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1. Myxoid glioneuronal tumor: Histopathologic, neuroradiologic, and molecular features in a single center series

Author

C. Caporalini, M. Scagnet, L. Giunti, V. Cetica, D. Mei, V. Conti, S. Moscardi, L. Macconi, F. Giordano, L. D'Incerti, L. Genitori, R. Guerrini, and A.M. Buccoliero

Subjects
Myxoid glioneuronal tumor, PDGFRA, Septum pellucidum, Dysembryoplastic neuroepithelial tumors, Neoplasms. Tumors. Oncology. Including cancer and carcinogens, RC254-282
Abstract

Background: Myxoid glioneuronal tumor (MGT) is a benign glioneuronal neoplasm recently introduced in the World Health Organization (WHO) classification of the central nervous system (CNS) tumors. MGTs are typically located in the septum pellucidum, foramen of Monro or periventricular white matter of the lateral ventricle. They were previously diagnosed as dysembryoplastic neuroepithelial tumors (DNT), showing histological features almost indistinguishable from classical cortical DNT. Despite that, MGTs have been associated with a specific dinucleotide substitution at codon 385 in the platelet-derived growth factor receptor alpha (PDGFRA) gene, replacing a lysine residue with either leucine or isoleucine (p. LysK385Leu/Iso). This genetic variation has never been described in any other CNS tumor. Materials and methods: Thirty-one consecutive tumors, previously diagnosed as DNTs at the Meyer Children's Hospital IRCCS between January 2010 and June 2021 were collected for a comprehensive study of their clinical, imaging, pathological features, and molecular profile. Results: In six out of the thirty-one tumors we had previously diagnosed as DNTs, we identified the recurrent dinucleotide mutation in the PDGFRA. All six tumors were typically located within the periventricular white matter of the lateral ventricle and in the septum pellucidum. We then renamed these lesions as MGT, according to the latest WHO CNS classification. In all patients we observed an indolent clinical course, without recurrence. Conclusion: MGT represent a rare but distinct group of neoplasm with a typical molecular profiling, a characteristic localization, and a relative indolent clinical course.

Published
2023
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2. MOP and NOP receptor interaction: Studies with a dual expression system and bivalent peptide ligands

Author

M. F. Bird, J. McDonald, B. Horley, J. P. O’Doherty, B. Fraser, C. L. Gibson, R. Guerrini, G. Caló, and D. G. Lambert

Subjects
Medicine, Science
Abstract

Opioids targeting mu;μ (MOP) receptors produce analgesia in the peri-operative period and palliative care. They also produce side effects including respiratory depression, tolerance/dependence and addiction. The N/OFQ opioid receptor (NOP) also produces analgesia but is devoid of the major MOP side effects. Evidence exists for MOP-NOP interaction and mixed MOP-NOP ligands produce analgesia with reduced side effects. We have generated a HEKMOP/NOP human expression system and used bivalent MOP-NOP and fluorescent ligands to (i) probe for receptor interaction and (ii) consequences of that interaction. We used HEKMOP/NOP cells and two bivalent ligands; Dermorphin-N/OFQ (MOP agonist-NOP agonist; DeNO) and Dermorphin-UFP101 (MOP agonist-NOP antagonist; De101). We have determined receptor binding profiles, GTPγ[35S] binding, cAMP formation and ERK1/2 activation. We have also probed MOP and NOP receptor interactions in HEK cells and hippocampal neurones using the novel MOP fluorescent ligand, DermorphinATTO488 and the NOP fluorescent ligand N/OFQATTO594. In HEKMOP/NOP MOP ligands displaced NOP binding and NOP ligands displaced MOP binding. Using fluorescent probes in HEKMOP/NOP cells we demonstrated MOP-NOP probe overlap and a FRET signal indicating co-localisation. MOP-NOP were also co-localised in hippocampal tissue. In GTPγ[35S] and cAMP assays NOP stimulation shifted the response to MOP rightwards. At ERK1/2 the response to bivalent ligands generally peaked later. We provide evidence for MOP-NOP interaction in recombinant and native tissue. NOP activation reduces responsiveness of MOP activation; this was shown with conventional and bivalent ligands.

Published
2022

3. Pharmacological Profile of Nociceptin/Orphanin FQ Receptors Interacting with G-Proteins and β-Arrestins 2.

Author

D Malfacini, C Ambrosio, M C Gro', M Sbraccia, C Trapella, R Guerrini, M Bonora, P Pinton, T Costa, and G Calo'

Subjects
Medicine, Science
Abstract

Nociceptin/orphanin FQ (N/OFQ) controls several biological functions by selectively activating an opioid like receptor named N/OFQ peptide receptor (NOP). Biased agonism is emerging as an important and therapeutically relevant pharmacological concept in the field of G protein coupled receptors including opioids. To evaluate the relevance of this phenomenon in the NOP receptor, we used a bioluminescence resonance energy transfer technology to measure the interactions of the NOP receptor with either G proteins or β-arrestin 2 in the absence and in presence of increasing concentration of ligands. A large panel of receptor ligands was investigated by comparing their ability to promote or block NOP/G protein and NOP/arrestin interactions. In this study we report a systematic analysis of the functional selectivity of NOP receptor ligands. NOP/G protein interactions (investigated in cell membranes) allowed a precise estimation of both ligand potency and efficacy yielding data highly consistent with the known pharmacological profile of this receptor. The same panel of ligands displayed marked differences in the ability to promote NOP/β-arrestin 2 interactions (evaluated in whole cells). In particular, full agonists displayed a general lower potency and for some ligands an inverted rank order of potency was noted. Most partial agonists behaved as pure competitive antagonists of receptor/arrestin interaction. Antagonists displayed similar values of potency for NOP/Gβ1 or NOP/β-arrestin 2 interaction. Using N/OFQ as reference ligand we computed the bias factors of NOP ligands and a number of agonists with greater efficacy at G protein coupling were identified.

Published
2015
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4. Pathogenic MAST3 variants in the STK domain are associated with epilepsy

Author

Elizabeth Roeder, Christelle Moufawad El Achkar, Amy L Schneider, Carmen Barba, Samuel F. Berkovic, Eva H. Brilstra, R Guerrini, Heather C. Mefford, Annalisa Vetro, Emily Bryant, Tiziana Pisano, Alison M. Muir, Gemma L. Carvill, P. Ian Andrews, Jennifer Rakotomamonjy, Elysa J. Marco, Hadassa Goldberg-Stern, Kyle R Christensen, Richard H. van Jaarsveld, Simone Mandelstam, Satoko Kumada, Rebecca O. Littlejohn, Berkley Schmidt, Lance H. Rodan, Kazuhiro Iwama, Ingrid E. Scheffer, Kirsty McWalter, Linda Laux, Patrick W. Carney, Alicia Guemez-Gamboa, Naomichi Matsumoto, Angus C. Nairn, William G. Wilson, Jessica Giannelli, John Millichap, and Egidio Spinelli

Subjects
Adult, Male, Adolescent, Biology, Protein Serine-Threonine Kinases, Corpus callosum, Article, Cohort Studies, Epilepsy, Mice, Young Adult, Neurodevelopmental disorder, medicine, Missense mutation, Animals, Humans, Amino Acid Sequence, Child, Gene, Exome sequencing, Genetics, Seizure types, HEK 293 cells, Genetic Variation, medicine.disease, Mice, Inbred C57BL, HEK293 Cells, Neurology, Female, Neurology (clinical), Microtubule-Associated Proteins, Follow-Up Studies
Abstract

Objective: The MAST family of microtubule-associated serine–threonine kinases (STKs) have distinct expression patterns in the developing and mature human and mouse brain. To date, only MAST1 has been conclusively associated with neurological disease, with de novo variants in individuals with a neurodevelopmental disorder, including a mega corpus callosum. Methods: Using exome sequencing, we identify MAST3 missense variants in individuals with epilepsy. We also assess the effect of these variants on the ability of MAST3 to phosphorylate the target gene product ARPP-16 in HEK293T cells. Results: We identify de novo missense variants in the STK domain in 11 individuals, including 2 recurrent variants p.G510S (n = 5) and p.G515S (n = 3). All 11 individuals had developmental and epileptic encephalopathy, with 8 having normal development prior to seizure onset at

