiescent state) and only rarely they are recruited in cell cycle to give rise to new neurons. In aging, the number as well as the neurogenic potentiality of aNCS progressively diminish until they disappear within the neurogenic niches. However, after brain injury, such as ischemic stroke or Traumatic Brain Injury (TBI), there is a strong increase in the activation and expansion of aNSCs, which actively participate in the neurorepair processes. The molecular mechanisms modulating the transition between quiescence and activation of aNSCs are very complex and are subjected of a wide range of studies aimed at enhancing the neurogenic potential of aNSCs to procrastinate brain aging and to increase the neuro-rigenerative responses following brain damage. In this context, our lab is involved in the study of the modulation of the gene p21, which represent one major regulators in the maintenance of the aNSCs quiescence. The purpose of our study is to specifically target the deletion of p21 in the aNSCs, with the goal to expand the NSC pool size and consequently enhance the adult neurogenesis in aging and after brain damage, in particular after TBI. To achieve these aims we developed a multi-approaches strategy including conditional cre-loxP models, adeno associated virus (AAV) specifically infecting aNSCs and containing interference sequences for p21 and the adjuvant proneurogenic action of physical activity.

3.5. Aging of neural stem cells: control by a network of cell cycle inhibitors (Btg1/2, p16Ink4a) and reversal by neurogenic stimuli (running, fluoxetine and diet polyphenols) (Tirone F., Micheli L.,D’Andrea G., Ceccarelli M. IBBC)

The continued production of new neurons during adulthood in the neurogenic niches (i.e., dentate gyrus of the hippocampus and subventricular zone) enhances the efficiency of episodic/associative memory, as shown by several laboratories including ours. We have found a network of cell cycle inhibitors (Tis21/Btg2, Btg1) that regulate quiescence and proliferation of stem and progenitor cells in adult and aging neurogenic niches. We have shown that neurogenic stimuli, i.e., running, antidepressant, diet, are able to counteract the reduced neurogenesis associated to aging, either physiological or induced by Btg1 knockout. Part of this network is also p16Ink4a, which we have shown to prevent during aging any exit from quiescence elicited by running; therefore, p16Ink4a protects the stem cells pool against depletion after stimulus during aging.

3.6 Postnatal early environment may affect the epigenome, stably improving or exacerbating behavioral and physiological phenotypes in mouse models of developmental disorders (D’Amato F.R., IBBC)

Early aversive environments (maltreatment, maternal separation, etc.) have deleterious effects on several aspect of offspring development. Few studies explore the effects of “positive” early environments. Environmental enrichment (social and physical) is usually applied to adults and young mouse models of different psychopathologies: positive temporary effects that reduce symptoms are usually detected, but they do not usually last for long time. Rearing newborns in enriched environment, when neural plasticity is maximal, might result in stable rescue of different psychopathologies. Due to infant immaturity at birth, social enrichment/deprivation can be perceived by pups (and mothers) even during vey early postnatal days and can affect later development. Molecular mechanisms responsible/associated to these altered phenotypes are investigated.
Prof. Marco Battaglia, CAMH, Toronto University
Prof. Rossella Ventura, Sapienza University, Rome
Dr. Bice Chini, IN CNR, Milano

3.7 Roles of non-coding RNAs in myotonic dystrophy type 1 (Falcone G., Cardinali B., Provenzano C., IBBC)

Myotonic dystrophy type 1 (DM1) is a neuromuscular disorder caused by an unstable (CTG)n repeat in the DM protein kinase (DMPK) gene. Expanded CUG-repeats have been demonstrated to be toxic per se, sequestering nuclear proteins and disrupting gene transcription and pre-mRNA alternative splicing. Non-coding RNAs such as microRNAs and circRNAs have been found to be important regulators of cellular physiology and pathology by using a variety of mechanisms. Our studies aim at identifying microRNAs and circRNAs that are dysregulated in muscles of DM1 patients and may contribute to the disease mechanisms. In addition, their potential to be used as biomarkers for disease staging and progression is being investigated. Key words: non-coding RNAs, microRNAs, circular RNAs, disease mechanisms, disease biomarkers
Fabio Martelli, IRCCS Policlinico San Donato, San Donato Milanese, Milan