Published
2021

6. P04.08 The role of SCN1A in glioblastomas and mixed neuronal glial tumors of pediatric age

Author

Carmen Barba, Annamaria Buccoliero, Davide Mei, Milena Guidi, R Guerrini, Sabrina Giglio, Iacopo Sardi, Cetica, Laura Giunti, De Gregorio, Lorenzo Genitori, and M Censullo

Subjects
Mixed Glioma, Cancer Research, Pathology, medicine.medical_specialty, business.industry, Central nervous system, Tissue membrane, Cancer, Pediatric age, medicine.disease, Poster Presentations, Epilepsy, medicine.anatomical_structure, Oncology, Glioma, medicine, Neurology (clinical), business, Glioblastoma
Abstract

BACKGROUND Low and high grade gliomas, are the most common pediatric central nervous system (CNS) tumors and they show an extremely broad range of clinical behavior. Pediatric glioma is distinct from its adult counterpart with different genetic/epigenetic profile and biological features. Recently, several studies have shown the involvement of voltage-gated Na+ channels (VGSC) in different types of cancer, including gliomas. VGSC are multimeric transmembrane complexes, responsible for membrane depolarization in excitable cells playing an important role also in cell proliferation, migration, apoptosis and differentiation. VGSC are therapeutic targets in cardiovascular and neurological disorders and, in cancers, they could be a novel target for the development of promising anticancer therapy MATERIAL AND METHODS Firstly, we explored the genetic background of 9 pediatric glioblastomas (1–9 pGBMs), through whole-exome sequencing (WES) using HiSeq1000 platform (Illumina) with paired-end approach. On the basis of our results, we extended our study in another group of 16 epileptogenic mixed neuronal-glial tumors of pediatric age, (WHO grade I and II), through an amplicon approach (TSCA) using MiniSeq System platform (Illumina) RESULTS We identified variants in SCN1A gene in 3/9 pGBMs: case 3 had c.5782C>G in tumor and blood; case 5 showed c.2278G>T and two mosaic variants (c.5933C>T, 22% and c.4942C>T, 14%); case 6 showed c.667G>T variant only in tumor, and not in other non tumoral tissues (blood, urine and buccal swab). No variants in SCN1A were identified in a group of 16 pediatric mixed gliomas CONCLUSION In this study, we explore the genetic background of two groups of pediatric neuroepithelial brain tumors, through Next generation sequencing approach. We identified only in pGBMs variants in SCN1A gene that encoded for VGSCs and is involved in a spectrum of early-onset epileptic encephalopathies. None of our mutated patients showed history of epilepsy. Now, it is not clear the significance of these variants in pGBMs but interestingly, these variants are present in pGBM and not in mixed gliomas. Further studies on a big cohort of patients are needed to establish if they could play a role in pGBMs aggressiveness, migration and progression. Moreover, VGSCs could be a pharmacological target in pGBMs treatment

Published
2019

7. Management of psychogenic non-epileptic seizures: a multidisciplinary approach

Author

S. Gasparini 1, 2, E. Beghi 3, E. Ferlazzo 1, 4, M. Beghi 5, V. Belcastro 6, K.P. Biermann 7, G. Bottini 8, G. Capovilla 9, R.A. Cervellione 10, V. Cianci 2, G. Coppola 11, C.M. Cornaggia 12, P. De Fazio1 3, S. De Masi 7, G. De Sarro 14, M. Elia 15, G. Erba 16, L. Fusco 17, A. Gambardella 1, V. Gentile1 8, A. T. Giallonardo 19, R. Guerrini 20, F. Ingravallo 22, A. Iudice 23, A. Labate 1, E. Lucenteforte 21, A. Magaudda 24, L. Mumoli 1, C. Papagno 25, G.B. Pesce 18, E. Pucci 27, P. Ricci 28, A. Romeo 29, R. Quintas 30, C. Sueri 2, G. Vitaliti 31, R. Zoia 32, U. Aguglia 1, Gasparini, S, Beghi, E, Ferlazzo, E, Beghi, M, Belcastro, V, Biermann, K, Bottini, G, Capovilla, G, Cervellione, R, Cianci, V, Coppola, G, Cornaggia, C, De Fazio, P, De Masi, S, De Sarro, G, Elia, M, Erba, G, Fusco, L, Gambardella, A, Gentile, V, Giallonardo, A, Guerrini, R, Ingravallo, F, Iudice, A, Labate, A, Lucenteforte, E, Magaudda, A, Mumoli, L, Papagno, C, Pesce, G, Pucci, E, Ricci, P, Romeo, A, Quintas, R, Sueri, C, Vitaliti, G, Zoia, R, Aguglia, U, Gasparini S., Beghi E., Ferlazzo E., Beghi M., Belcastro V., Biermann K.P., Bottini G., Capovilla G., Cervellione R.A., Cianci V., Coppola G., Cornaggia C.M., De Fazio P., De Masi S., De Sarro G., Elia M., Erba G., Fusco L., Gambardella A., Gentile V., Giallonardo A.T., Guerrini R., Ingravallo F., Iudice A., Labate A., Lucenteforte E., Magaudda A., Mumoli L., Papagno C., Pesce G.B., Pucci E., Ricci P., Romeo A., Quintas R., Sueri C., Vitaliti G., Zoia R., and Aguglia U.

Subjects
Adult, Male, medicine.medical_specialty, conversion disorder, EEG, epilepsy, EEG, conversion disorder, epilepsy, Psychophysiologic Disorder, NO, 03 medical and health sciences, Epilepsy, 0302 clinical medicine, Seizures, Psychogenic non-epileptic seizures, medicine, Psychogenic disease, Humans, 030212 general & internal medicine, Psychiatry, Child, Conversion disorder, business.industry, Electroencephalography, Evidence-based medicine, Bioethics, medicine.disease, Personality disorders, Psychophysiologic Disorders, Neurology, Neurology (clinical), Anxiety, Female, medicine.symptom, business, 030217 neurology & neurosurgery, Human, conversion disorder, EEG, epilepsy, Adult, Child, Electroencephalography, Female, Humans, Male, Psychophysiologic Disorders, Seizures
Abstract

The International League against Epilepsy (ILAE) proposed a diagnostic scheme for psychogenic non-epileptic seizure (PNES). The debate on ethical aspects of the diagnostic procedures is ongoing, the treatment is not standardized and management might differ according to age group. The objective was to reach an expert and stakeholder consensus on PNES management. A board comprising adult and child neurologists, neuropsychologists, psychiatrists, pharmacologists, experts in forensic medicine and bioethics as well as patients’ representatives was formed. The board chose five main topics regarding PNES: diagnosis; ethical issues; psychiatric comorbidities; psychological treatment; and pharmacological treatment. After a systematic review of the literature, the board met in a consensus conference in Catanzaro (Italy). Further consultations using a model of Delphi panel were held. The global level of evidence for all topics was low. Even though most questions were formulated separately for children/adolescents and adults, no major age-related differences emerged. The board established that the approach to PNES diagnosis should comply with ILAE recommendations. Seizure induction was considered ethical, preferring the least invasive techniques. The board recommended looking carefully for mood disturbances, personality disorders and psychic trauma in persons with PNES and considering cognitive-behavioural therapy as a first-line psychological approach and pharmacological treatment to manage comorbid conditions, namely anxiety and depression. Psychogenic non-epileptic seizure management should be multidisciplinary. High-quality long-term studies are needed to standardize PNES management.

Published
2019
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8. Overview of presurgical assessment and surgical treatment of epilepsy from the Italian League Against Epilepsy

Author

R. Guerrini, M. Scerrati, G. Rubboli, V. Esposito, G. Colicchio, M. Cossu, C. E. Marras, L. Tassi, M. P. Canevini, P. Quarato, F. Giordano, T. Granata, F. Villani, M. Giulioni, P. Scarpa, V. Barbieri, G. Bottini, A. D. Sole, G. Vatti, R. Spreafico, G. L. Russo, C. f. Epilepsy, TINUPER, PAOLO, R. Guerrini, M. Scerrati, G. Rubboli, V. Esposito, G. Colicchio, M. Cossu, C. E. Marra, L. Tassi, P. Tinuper, M. P. Canevini, P. Quarato, F. Giordano, T. Granata, F. Villani, M. Giulioni, P. Scarpa, V. Barbieri, G. Bottini, A. D. Sole, G. Vatti, R. Spreafico, G. L. Russo, and C. f. Epilepsy

Subjects
methods, Treatment Outcome, medicine.medical_specialty, Epilepsy, business.industry, League, medicine.disease, Neurosurgical Procedures, Treatment Outcome, Neurology, diagnosis/epidemiology/surgery, Humans, Neurosurgical Procedure, Preoperative Care, Physical therapy, Medicine, Humans, Medical physics, Epilepsy surgery, Neurology (clinical), business, Surgical treatment, methods, Preoperative Care
Abstract

The Commission for Epilepsy Surgery of the Italian League Against Epilepsy (LICE) presents an overview of the techniques and methodologies of presurgical evaluation and of the surgical treatment of epilepsies. This overview is the result of the experience developed in the past years in the major Italian centers where programs of epilepsy surgery have been established, and it has the aim of offering a quick and easy reference tool for those involved in the treatment of patients with epilepsy. The sharing of different experiences has the additional aim of conforming and disseminating the employed techniques as well as the methods of selection and evaluation of patients. The synthetic coverage of the main issues concerning the presurgical workup and the available surgical options will hopefully provide a framework that may integrate and develop the contributions of every single center, in one of the more complex, challenging, and dynamic areas of neurological sciences.

Published
2013

9. Delta opioidmimetic antagonists: prototypes for designing a new generation of ultraselective opioid peptides

Author

S. Salvadori, M. Attila, G. Balboni, C. Bianchi, S. D. Bryant, CRESCENZI, ORLANDO, R. Guerrini, PICONE, DELIA, T. Tancredi, TEMUSSI, PIERO ANDREA, L. H. Lazarus, S., Salvadori, M., Attila, G., Balboni, C., Bianchi, S. D., Bryant, Crescenzi, Orlando, R., Guerrini, Picone, Delia, T., Tancredi, Temussi, PIERO ANDREA, and L. H., Lazarus

Subjects
congenital, hereditary, and neonatal diseases and abnormalities, Magnetic Resonance Spectroscopy, opioidmimetic, antagonist, peptide, dimethyltyrosine, tetrahydroisoquinolinecarboxylate, Narcotic Antagonists, Guinea Pigs, Molecular Sequence Data, In Vitro Techniques, Spectrometry, Mass, Fast Atom Bombardment, Substrate Specificity, Amphibians, Mice, Structure-Activity Relationship, Ileum, Receptors, Opioid, delta, Tetrahydroisoquinolines, mental disorders, Animals, Amino Acid Sequence, Skin, Receptors, Opioid, kappa, Dipeptides, Isoquinolines, Kinetics, Drug Design, Tyrosine, Indicators and Reagents, human activities, Oligopeptides, Research Article, Muscle Contraction
Abstract

BACKGROUND: Tyr-Tic (1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid) and Tyr-Tic-Ala were the first peptides with delta opioid antagonist activity lacking Phe, considered essential for opioid activity based on the N-terminal tripeptide sequence (Tyr-D-Xaa-Phe) of amphibian skin opioids. Analogs were then designed to restrain the rotational flexibility of Tyr by the substitution of 2,6-dimethyl-L-tyrosine (Dmt). MATERIALS AND METHODS: Tyr and Dmt peptides were synthesized by solid phase and solution methods using Fmoc technology or condensing Boc-Dmt-OH or Boc-Tyr(But)-OH with H-L-Tic-OBut or H-D-Tic-OBut, respectively. Peptides were purified (> 99%) by HPLC and characteristics determined by 1H-NMR, FAB-MS, melting point, TLC, and amino acid analyses. RESULTS: H-Dmt-Tic-OH had high affinity (Ki delta = 0.022 nM) and extraordinary selectivity (Ki mu/Ki delta = 150,000); H-Dmt-Tic-Ala-OH had a Ki delta = 0.29 nM and delta selectivity = 20,000. Affinity and selectivity increased 8700- and 1000-fold relative to H-Tyr-Tic-OH, respectively. H-Dmt-Tic-OH and H-Dmt-Tic-NH2 fitted one-site receptor binding models (eta = 0.939-0.987), while H-Dmt-Tic-ol, H-Dmt-Tic-Ala-OH and H-Dmt-Tic-Ala-NH2 best fitted two-site models (eta = 0.708-0.801, F 18.9-26.0, p < 0.0001). Amidation increased mu affinity by 10- to 100-fold and acted synergistically with D-Tic2 to reverse selectivity (delta-->mu). Dmt-Tic di- and tripeptides exhibited delta antagonist bioactivity (Ke = 4-66 nM) with mouse vas deferens and lacked agonist mu activity (> 10 microM) in guinea-pig ileum preparations. Dmt-Tic analogs weakly interacted with kappa receptors in the 1 to > 20 microM range. CONCLUSIONS: Dmt-Tic opioidmimetic peptides represent a highly potent class of opioid peptide antagonists with greater potency than the nonopioid delta antagonist naltrindole and have potential application as clinical and therapeutic compounds.

Published
1995

10. Education and self-assessment

Author

R Guerrini, S Brown, L Sander, N Pickard, P Thompson, and P Crawford

Subjects
Self-assessment, Epilepsy, Neurology, business.industry, Clinical Neurology, medicine, Neurology (clinical), General Medicine, medicine.disease, business, Clinical psychology
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11. Simultaneous targeting of multiple opioid receptors: a strategy to improve side-effect profile.

Author

N. Dietis, R. Guerrini, G. Calo, S. Salvadori, D. J. Rowbotham, and D. G. Lambert

Subjects
*OPIOID receptors, *TARGETED drug delivery, *DRUG side effects, *MORPHINE, *ANALGESICS, *DRUG tolerance, *PHARMACOLOGY, *MEDICAL literature
Abstract

Opioid receptors are currently classified as µ (mu: mOP), δ (delta: dOP), κ (kappa: kOP) with a fourth related non-classical opioid receptor for nociceptin/orphainin FQ, NOP. Morphine is the current gold standard analgesic acting at MOP receptors but produces a range of variably troublesome side-effects, in particular tolerance. There is now good laboratory evidence to suggest that blocking DOP while activating MOP produces analgesia (or antinociception) without the development of tolerance. Simultaneous targeting of MOP and DOP can be accomplished by: (i) co-administering two selective drugs, (ii) administering one non-selective drug, or (iii) designing a single drug that specifically targets both receptors; a bivalent ligand. Bivalent ligands generally contain two active centres or pharmacophores that are variably separated by a chemical spacer and there are several interesting examples in the literature. For example linking the MOP agonist oxymorphone to the DOP antagonist naltrindole produces a MOP/DOP bivalent ligand that should produce analgesia with reduced tolerance. The type of response/selectivity produced depends on the pharmacophore combination (e.g. oxymorphone and naltrindole as above) and the space between them. Production and evaluation of bivalent ligands is an emerging field in drug design and for anaesthesia, analgesics that are designed not to be highly selective morphine-like (MOP) ligands represents a new avenue for the production of useful drugs for chronic (and in particular cancer) pain. [ABSTRACT FROM AUTHOR]

Published
2009
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12. Drug Prescription and Delirium in Older Inpatients: Results From the Nationwide Multicenter Italian Delirium Day 2015-2016