3.8 Synaptic dysfunctions and potential therapies in Fragile X syndrome (Catania M.V., D’Antoni S. IRIB)

Our research is focused on pathophysiological mechanisms of Fragile X Syndrome (FXS) and other neurodevelopmental disorders characterized by Intellectual Disability and autism, with the final goal of identifying possible pharmacological treatments. We found that in the synapses of the Fmr1 KO mouse model of FXS, metabotropic glutamate receptors type 5 (mGlu5) are less coupled to the scaffolding/effector protein Homer, with consequences for the mobility of mGlu5 receptors and their functional interaction with the NMDA receptors. We provide also evidence that activation of serotonin 5-HT7 receptor agonists improves pathological phenotypes of the Fmr1 KO mouse, namely altered morphology of dendritic spines and audiogenic seizures susceptibility. We are currently extending our findings to other diseases by studying the expression of different receptors and scaffolding proteins in specimens from human brains.
Dr. Eleonora Aronica, Dept of (Neuro) Pathology, Academic Medical Center, Amsterdam, The Netherlands;
Dr. Sebastiano Musumeci, Dr. Corrado Romano, Dr. Carmela Maria Bonaccorso, Dr. Michela Spatuzza, Oasi Research Institute Troina (EN);
Prof. Ferdinando Nicoletti, Dr. Giuseppe Battaglia, Università La Sapienza, Roma - IRCCS Neuromed, Pozzilli (IS);
Dr. Barbara Bardoni, IPMC, CNRS UMR6097, Valbonne, France
Prof. Marcello Leopoldo, Dr. Enza Lacivita, Università di Bari

3.9 Neurodevelopmental rare diseases: potassium channels mutations and ncRNAs in Zimmermann-Laband and FHEIG syndromes. (Parisi C., IBBC)

Zimmermann-Laband and FHEIG syndromes are neurodevelopmental genetic rare diseases with overlapping clinical features. A high proportion of mutations reside on K+ channels. We work on the functional characterization of KCNH1 and KCNK4 mutations in human patients fibroblasts, focusing on primary cilium signalling through Hedgehog pathway and its regulation by microRNAs.
Prof. Viviana Caputo Dept of Experimental Medicine, Sapienza University/CSS Mendel Rome, IT

RESEARCH AREA 4.  Innovative diagnostic and therapeutic strategies
The development of new tools and technologies for early diagnosis and therapeutic treatments is a fast-growing research area that has the ambitious goal to improve clinical practice of neurological diseases and neurological damage. Meeting this need requires that multidisciplinary knowledge translates into clinical applications in order to identify new in vivo diagnostic/prognostic tools and develop techniques enabling accurate, non-invasive, early disease diagnosis, as well as prediction of response to therapy. This is expected to lead to improved health outcomes and, in the long run, to contribute to improved sustainability of the health care system. 
IBPM researchers are active in the development of new diagnostic and therapeutic tools for fighting neurological disorders and tumours of the central nervous systems.
Abstracts of research projects

4.1 Adeno associated viral delivery system targeting nervous system (Onori A., Passananti C., Pisani C., Corbi N.-IBPM)

Recombinant adeno-associated virus (rAAV) vectors are versatile tools for gene transfer to the nervous system and in particular for central nervous system (CNS). Recombinant AAVs exhibit important advantages, including: safety profile due to the non-pathogenic nature of their wild-type form, stable transgene expression in post-mitotic cells, low risk of insertion mutagenesis, minimal immune responses and capsid-dependent tissue-specific tropism. Over the last years, twelve natural serotypes and more than one hundred variants of AAVs have been detected and isolated from humans and other primates. In particular, specific capsid serotypes, including serotype "9" bypass the blood-brain barrier, raising the possibility of intravascular administration as a non-invasive delivery route to achieve widespread CNS gene expression. To further lower any possible host immune response the use of peculiar AAV serotypes can be combined with tissue specific promoter/enhancer regions. We have successfully designed, produced and tested several rAAV vectors containing different human tissue specific promoters/enhancers. We intend to develop additional rAAV vectors carrying human promoter/enhancer regions exhibiting high nervous system tissue specificity. The AAV vector administration has been tested in several clinical trials for motor neuron disorders, lysosomal storage disorders, neurotransmitter disorders, glycogen storage disorders, neurodegenerative disorders and eye disorders.
Di Certo M.G., Gabanella F. (Institute of Cellular Biology and Neurobiology (IBCN) CNR Rome)