Author

Aloisi, Gaetano, Marengoni, Alessandra, Morandi, Alessandro, Zucchelli, Alberto, Cherubini, Antonio, Mossello, Enrico, Mario, Bo, Di Santo Simona, Mazzone, Andrea, Trabucchi, Marco, Cappa, Stefano, Fimognari, Filippo, Antonelli Incalzi Raffaele, Gareri, Pietro, Perticone, Francesco, Campanini, Mauro, Montorsi, Marco, Latronico, Nicola, Zambon, Antonella, Bellelli, Giuseppe, Rispoli, Vincenzo, Malara, Alba, Spadea, Fausto, Di Cello Serena, Ceravolo, Francesco, Fabiano, Francesco, Chiaradia, Giuseppe, Gabriele, Amedeo, Lenino, Peluso, Andrea, Taristano, Settembrini, Vincenzo, Capomolla, Domenico, Citrino, Antonella, Scriva, Antonietta, Bruno, Irene, Secchi, Roberto, De Martino Eugenio, Muccinelli, Roberto, Lupi, Gerardo, Paonessa, Patrizio, Fabbri, Andrea, Passuti Maria Teresa, Castellari, Sofia, Andrea, Po, Gaggioli, Guido, Varesi, Massimo, Moneti, Paolo, Capurso, Sebastiano, Latini, Vincenzo, Ghidotti, Stefano, Riccardelli, Francesco, Macchi, Maurizio, Rigo, Rosaria, Claudio, Pascale, Bosio, Angelo, Cerrato, Flavio, Bardelli, Benedetta, Boffelli, Stefano, Cassinadri, Angela, Franzoni, Simone, Spazzini, Elena, Andretto, Daniela, Tonini, Gabriele, Andreani, Laura, Coralli, Mirco, Balotta, Antonio, Cancelliere, Roberto, Ballardini, Giorgio, Simoncelli, Margherita, Mancini, Annarita, Strazzacapa, Mara, Fabio, Stefano, De Filippi Francesco, Giudice, Chiara, Dentizzi, Cosimo, Azzini, Margherita, Cazzadori, Marco, Mastroeni, Valeria, Bertassello, Paolo, Santana, Helena, Benati, Claudia, Nesta, Elisa, Tobaldini, Chiara, Guerini, Fabio, Tambara, Elena, Mombelloni, Paolo, Fontanini, Fulvio, Lassa, Gabriella, Lassa, Pizzorni, Crosio, Oliverio, Martina, Del, Grosso, Luciano, Luca, Giavedoni, Cristina, Bidoli, Giuliano, Mazzei, Bruno, Corsonello, Andrea, Fusco, Sergio, Vena, Silvio, De Vuono Tommaso, Maiuri, Giorgio, Fimognari Filippo Luca, Arone, Andrea, Sgro, Giovanni, Nicolazzo, Rossella, Castegnaro, Eugenio, De Rosa Salvatore, Sechi Bazzano Rossella, Benvenuti, Enrico, Del Lungo Ilaria, Giardini, Sante, Giulietti, Chiara, Di Bari Mauro, Barghini, Eleonora, Paoli, Martina, Fiordelli, Irene, Barucci, Riccardo, Sgrilli, Federica, Bertoletti, Erik, D'Amico, Ferdinando, Caronzolo, Francesco, Grippa, Alessandro, Lombardo, Giuseppina, Pipicella, Tiziana, Saitta, Antonino, Corica, Francesco, Cucinotta, Valeria, Giaritta, Prestipino, Larosa, Daniela, Cucinotta, Domenico, Basile, Giorgio, March, Albert, Nitti Maria Teresa, Felici, Alessandro, Pavan, Silvia, Piazzani, Fabrizio, Lunelli, Alessandra, Dimori, Sergio, Margotta, Alessandro, Soglia, Tiziano, Postacchini, Demetrio, Brunelli, Roberto, Santini, Silvia, Francavilla, Monia, Macchiati, Ilenia, Sorvillo, Francesca, Giuli, Cinzia, Mecocci, Patrizia, Longo, Annalisa, Addesi, Desiree, Rosa Paola Cerra, Bencardino, Giuseppe, Falbo, Tania, Grillo, Nadia, Filice, Marco, Mirella, Filippo, Fanto, Fausto, Isaia, Gianluca, Pezzilli, Stella, Bergamo, Daniele, Furno, Elisabetta, Rrodhe, Sokol, Lucarini, Simonetta, Dijk, Babette, Dall'Acqua, Francesca, Cappelletto, Francesco, Calvani, Donatella, Becheri, Dimitri, Mottino, Giuseppe, Mitidieri, Costanza, Antonio, Vito, Bartalucci, Francesca, Magherini, Lorenza, Malin, Novella, Boni, Franca, Gambardella Paoli Martina Lucia, Valente, Carlo, Bracali, Ilaria, Bracali, Foschini, Alice, Porrino, Paola, Ceci, Giacomo, Bottignole, Giuliana, Tibaldi, Michela, Coppo, Eleonora, Ettore, Evaristo, Camellini, Cinzia, Servello, Adriana, Grassi, Alessandro, Rozzini, Renzo, Tironi, Sara, Grassi Maria Grazia, Troisi, Elio, Caltagirone, Carlo, Franchini, Flaminia, Ratto, Federica, Pavoni, Beatrice, Toniolo, Sofia, Gabutto, Anna, Quazzo, Loredana, Rosatello, Annalisa, Suraci, Domenico, Tagliabue, Benedetta, Perrone, Chiara, Ferrara, Lucia, Castagna, Alberto, Tremolada Maria Luisa, Castoldi, Giuseppe, Barbero, Stefano, Oltramonti, Davide, Piano, Simonetta, Serviddio, Gaetano, Lo Buglio Aurelio, Gurrera, Tiziana, Merlo, Valeria, Rovai, Carla, Cotroneo Antonino Maria, Carlucci, Rosaria, Abbaldo, Anna, Monzani, Fabio, Qasem Ahmad Amedeo, Bini, Giacomo, Tafuto, Silvia, Galli, Giovanni, Bruni Amalia Cecilia, Mancuso, Giovanna, Mancuso, Gerardo, De Luca Giuseppe Massimiliano, Calipari Daniela., Bernardini, Bruno, Corsini, Carla, Ciccarelli, Michele, Dal Farra Sara, Dal, Farra, Cagnin, Annachiara, Fragiacomo, Federica, Pompanin, Sara, Amodio, Piero, Cagnin, Marco, Zurlo, Amedeo, Guerra, Gianluca, Pala, Marco, Menozzi, Luca, Delli Gatti Chiara, Magon, Stefania, Manfredini, Roberto, De Giorgi Alfredo, Fabbian, Fabio, Tiseo, Ruana, Misurati, Elisa, Boari, Benedetta, Molino, Christian, Gallerani, Massimo, Di Francesco Vincenzo, Faccioli, Silvia, Pellizzari, Luca, Fontana, Giorgia, Barbagallo, Giuseppe, Lunardelli Maria Lia, Martini, Emilio, Ferrari, Eleonora, Macchiarulo, Maria, Corneli, Maria, Bacci, Monica, Battaglia, Giuseppe, Anastasio, Luigi, Lo Storto Mario Sostene, Seresin, Chiara, Simonato, Matteo, Loreggian, Michele, Cestonaro, Fausta, Durando, Mario, Latella, Raffaele, Mazzoleni, Marta, Russo, Giuseppe, Ponte, Martino, Valchera, Alessandro, Salustri, Giuseppe, Petritola, Donatella, Costa, Alfredo, Sinforiani, Elena, Cotta Matteo Ramusino, Pizio Renato Nicola, Cester, Alberto, Formilan, Marino, Bonometto, Pietro, Carbone, Pasqualina, Cazzaniga, Ilaria, Appollonio, Ildebrando, Cereda, Diletta, Stabile, Andrea, Xhani, Rubjona, Acampora, Roberto, Tremolizzo, Lucio, Pieruzzi, Federico, Ciaccio, Antonio, Pontecorvi, Valerio, Burti, Cesare, Mokini, Zhirajr, Vitale, Giovanni, Amigoni, Maria, Sparacino, Mariaelena, Bottacchi, Edo, Bucciantini, Elisabetta, Di Giovanni Marco, Franchi, Fabrizio, Lucchetti, Lucio, Mariani, Claudio, Grande, Giulia, Rapazzini, Piero, Mauri, Marco, Romanelli, Giuseppe, Nicosia, Franco, Margola, Alessio, Bonardelli, Stefano, Porcella, Laura, Portolani, Nazario, Concoreggi, Carlo, Grassini, Chiara, Salvi, Andrea, Bianchetti, Luca, Spagnoli, Francesca, Apuzzo, Roberto, Fontanella, Ceraso, Anna, Castelli, Francesco, Fugazza, Luciano, Guerrini, Chiara, De Paduanis Giovanna, Iallonardo, Lucia, Palumbo, Pasquale, Zuliani, Giovanni, Ortolani, Beatrice, Capatti, Eleonora, Soavi, Cecilia, Bianchi, Lara, Francesconi, Daniela, Miselli, Agata, Brombo, Gloria, Romagnoli, Tommaso, Pazzaglini, Chiara, Miselli Maria Agata, Dall'Agata, Marco, Suardi, Teresa, Zaccarini, Cinzia, Riva, Manuela, Mirra, Gianluca, Muti, Ettore, Bottura, Renato, Moretti, Gianpaolo, Secreto, Piero, Bisio, Erika, Cecchettani, Marco, Naldi, Tamara, Pallavicino, Alessandra, Pugliese, Michela, Iozzo Rosaria Cosima, Grassi, Guido, Bombelli, Michele, Delloro, Raffaella, Quarti Trevano Fosca, Giussani Carlo Giorgio, Paterno, Giovanni, Contro, Ernesto, Mannironi, Antonio, Giorli, Elisa, Oberti, Sara, Fierro, Brigida, Piccoli, Tommaso, Giacalone, Fabio, Mandas, Antonella, Serchisu, Luca, Costaggiu, Diego, Pinna, Elisa, Orru, Francesca, Mannai, Martina, Cordioli, Zeno, Pelizzari, Luca, Turcato, Emanuela, Arduini, Pietro, Cacace, Carlo, Chiloiro, Roberta, Cimino, Rosella, Ruberto, Carmen, Ruotolo, Giovanni, Greco, Laura, Dal Santo Pierluigi, Andriolli, Antonino, Burattin, Giuseppe, Rossi, Laura, Andreolli, Antonino, Cervati, Giovanna, Catalano, Giuseppe, Tezza, Fabiana, Pizzardini, Maddalena, Stievano, Laura, Crippa, Patrizia, Aloisio, Paola, Di Monda Tiziana, Malighetti, Alessandro, Galbassini, Gloria, Ivaldi, Claudio, Russo Anna Maria, Bennati, Ettore, Pino, Elisabetta, Zavarise, Gianmaria, Pesci, Alberto, Suigo, Giulia, Faverio, Paola, Gori, Andrea, Perego Sabrina Perego, Zanasi, Massimo, Moniello, Giovanni, Rostagno, Carlo, Cartei, Alessandro, Polidori, Gianluca, Ungar, Andrea, Melis Maria Ramona, Martellini, Eleonora, Torrini, Monica, Giordano, Antonella, Leone, Giovanna, Migliorini, Marta, Caramelli, Francesca, Battiston, Bruno, Berardino, Maurizio, Cavallo, Simona, Masse, Alessandro, Santoro, Anna, Lombardi, Bruna, D'Ippolito, Pierpaolo, Furini, Angela, Villani, Daniele, Raimondi, Clara, Guarneri, Massimo, Paolucci, Stefano, Bassi, Andrea, Coiro, Paola, De Angelis Domenico, Morone, Giovanni, Venturiero, Vincenzo, Palleschi, Lorenzo, Raganato, Paolo, Di Niro Giuseppina, Casini Anna Rosa, Bove, Loredana, Imoscopi, Alessandra, Isaia, Giancarlo, Tibaldi, Vittoria, Calvi, Elisa, Clementi, Carlotta, Zanocchi, Mauro, Agosta, Luca, Criasia, Antonio, Spertino, Elena, Nortarelli, Antonella, Provenzano, Giuseppe, Principato, Pietro, Rizzo, Anita, Cellura, Eleonora, Mari, Daniela, Romano Federica Ylenia, Rosini, Francesca, Mansi, Marta, Rossi, Silvia, Riccardelli, Alex, Inzaghi, Lorenzo, Bonini, Giulia, Rossi, Paolo, Potena, Alfredo, Lichii, Mihaela, Candiani, Tiziana, Grimaldi, William, Bertani, Emiliano, Porta, Alessandra, Calogero, Pietro, Pinto, Daniela, Bernardi, Roberto, Nicolino, Francesco, Galetti, Caterina, Gianstefani, Alice, Corvalli, Giulia, Mulazzani, Lorenzo, Odetti, Patrizio, Monacelli, Fiammetta, Prefumo, Matteo, Canepa, Marta, Minaglia, Cecilia, Paolisso, Giuseppe, Rizzo Maria Rosaria, Prestano, Raffaele, Dalise Anna Maria, Barra, Davide, Dal Bosco Livio, Asprinio, Vincenzo, Dallape, Luciana, Maina, Paola, Perina, Elisa, Bartoli Rossi Isaura, Pluderi, Alice, Maina, Antonella, Pecoraro, Elisabetta, Sciarra, Michela, Prudente, Angela, Mete, Francesca, Ventura, Manuel, Cesari, Luisella, Pernigotti Luigi Maria, Santangelo, Tina, Benini, Lucia, Levato, Francesco, Mhiuta, Victor, Alius, Florin, Davidoaia, Diana, Giardini, Vittorio, Garancini, Mattia, Bellamoli, Claudio, Terranova, Luciano, Bozzini, Claudia, Tosoni, Paolo, Provoli, Emma, Cascone, Luisa, Dioli, Andrea, Ferrarin, Gianfranco, Bucci, Adelmo, Bua, Guido, Fenu, Sara, Bianchi, Giovanna, Casella, Silvia, Romano, Valentina, Poli, Maurizio, Mascherona, Ilenia, Belotti, Gloria, Cavaliere, Sabina, Cuni, Estella, Merciuc, Nina, Oberti, Rosella, Veneziani, Silvia, Capoferri, Emanuela, De Bernardi Elisabetta, Colombo, Katia, Bravi, Marco, Negrinotti, Nicoletta, D'Arcangelo, Paolo, Montenegro, Nicola, Montanari, Roberto, Lamanna, Pierpaolo, Gasperini, Beatrice, Montesi, Isabella, Diotallevi, Stefania, Altobelli, Gaia, Cacinaro, Filippo, Palama, Chiara, Di Emidio Chiara, Scarpini, Elio, Arighi, Andrea, Scrimieri, Antonia, Fumagalli, Giorgio, Basilico, Paola, Migone De Amicis Margherita, Mancarella, Marta, Maira, Diletta, Granata, Antonino, Ranalli, Claudia, Cammilli, Alessandra, Cavallini Maria Chiara, Tricca, Manola, Natella, Daniela, Gabbani, Luciano, Tesi, Francesca, Martella, Letizia, Imbrici, Rosalba, Pacchioni, Maurizio, Gianbattista, Guerrini, Scotuzzi Anna Maria, Sozzi, Ferdinando, Valenti, Luigi, Chiarello, Antonino, Monella, Monia, Pilotto, Alberto, Prete, Camilla, Senesi, Barbara, Meta Anna Cristina, Pendenza, Enrico, Pasqualetti, Giuseppe, Polini, Antonio, Tognini, Sara, Ballino, Elena, Dell'Aquila, Giuseppina, Gasparrini Pina Maria, Marotti, Elisabetta, Migale, Monica, Falsetti, Lorenzo, Salvi, Aldo, Toigo, Gabriele, Ceschia, Giuliano, Rosso, Alessia, Tongiorgi, Chiara, Scarpa, Cristina, De Dominicis Luigino, Pucci, Eugenio, Renzi, Sara, Cartechini, Elisabetta, Tomassini Pia Francesca, Del Gobbo Maurizio, Ugenti, Francesca, Romeo, Pasquale, Nardelli, Anna, Lauretani, Fulvio, Visioli, Sandra, Montanari, Ilaria, Ermini, Francesca, Giordano, Antonio, Pigato, Giorgio, Simeone, Emilio, Colameco, Franco, Cecamore, Antonella, Scurti, Rosa, Lupinetti Maria Cristina, Barbujani, Mario, Perazzi, Beatrice, Giampieri, Marina, Amoruso, Raffaele, Piccinini, Maristella, Ferrari, Camilla, Gambetti, Claudio, Sfrappini, Mario, Semeraro, Letizia, Striuli, Rinaldo, Mariani, Claudia, Pelliccioni, Giuseppe, Marinelli, Donatella, Fabi, Katia, Rossi, Tommaso, Pesallaccia, Martina, Sabbatini, Debora, Gobbi, Beatrice, Cerqua, Raffaella, Tagliani, Giancarla, Schlauser, Elena, Caser, Luciano, Caramello, Elisa, Sandigliano, Franca, Rosso, Giorgio, Bendini, Chiara, Davoli, Maria, Casella, Monica, Prampolini, Raffaella, Scevola, Moreno, Vitale, Enrico, Brugiolo, Roberto, Brugiolo, Fagherazzi, Carlo, Scarpa, Alberto, Zara, Daniela, Bertozzo, Giulia, Granziera, Serena, Barazzuol, Michele, Maugeri, Domenico, Sorace, Rosaria, Anzaldi, Massimiliano, De