4.2 DMD: Innovative therapeutic strategy for Duchenne Muscular Dystrophy by AAV-mediated delivery of Zinc Finger Artificial Transcription Factors (ZF-ATFs). (Onori A., Passananti C., Pisani C., Corbi N.-IBPM)

Up-regulation of the dystrophin-related gene "utrophin" is a promising therapeutic strategy for the treatment of Duchenne Muscular Dystrophy (DMD). In order to re-program the utrophin expression level in muscle, we engineered the artificial ZF-ATF named "JZif1" that targets and up-regulates utrophin promoter. JZif1, delivered by AAV viral vectors, induces remarkable amelioration of the pathological phenotype in dystrophic mice (mdx). The molecular mechanisms underlying ZF-ATF induced muscle functional rescue is partially explained by the ZF-ATFs positive impact on the neuromuscular Junction (NMJ). Our results candidate our ZF-ATFs as novel therapeutic molecules for DMD treatment.
Mattei E., Strimpakos G. Institute of Cellular Biology and Neurobiology (IBCN) CNR, Rome.
Fanciulli M., SAFU, Translational Research Area, Regina Elena National Cancer Institute, Rome.

4.3 Exosomes in the neurodegenerative diseases (Alzheimer’s and Parkinson’s diseases) (Fiorucci G.IBPM)

Extracellular vesicles (EVs) comprise a wide variety of membrane-limited vesicles released from cells. Exosomes are the best characterized EVs. Neuronal cells release exosomes and for this reason these EVs have been proposed as novel means for intercellular communication which takes part to the transmission of pathogenic proteins in the neurological diseases, such as Alzheimer’s and Parkinson’s diseases.
In both diseases exosomes can propagate the misfolded proteins, but there is also evidence that EVs can have a protective role in neurodegeneration. The possible effects of exosome-associated pathogenic proteins on the recipient cells are important: how these proteins are internalized and how they induce protein aggregation and neurodegeneration in the recipient cells.
Moreover, exosomes can deliver microRNAs. Exosome-mediated microRNA signature could be utilized in the diagnosis of these diseases.
On the basis of these evidence, our aim is to deeply investigate the role of exosomes in the pathogenesis and in the progression of neurodegenerative diseases for a concrete use of exosomes for diagnosis and as novel therapeutic approach in these diseases.
Chiantore M.V. (ISS)
Romeo G., Dr. Mangino G., Dr. Iuliano M. (Università Sapienza Roma)

4.4 Ferritin nanoparticles for innovative therapies against brain tumors (Falvo E., Colotti G., Ilari A., Morea V., Ceci P. IBPM)

We will develop nanoparticles based on the human protein ferritin to selectively deliver therapeutic and/or diagnostic agents to brain tumors. The human protein ferritin is endowed with a number of favourable properties that make it an ideal nanocarrier for biomedical applications: it is present both in plasma and within cells in physiological conditions, scarcely immunogenic, non-toxic and long circulating in vivo; it can incorporate small compounds, shielding them from the external environment; it is imported in all cell types by a specific receptor, which is overexpressed by many types of cancer cells; it can be produced in high yields with low costs in bacterial cells. As far as delivery to brain regions is concerned, ferritin-based nanoparticles have been recently shown to be able to cross the endothelium, epithelium, and blood-brain barrier layers. Additionally, in brain tumors, such as glioblastoma, the integrity of the blood-brain barrier is compromised, allowing an increased passage of proteins between peripheral vasculature and central nervous system.