Gesu Roberto, Morrone, Giuseppe, Davolio, Federica, Fabbo, Andrea, Palmieri, Marina, Forti, Paola, Pirazzoli, Luca, Fabbri, Elisa, Terenzi, Laura, Bergolari, Federica, Wenter, Christian, Ruffini, Ingrid, Insam, Miriam, Abraham, Elisabeth, Kirchlechner, Christine, Lasco, Antonino, Arena Maria Grazia, Parise, Pasquale, Boccali, Andrea, Amici, Serena, Gambacorta, Maurizia, Ferrari, Alberto, Lasagni, Anna, Lovati, Roberto, Giovinazzo, Francesca, Kimak, Elzbieta, Zappa, Paolo, Medici, Francesco, Lo Castro Marika, Mauro, Flavia, De Luca Alessandro, Sancesario, Giuseppe, Martorana, Alessandro, Scaricamazza, Beatrice, Di Lorenzo Francesco, Liguori, Claudio, Vita, Natale, Giomi, Mirna, Forte, Floriana, Padovani, Alessandro, Rozzini, Luca, Caratozzolo, Salvatore, Cottino, Maria, Vitali, Silvia, Marelli, Eleonora, Tripi, Gabriele, Miceli, Salvatore, Urso, Giovanni, Grioni, Giuseppe, Vezzadini, Giuliana, Misaggi, Giulia, Forlani, Chiara, Avanzi, Stefano, Saulle, Serena, Casarini, Claudia, Viggiano, Marilena, Alberto, Luisa, Ghianda, Diego, Giordano, Alessandro, Iemolo, Francesco, Giordano, Antonello, Sanzaro, Enzo, D'Asta, Gabriele, Proietto, Maria, Carnemolla, Anna, Razza, Grazia, Spadaro, Daniela, Bertolotti, Marco, Mussi, Chiara, Neviani, Francesca, Chiesa, Roberto, Guerzoni, Valentina, Morselli, Linda, Venturelli, Francesca, Tarozzi, Adriano, Balestri, Francesca, Mannarino, Giulio, Bigolari, Michela, Natale, Alessia, Grassi, Simona, Bottaro, Cinzia, Stefanelli, Sara, Bovone, Ugo, Tortorolo, Umberto, Quadri, Roberto, Leone, Giuseppe, Ponzetto, Maria, Frasson, Paola, Annoni, Giorgio, Bruni, Adriana, Confalonieri, Roberto, Corsi, Maurizio, Moretti, Daniela, Teruzzi, Fabiola, Umidi, Simona, Mazzola, Paolo, Perego, Sabrina, Persico, Ilaria, Olivieri, Giulia, Bonfanti, Alessandra, Hajnalka, Szabo, Galeazzi, Marianna, Massariello, Francesca, Anzuini, Alessandra, Caffarra, Paolo, Barocco, Federica, Spallazzi, Marco, Ceda Gian Paolo, Morganti, Simonetta, Artoni, Andrea, Chioatto, Paolo, Bortolamei, Sandra, Soattin, Lucia, Beneamino, Borelli, Barilaro, Giuseppe, Bertazzoli, Marco, Rota, Elisabetta, Adobati, Annamaria, Zuccher, Paola, Dal Fabbro Angela, Lo Nigro Ambra, Franchetti, Lorena, Toniolo, Marika, Marcuzzo, Cinzia, Rollone, Marco, Guerriero, Fabio, Sgarlata, Carmelo, Anna, Santoro, Zatti, Giovanni, Piatti, Massimiliano, Graci, Jole, Benati, Giuseppe, Boschi, Federica, Biondi, Mario, Fiumi, Nicoletta, Erika, Tamburini, Locatelli Sergio Mario, Mauri, Sabrina, Beretta, Mauro, Margheritis, Laura, Desideri, Giovanbattista, Liberatore, Ester, Carucci Anna Cecilia, Bonino, Paolo, Caput, Margherita, Antonietti Maria Paola, Polistena, Giuseppe, De la Pierre Franz, Mari, Marcello, Massignani, Paola, Tombesi, Fabio, Selvaggio, Fabio, Verbo, Brunella, Bodoni, Paolo, Marchionni, Niccolo, Sabatini, Tony, Mussio, Eleonora, Magni, Eugenio, Bianchetti, Angelo, Crucitti, Andrea, Titoldini, Giulia, Cossu, Beatrice, Fascendini, Sara, Licini, Cristina, Tomasoni, Angela, Calderazzo, Massimo, Tropiano, Daniela, Lagana, Valentina, Melotti Rita Maria, Lilli, Albina, Buda, Simona, Adversi, Marco, Noro, Gabriele, Turco, Renato, Chiara, Ubezio M., Mantovani Anna Roberta, Viola Maria Cristina, Serrati, Carlo, Pretta, Stefano, Infante, Maria, Gentile, Simona, D'Ambrosio, Viviana, Mazzanti, Paolo, Brambilla, Cristina, Sportelli, Silvia, Platto, Caterina, Faraci, Bianca, Quattrocchi, Daniela, Pisu, Cristina, Sicuro, Francesca, Del Grosso Luciano Luca, Zagnoni, Piergiuseppe, Ghiglia, Stefania, Mosca, Massimiliano, Corazzin, Ileana, Deola, Mariangela, Biagini Carlo Adriano, Bencini, Francesca, Cantini, Claudia, Tonon, Elisabetta, Pierinelli, Silvia, Onofrj, Marco, Thomas, Astrid, Borbone, Filomena, Bonanni, Laura, Cacchio, Gabriella, Comi, Giancarlo, Magnani, Giuseppe, Santangelo, Roberto, Mazzeo, Salvatore, Caso, Francesca, Cecchetti, Giordano, Sant’Angelo, Roberto, Barbieri, Cristina, Giroldi, Liviana, Bandini, Fabio, Masina, Marco, Malservisi, Simona, Cicognani, Annalena, Ricca, Laura, Piccininni, Maristella, Tassinari, Tiziana, Brogi, Davide, Sugo, Annalisa, Filippi, Alessandra, Manfredi, Sonia, Vanni, Valerio, Usai carlo Andrea, Colombo, Enrico, Renna Vera Francesca, Sauchella, Assunta, Prete, Mauro, Bisagni, Pietro, Masini, Roberto, Carrabetta, Salvatore, Barone, Antonella, Razzano, Monica, Imperoli, Giuseppe, Bini, Angela, Serra, Francesco, D’Agostino, Valeria, Gianolla, Federico, Pietrangeli, Lucia, Velardi, Antonella, Di Cello Elisabetta, Rosati, Cristina, Casali, Nadia, Casali, Sessa, Maria, Abruzzi, Luciano, Costanzi, Chiara, Bini, Paolo, Pignata, Michele, Buonaugurio, Enrico, Vollery, Maria, Carrieri, Giovanni, Cioni, Giorgio, Toschi, Andrea, Metra, Marco, Ranieri, Piera, Ceccon, Anna, Magrin, Laura, Marin, Sara, Barbara, Sofia, Ghedini, Laura, Moroni, Matteo, Paragona, Marco, Pallotti Maria Caterina, Rota, Carla, Gottardi, Federica, Tomasoni, Clara, Cappuccio, Melania, Bonini, Giampaolo, Guerini, Stefano, Guerini, Valeria, Merlaj, Lucia, Tovaglieri, Maurizio, Antonio, Grillo, Francesca, Arenare, Mineo, Tonino, Kanah, David, Vianello, Giorgio, Vianello, Piero, Balducci, Ubaldo, Sidoti, Vincenzo, Montanari, Stefano, Murelli Flavio Teodolindo Busonera, Albanese, Paolo, Maselli, Monica, Bolzetta, Francesco, Fabris, Roberto, Durante-Mangoni, Emanuele, Testoni, Michela, Di Stefano Fabio, Seccia, Loredana, Sonzini, Valeria, Fabiano, Maria, Di Giorgio Annabella, De Cosmo Salvatore, Greco, Antonio, D’Onofrio, Grazia, Sancarlo, Daniele, Resta, Gianluca, Girardello, Renzo, Girardello, Minervini, Sergio, Boni, Morena, Vitali, Mariagrazia, Pizzoni, Marina, De Colle Paolo, Frattola, Alessandra, Orlandini, Francesco, La Regina Micaela, Padulo, Filomena, Margheriti, Cristina, Rolano, Dario, Moscatelli, Giancarla, Radaelli, Guido, Montini, Elena, Prete, Cosimo, Novello, Marileda, Bramuzzo, Igor, Bertin, Nicole, Rinaldo, Elena, Paolillo, Ciro, Riccardi, Angela, Benedetti, Claudia, Rizzi, Barbara, Montagna, Francesca, Cutaia, Chiara, Baca, Oliver, Colombo, Mauro, Procino, Giuseppe, Tarara, Rosaria, Dell’Acqua, Davide, Cislaghi, Stefania, Cairati, Marco, Porta, Massimo, Scaglione, Luca, De Feo Martina, Vertolli, Paola, Salvati, Lia, Cinotti, Sandro, De Santis Valentina, Biferi, Erminia, Cheli, Paola, Rebizzo, Romina, Salvatore, Minisola, Luciano, Colangelo, Abete, Pasquale, Liguori, Ilaria, Curcio, Francesca, Spassini, Guglielmo, Engheben, Marco, Rotunno, Sara, Rotunno, Arosio, Paola, Angelini, Claudio, Reggiani, Francesco, Cappelli, Alessandro, Marcheselli, Simona, Fratticci, Lara, Ranzani, Paola, Cesarini, Simonetta, Cerini, Antonella, Gallucci, Fernando, Serra, Emanuela, Sola, Mariolina, Delitala, Alessandro, Pes, Chiara, Lobianco, Giulia, Giani, Alessandro, Famularo, Simone, Sandini, Marta, Pinotti, Enrico, Gianotti, Luca, Battuello, Antonella, Pintore, Giulia, Rossi, Andrea, Rubele, Sofia, Sant, Selena, Vignati, Marco, Clerici, Danila, Rosa, Fabio, Bandirali Maria Pia, Cattaneo, Nicoletta, Boffi, Laura, Avalli, Leonello, Scanziani, Margherita, De Notaris Stefania, Del Santo Pierluigi, Del, Santo, Catalano, Felice, Fogli, Danilo, Di Bella Giovanna, Landi, Francesco, Salini, Sara, Brunetti Maria Angela, Cappa, Giorgetta, Galvagno, Giovanni, Cena, Paola, Bruno, Gerardo, Raspo, Silvio, Semeraro, Pedrotti, Sabrina, Palmieri Vincenzo Ostilio, Palasciano, Giuseppe, Belfiore, Anna, Portincasa, Piero, Sabbà, Carlo, Solfrizzi, Vincenzo, D’Introno, Alessia, Valiani, Vincenzo, Bologna, Carolina, Ciarambino, Tiziana, Magnani, Turelli, Paola, Pazzaglia, Ugo, Rodella, Federica, Piana, Giacomo, Castellano, Maurizio, Garelli, Anna, Garelli, Casella, Elisa, Campana, Federica, Coschigliano Maria Antonietta, Marinangeli, Luigi, Lorico, Fabio, Bazzano, Salvatore, Menculini, Giuseppe, Gelosa, Giorgio, Viviana Ambrogio Teresa, Piras, Valentina, Ciricugno, Andrea, Bollari, Alessandra, Coen, Daniele, Magliola, Renata, Milanesio, Deborah, Muzzulini Carlo Lorenzo, Fogliacco, Paolo, Marinella, Turla, Cotelli Maria Sofia, Bianchi, Marta, Siano, Pietro, Capo, Giuseppe, Napoletano, Rosa, Politi, Cecilia, Mancini, Concetta, Del Buono Corrado, De Bartolomeo Giuseppe, Martinelli, Addolorata, Cefalogli, Carmen, Cozzi, Roberto, Virtuani Angelo Giovanni, Moschettini, Gianfilippo, Mastroianni, Franco, Roglia, Daniela, D'Amico, Gabriella, Palella, Mirella, Endrizzi, Cristina, Trotta, Lucia, Zanetti, Orazio, Terazzi, Emanuela, Sacchetti, Marta, Fleetwood, Thomas, Tondo, Giacomo, Di Fazio Ignazio, Bruni, Andrea, Orsitto, Giuseppe, Fabbro, Emanuela, D’Imporzano, Elena, Casanova, Anna, Bertolio, Serena, Nervo, Silvestri, Roberto, Semproni, Elena, Pintus, Manuela, Aloe, Francesca, Tagliaccica, Angelo., Aloisi G., Marengoni A., Morandi A., Zucchelli A., Cherubini A., Mossello E., Bo M., Di Santo S.G., Mazzone A., Trabucchi M., Cappa S., Fimognari F.L., Incalzi R.A., Gareri P., Perticone F., Campanini M., Montorsi M., Latronico N., Zambon A., Bellelli G., Rispoli V., Malara A., Spadea F., Di Cello S., Ceravolo F., Fabiano F., Chiaradia G., Gabriele A., Lenino P., Andrea T., Settembrini V., Capomolla D., Citrino A., Scriva A., Bruno I., Secchi R., De Martino E., Muccinelli R., Villa Del Sole R.S.A., Lupi G., Paonessa P., Fabbri A., Passuti M.T., Castellari S., Po A., Gaggioli G., Varesi M., Moneti P., Capurso S., Latini V., Ghidotti S., Riccardelli F., Macchi M., Rigo R., Claudio P., Angelo B., Flavio C., Benedetta B., Boffelli S., Cassinadri A., Franzoni S., Spazzini E., Andretto D., Tonini G., Andreani L., Coralli M., Balotta A., Cancelliere R., Ballardini G., Simoncelli M., Mancini A., Strazzacapa M., Fabio S., De Filippi F., Giudice C., Dentizzi C., Azzini M., Cazzadori M., Mastroeni V., Bertassello P., Benati H.S.C., Nesta E., Tobaldini C., Guerini F., Elena T., Mombelloni P., Fontanini F., Gabriella L., Pizzorni C., Oliverio M., Del Grosso L.L., Giavedoni C., Bidoli G., Mazzei B., Corsonello A., Fusco S., Vena S., De Vuono T., Maiuri G., Luca F.F., Andrea A., Giovanni S., Rossella N., Castegnaro E., De Rosa S., Benvenuti E., Del Lungo I., Giardini S., Giulietti C., Mauro D.B., Eleonora B., Martina P., Irene F., Riccardo B., Federica S., Bertoletti E., D'Amico F., Caronzolo F., Grippa A., Lombardo G., Pipicella T., Antonino S., Francesco C., Valeria P.G., Daniela L., Domenico C., Giorgio B., March A., Nitti M.T., Felici A., Pavan S., Piazzani F., Lunelli A., Dimori S., Margotta A., Soglia T., Postacchini D., Brunelli R., Santini S., Francavilla M., Macchiati I., Sorvillo F., Giuli C., Mecocci P., Longo A., Addesi D., Rosa P.C., Bencardino G., Falbo T., Grillo N., Marco F., Mirella F., Fanto F., Isaia G., Pezzilli S., Bergamo D., Furno E., Rrodhe S., Lucarini S., Dijk B., Dall'Acqua F., Calvani D., Becheri D., Giuseppe M., Costanza M., Vito A., Francesca B., Magherini L., Novella M., Franca B., Gambardella P.M.L., Valente C., Ospedale N., Ilaria B., Alice F., Porrino P., Ceci G., Giuliana B., Michela T., Eleonora C., Ettore E., Camellini C., Servello A., Grassi A., Rozzini R., Tironi S., Grassi M.G., Troisi E., Carlo C., Gabriella D.S.S., Flaminia F., Federica R., Beatrice P., Sofia T., Gabutto A., Quazzo L., Rosatello A., Suraci D., Tagliabue B., Perrone C., Ferrara L., Castagna A., Tremolada M.L., Giuseppe C., Stefano B., Davide O., Piano S., Serviddio G., Lo Buglio A., Gurrera T., Merlo V., Rovai C., Cotroneo A.M., Carlucci R., Abbaldo A., Monzani F., Qasem A.A., Bini G., Tafuto S., Galli G., Bruni A.C., Mancuso G., Calipari D., Bernardini B., Corsini C., Michele C., Sara D.F., Cagnin A., Fragiacomo F., Pompanin