4.6 GBM: Characterization of Axitinib treatment on glioblastoma angiogenesis (Falchetti ML, -IBCN)

Axitinib, a tyrosine kinase inhibitor, works as a specific inhibitor of Vascular Endothelial Growth Factor Receptors 1, 2 and 3. It is FDA-approved for the treatment of metastatic renal cell carcinoma. In vitro, chronic treatment with Axitinib induces senescence in different glioblastoma cell lines. It is known that, following radiotherapy, endothelial cells of GBM patients may undergo cellular senescence releasing a variety of pro-inflammatory chemokines and citokines eventually resulting in a huge modification of their SASP (Senescence Associated secretory Phenotype), able to strongly affect tumor microenvironment. Starting from these premises, we want to perform a characterization of the consequences of Axitinib treatment on endothelial cells. We are currently performing a gene expression profiling by RNAseq of HUVEC cells treated with Axitinib.

4.7 Isolated peptides from mt-leucyl-tRNA synthetase as novel therapeutic instruments against neurodegenerative mitochondriopathies (Morea V., Colotti G., Ceci P.-IBPM)

Mitochondrial encephalopathy, lactic acidosis, and stroke (MELAS) syndrome and myoclonic epilepsy with ragged red fibers (MERRF) syndrome are multisystem and progressive neurodegenerative disorders due to mutations in mitochondrial DNA genes (mt-tRNALeuUUR and mt-tRNALys, respectively). MELAS neurodegeneration involves the cerebellar purkinje layer and cortical neurons, while MERRF neurodegeneration involves the cerebellar purkinje layer and dentate nucleus.We have identified several short peptides derived from human mitochondrial LeuRS (leucyl tRNA synthetase) that represent attractive new candidates for future therapeutic applications against MELAS and MERRF syndromes: overexpression of LeuRS-derived peptides is able to rescue the defective phenotype of cells bearing mt-tRNA mutations that cause MELAS and MERRF. Additionally, LeuRS-derived peptides directly interact with the mutated tRNAs and stabilize their functional conformations. We will implement strategies to deliver LeuRS-derived peptides to the mitochondria and will identify small non-peptide molecules able to both mimic the biological activity of the peptides and spontaneously diffuse to the mitochondrial matrix.
D’Amati G., Prof. Giordano C. (Sapienza University)
Cantatore P. (Bari University)
Bresciani A. (IRBM)

4.8 LTL as a biomarker in MCI and AD diseases (Scarabino D.-IBPM)

The main interest is to investigate susceptibility genes for age-related complex neurodegenerative diseases, in particular Alzheimer's disease (AD) and Mild Cognitive Impairment (MCI). Alzheimer's disease (AD) is a neurodegenerative disease associated with a drastic decline in life expectancy. Mild Cognitive Impairment (MCI), a clinical entity considered prodromal of AD, is characterized by memory alterations in subjects with normal global cognitive function. Our recent data have shown that some polymorphisms of the telomerase genes TERC and TERT are involved in susceptibility to AD confirming the relationship between the onset of AD / telomerase / telomere length. Numerous studies have addressed the analysis of LTL in neurodegenerative diseases, the overall data suggest that LTL could be a marker for cellular pathology in neurodegenerative diseases. Moreover, in a recent investigation, we have shown that LTL measurement may be a useful marker to follow the dementia progression . The possibility of detecting and following the disease before the onset of the most serious symptoms, could provide a window of opportunity for actions aimed at preventing or delaying the disease.
Corbo R.M. (Department of Biology and Biotechnology, Sapienza University)
Gambina G., Brogio E. (Alzheimer's Disease Center, Department of Neuroscience, University and Hospital of Verona)
Businaro R. (Department of Medico-Surgical Sciences and Biotechnologies, Sapienza University)
Maida C., Gaudio M.R. (S. Giovanni –Addolorata Hospital, Rome)
Mantuano E.,Veneziano L. (CNR Institute of Translational Pharmacology (IFT))

4.9 Pathological memory: from disease’ mechanisms to therapeutic strategies (De Leonibus E-IBCN).

We study the neurobiology of learning and memory in normal and pathological conditions, which include aging, ageing associated neurodegenerative disorders (Alzheimer’ disease, Parkinson’ disease and synucleinopathies), paediatric neurodegenerative disorders (lysosomal storage disorders), and neurodevelopmental disorders (autism, ADHD, and schizophrenia). The goal of our projects is to identify early cognitive deficits preceding the onset of neuronal loss in neurodegenerative disorders; working at this early stage we can identify the synaptic and molecular disease’ mechanisms leading to neuronal loss. This information is used to identify novel restorative or symptomatic therapeutic approaches (pharmacological, gene-therapy or the combination of the two) to correct the cognitive deficits and/or preventing the conversion to dementia-like symptoms.
Techniques routinely used in the lab: 1. In vivo: Behavioural testing probing different types of memory in rodents (mice and rats), cannula brain permanent implantation, optogenetics and we are setting up electroencefalografic recording in mice. 2. Ex-vivo: Western blot, brain tissue fractionations, immunohistochemistry, immunofluorescence, RNAseq and Mass Spectrometry in collaboration with the Telethon Institute of Genetics and Medicine (TIGEM) bioinformatic core; 3. In vitro: primary neuronal cultures and directed reprogrammed neurons from mouse or human fibroblasts, adeno-associated viral vector production in collaboration with TIGEM-aav core.
Mele A. (University of Roma,Sapienza), Calabresi P., (University of Perugia) Picconi B. (University San Raffaele la Pisana), Ballabio A., Alessandro Fraldi A. (TIGEM), Jezek K.(University of Pilsen, Chzec Republic),Di Angelantoio S. Ruocco G. (IIT, Sapienza), Gardoni F. (University of Milano), Sulzer D. (Columbia University, USA)

4.10 QMT: Modulation of neurotrophins’ and cytokines’ levels in human subjects (Caserta M., Verdone L.-IBPM)

This field of research is related to the analysis of the effects of Quadrato Motor Training, a specific motor-cognitive training, on the levels of relevant molecular markers in humans: neurotrophins, such as Brain-derived Neurotrophic Factor (BDNF) and Nerve Growth Factor (NGF), both known to be involved in the development and function of a wide variety of neuron populations, and cytokines as markers of neuroinflammation. Experiments enrolling different groups of healthy volunteers have shown that the training induces changes of proBDNF and proNGF salivary levels. In a subset of participants, proBDNF increase was shown to correlate with changes in morphological parameters at the level of defined brain areas, as observed by fMRI performed in collaboration with the Department of Physiology and Pharmacology at Sapienza University. The recent implications of neurotrophins and their receptors in the pathogenesis and therapy of neurodegenerative diseases suggest this training could have an impact on their prevention and treatment.
Ben-Soussan T.D. (Research Institute for Neuroscience, Education and Didactics, Patrizio Paoletti Foundation)
Venditti S. (Department of Biology and Biotechnology, Sapienza University Rome)
Raffone A. (Department of Psychology, Sapienza University of Rome)

4.11 Epigenetic drugs as therapeutic approach in DMD (Mozzetta C. -IBPM)

Duchenne Muscular Dystrophy (DMD) is the most severe form of dystrophy that leads to progressive muscle weakness because of a gradual replacement of functional muscle with fat and fibrotic scars. Pharmacological therapies for DMD should therefore aim to counteract this fibro-adipogenic degeneration and to promote the compensatory regeneration to slow down progression of pathology. Previous works proved pre-clinical efficacy of pan-histone deacetylase inhibitors (HDACi) in the treatment of murine models of DMD, showing the ability of HDACi to counter disease progression and induce functional and morphological recovery. These studies paved the way for an epigenetic therapy as a potential therapeutic approach in DMD prompting our interest in exploring pre-clinical efficacy of more selective compounds, such as specific inhibitors for class I HDACs (I-HDACi). Moreover, other epigenetic modifiers, such as Histone Lysine Methyltransferases (KMTs) are emerging as particularly relevant in myogenesis, given their high specificity for precise histone residues, the development of KMTs specific inhibitors might be a strategy to increase selectivity of epigenetic pharmacology in the context of DMD.
IRBM (Pomezia, Italy)
Dr. Arianna Rinaldi (Department of Biology and Biotechnology “C. Darwin”, Sapienza University Rome)
Prof. Maria Egle De Stefano (Department of Biology and Biotechnology “C. Darwin”, Sapienza University Rome)

4.12 AD: mitochondrial dysfunction and transplantation therapy (Di Carlo M., Picone P., Galizzi G., Nuzzo D. IRIB)

Studies on mitochondrial dysfunction as early event in neurodegenerative diseases including Alzheimer’s disease. In particular, mitochondrial dynamics, biogenesis, homeostasis and degradation (mitophagy) are analyzed by using specific biomarkers and imaging techniques. Use of antioxidant molecules and insulin as therapeutic approach to prevent or reduce mitochondrial dysfunction is contemplated. It is also studied the role of mitochondria-associated ER membranes (MAMs) in the control of calcium perturbation, signal transduction, mitochondrial and ER stress, mitochondrial dysfunction, and Amyloid Precursor Protein (APP) cleavage. A new approach, called mitochondrial transplantation, based on the possibility of replacing damaged mitochondria with healthy exogenous mitochondria, is currently being studied. Development of techniques to isolate and characterize neuro-microvesicles to be used as mitochondria delivery carrier are considered.
Dr. Donatella Bulone and Pierluigi San Biagio Biophysic Institute (IBF), CNR, Palermo; Prof. Valeria Vetri Chemistry Physics Department (DCF) Palermo University

4.13 CRISPR/Cas9-mediated gene editing in in vitro and in vivo models of Myotonic Dystrophy type 1: assessment of efficiency, safety and therapeutic effect of CTG‐repeat deletion. (Falcone F., Cardinali C., Provenzano C., Mandillo S., Golini E., Strimpakos G., IBBC)

Myotonic dystrophy type 1 (DM1) is a dominantly inherited, multisystemic disorder caused by expanded CTG repeats in the 3’UTR of the DMPK gene. DMPK mutated transcript accumulates into nuclear foci that affect the localization and activities of RNA-binding proteins involved in splicing regulation. No effective therapy is yet available for DM1. Our project focuses on the design, application and molecular characterization of a CRISPR/Cas9-mediated gene editing approach for the permanent elimination of the toxic mutant repeats in both in vitro and in vivo disease models, in order to reverse the pathologic phenotype. A deep understanding of the efficiency and specificity of the CRISPR/Cas9 gene editing strategy is essential to evaluate its potential application as gene therapy in DM1patients.
Key words: CRISPR/Cas9, DM1 models, repeat expansion disease, gene therapy, adeno-associated virus (AAV)
Fabio Martelli, IRCCS Policlinico San Donato, San Donato Milanese, Milan
Genevieve Gourdon, Institut de Myologie, Paris, France

4.14 Neurodegenerative and neuromuscular diseases, Innovative diagnostic and therapeutic strategies (Manni L., Soligo M. ITP)

Nerve growth factor has been indicated as possible therapeutic for neurodegenerative diseases. We recently explored its therapeutic efficacy after intranasal delivery in pediatric patients affected by severe brain traumas. However, the predominant form of NGF expressed in the brain is proNGF. The physiology of the different proNGF variants is still unexplored. We aimed our recent work at exploring proNGF variants production, processing, release and action, in animal models of neurodegeneration. We recently demonstrated, both in vitro and in vivo, that different native proNGF variants may challenge different receptor(s), exerting selective biological roles. We also recently received financial support to explore the role of different proNGFs as surrogate biomarkers in pediatric traumatic brain injury.
European Brain Research Institute (EBRI), Rome, Italy
Department of Biology and Biotechnology “Charles Darwin”, Sapienza University, Rome, Italy
Department of Biology, University of Rome Tor Vergata, Rome, Italy
Institute of Pediatrics, Università Cattolica del Sacro Cuore, Rome – Italy
Pediatric Intensive Care Unit, Policlinico A. Gemelli, Rome, Italy
Division of Rheumatology and Immuno-Rheumatology Research Laboratories, Bambino Gesù Children’s Hospital, Rome, Italy
Dept of Physiology and Pharmacology, Karolinska Institute, Stockholm, Sweden.

4.15 A multi-omics perspective of neurodegeneration: new clues for personal diagnostics and therapies (Cavallaro S., Guarnaccia M., Gentile G., Spampinato A.G. IRIB)

Our research group investigates neurodegeneration through a systems biology approach and multi-omics integration. Different layers of genomic information arising from the analysis of human pathology, together with in vitro and in vivo models of neurodegenerative disorders, are allowing a systems biology portrait of neurodegeneration. This approach is not only helping to identify key drivers and pathways underlying neurodegeneration, but is laying the foundation for a molecular taxonomy and patient-tailored therapies. This new perspective allows an innovative pharmacology focused on downstream targets, networks and transcriptional modules controlling neuronal fate. The final goal of our research is to develop breakthrough therapies for neurodegenerative diseases through integration of diverse omics data, stringent and reliable drug target selection and validation.
Dr. Cinzia Severini, CNR-IBBC, Rome, Italy.
Dr. Cinzia Volontè, CNR- IASI, Rome, Italy.
Prof. Eleonora Aronica, University of Amsterdam, The Netherlands.
Prof. Velia D’Agata, University of Catania, Italy.
Prof. Francesca Luisa Conforti, University of Calabria, Cosenza, Italy.

4.16 Olfactory Ensheathing Cells, a glial stem cell type, as promising tool for cell therapy (Pellitteri R.- IRIB)

Cell therapy has attracted considerable interest as a promising approach for repair in neurodegenerative diseases. Olfactory Ensheathing Cells (OECs), are glial cells able to secrete trophic factors, to exert neuroprotective effects and to promote plasticity in the lesioned area. OECs have capability to express stem cell marker, such as nestin, showing stem cell characteristics, indicating them as useful potential clinical agents to support nerve injured. In the recent years, several combined treatments have been studied: they include the use of OECs plus phitochemical molecules (curcumin), or neuropeptides, (ghrelin), or use of OECs plated on silicon-carbide substrate, a highly biocompatible material. Therefore, OECs might be considered as clinical tool for lesioned neural areas, representing a future perspective of novel cell therapies that would to be useful for the neuro-degenerated patients.
Proff. Antonella Russo, Stefania Stanzani, Rosario Giuffrida, Debora Lo Furno - Department of Biomedical and Biotechnological Sciences, University of Catania, Catania, Italy.
Cristina Russo, Ph.D student in Neuroscience – Department of Biomedical and Biotechnological Sciences, University of Catania, Catania, Italy.
Proff. Agata Campisi, Teresa Musumeci - Dep of Pharmaceutical Sciences, Section of Biochemistry, University of Catania, Italy.
Prof. Vincenzo Silani, Dr. Patrizia Bossolasco, Dr. Lidia Cova - Dep. Neurology and Laboratory of Neuroscience, IRCCS, Istituto Auxologico Italiano, Milan (Italy).
Prof. Maria Teresa Moreno-Flores - Dip. Anatomia, Histologia y Neurociencia Facultad de Medicina, Universidad Autonoma de Madrid, Madrid (Spain)

4.17 Therapeutic effects of novel drugs for spinal muscular atrophy (Crescimanno G., Marrone O., IRIB)

Spinal muscular atrophy (SMA) is characterized by reduction of the survival motor neuron protein (SMN), which is normally found mainly in the spinal cord and plays a critical role in the survival of spinal motor neurons. In this disease, skeletal muscles become progressively hypotrophic and weaker. Eventually, respiratory muscles impairment leads to death. Two drugs for SMA have been recently developed. Nusinersen, which was developed with «anti-sense RNA» biotechnology, seems to be less effective on respiratory than on other skeletal muscles; its administration was often associated with respiratory tract infections and atelectasis. Risdiplam, an investigational SMN2 splicing modifier, is expected to be more effective on respiratory function. We are interested to evaluate long-term effects of these drugs in patients with SMA, paying particular attention to the performance of respiratory muscles.
NEMO Center Milano and Messina