S., Piero A., Marco C., Zurlo A., Guerra G., Pala M., Menozzi L., Delli Gatti C., Magon S., Roberto M., Alfredo D.G., Fabio F., Ruana T., Elisa M., Christian M., Marco P., Massimo G., Di Francesco V., Faccioli S., Pellizzari L., Giorgia F., Barbagallo G., Lunardelli M.L., Martini E., Ferrari E., Macchiarulo M., Corneli M., Bacci M., Battaglia G., Anastasio L., Lo Storto M.S., Seresin C., Simonato M., Loreggian M., Cestonaro F., Durando M., Latella R., Mazzoleni M., Russo G., Ponte M., Valchera A., Salustri G., Petritola D., Costa A., Sinforiani E., Cotta M.R., Pizio R.N., Cester A., Formilan M., Pietro B., Carbone P., Cazzaniga I., Appollonio I., Cereda D., Stabile A., Xhani R., Acampora R., Tremolizzo L., Federico P., Antonio C., Valerio P., Cesare B., Zhirajr M., Giovanni V., Maria A., Mariaelena S., Bottacchi E., Bucciantini E., Di Giovanni M., Franchi F., Lucchetti L., Mariani C., Grande G., Rapazzini P., Marco M., Romanelli G., Franco N., Alessio M., Nicola L., Laura P., Nazario P., Chiara G., Soccorso P., Andrea S., Luca B., Francesca S., Roberto A., Anna C., Infettive M., Fugazza L., Guerrini C., De Paduanis G., Iallonardo L., Palumbo P., Zuliani G., Ortolani B., Capatti E., Soavi C., Bianchi L., Francesconi D., Miselli A., Gloria B., Tommaso R., Chiara P., Agata M.M., Dall'Agata M., Luca M., Gianluca G., Suardi T., Zaccarini C., Manuela R., Mirra G., Muti E., Bottura R., Gianpaolo M., Secreto P., Bisio E., Cecchettani M., Naldi T., Pallavicino A., Pugliese M., Iozzo R.C., Grassi G., Michele B., Dell'Oro R., Fosca Q.T., Giorgio G.C., Giovanni P., Ernesto C., Mannironi A., Giorli E., Oberti S., Fierro B., Piccoli T., Giacalone F., Mandas A., Serchisu L., Costaggiu D., Pinna E., Orru F., Mannai M., Cordioli Z., Pelizzari L., Turcato E., Arduini P., Cacace C., Chiloiro R., Cimino R., Ruberto C., Giovanni R., Pietro G., Laura G., Alberto C., Carmen R., Dal Santo P., Andriolli A., Burattin G., Rossi L., Antonino C.G.A., Tezza F., Maddalena P., Laura S., Crippa P., Aloisio P., Di Monda T., Malighetti A., Galbassini G., Ivaldi C., Russo A.M., Bennati E., Pino E., Zavarise G., Pesci A., Suigo G., Faverio P., Andrea G., Sabrina P., Zanasi M., Moniello G., Rostagno C., Cartei A., Polidori G., Ungar A., Melis M.R., Martellini E., Enrico M., Monica T., Antonella G., Giovanna L., Migliorini M., Caramelli F., Battiston B., Berardino M., Cavallo S., Alessandro M., Anna S., Lombardi B., D'Ippolito P., Furini A., Villani D., Clara R., Guarneri M., Paolucci S., Bassi A., Coiro P., De Angelis D., Morone G., Venturiero V., Palleschi L., Raganato P., Di Niro G., Rosa C.A., Loredana B., Imoscopi A., Tibaldi V., Bottignole G.G., Calvi E., Clementi C., Zanocchi M., Agosta L., Criasia A., Spertino E., Nortarelli A., Provenzano G., Principato P., Rizzo A., Cellura E., Mari D., Romano F.Y., Rosini F., Mansi M., Rossi S., Riccardelli A., Inzaghi L., Bonini G., Rossi P., Potena A., Lichii M., Candiani T., Grimaldi W., Bertani E., Alessandra P., Calogero P., Pinto D., Bernardi R., Nicolino F., Galetti C., Gianstefani A., Giulia C., Lorenzo M., Odetti P., Monacelli F., Prefumo M., Fiammetta M., Canepa M., Minaglia C., Paolisso G., Rizzo M.R., Prestano R., Dalise A.M., Barra D., Dal Bosco L., Asprinio V., Dallape L., Perina E., Bartoli I.R., Pluderi A., Maina A., Pecoraro E., Sciarra M., Prudente A., Paola M., Francesca M., Manuel V., Luisella C., Maria P.L., Tina S., Benini L., Levato F., Mhiuta V., Alius F., Davidoaia D., Giardini V., Garancini M., Bellamoli C., Terranova L., Bozzini C., Tosoni P., Provoli E., Cascone L., Dioli A., Ferrarin G., Bucci A., Bua G., Fenu S., Bianchi G., Casella S., Romano V., Maurizio P., Mascherona I., Belotti G., Cavaliere S., Cuni E., Merciuc N., Oberti R., Veneziani S., Capoferri E., De Bernardi E., Colombo K., Bravi M., Nicoletta N., D'Arcangelo P., Montenegro N., Montanari R., Lamanna P., Gasperini B., Isabella M., Stefania D., Gaia A., Filippo C., Palama C., Di Emidio C., Scarpini E., Arighi A., Fumagalli G., Basilico P., De Amicis Margherita M., Marta M., Diletta M., Granata A., Ranalli C., Cammilli A., Cavallini M.C., Tricca M., Natella D., Gabbani L., Tesi F., Martella L., Imbrici R., Guerrini G., Scotuzzi A.M., Sozzi F., Valenti L., Chiarello A., Monia M., Pilotto A., Prete C., Senesi B., Meta A.C., Pendenza E., Pasqualetti G., Polini A., Tognini S., Ballino E., Dell'Aquila G., Gasparrini P.M., Marotti E., Migale M., Scrimieri A., Falsetti L., Salvi A., Toigo G., Ceschia G., Rosso A., Tongiorgi C., Scarpa C., De Dominicis L., Pucci E., Renzi S., Cartechini E., Tomassini P.F., Del Gobbo M., Ugenti F., Romeo P., Nardelli A., Lauretani F., Visioli S., Montanari I., Ermini F., Giordano A., Pigato G., Simeone E., Colameco F., Cecamore A., Scurti R., Lupinetti M.C., Barbujani M., Perazzi B., Giampieri M., Amoruso R., Piccinini M., Ferrari C., Gambetti C., Sfrappini M., Semeraro L., Striuli R., Pelliccioni G., Marinelli D., Fabi K., Rossi T., Pesallaccia M., Sabbatini D., Gobbi B., Cerqua R., Tagliani G., Schlauser E., Caser L., Caramello E., Sandigliano F., Rosso G., Ferrari A., Bendini C., Luisa D.M., Casella M., Prampolini R., Scevola M., Vitale E., Roberto B., Carlo F., Sergio F., Alberto S., Daniela Z., Giulia B., Serena G., Maugeri D., Sorace R., Anzaldi M., De Gesu R., Morrone G., Davolio F., Fabbo A., Palmieri M., Zoli M., Forti P., Pirazzoli L., Fabbri E., Terenzi L., Bergolari F., Wenter C., Ruffini I., Insam M., Abraham E., Kirchlechner C., Cucinotta D., Antonino L., Basile G., Grazia A.M., Parise P., Boccali A., Amici S., Gambacorta M., Lasagni A., Lovati R., Giovinazzo F., Kimak E., Zappa P., Medici F., Lo Castro M., Mauro F., De Luca A., Sancesario G., Martorana A., Scaricamazza B., Toniolo S., Di Lorenzo F., Liguori C., Lasco A., Vita N., Giomi M., Forte F., Padovani A., Rozzini L., Ceraso A., Salvatore C., Cottino M., Vitali S., Marelli E., Tripi G., Miceli S., Urso G., Grioni G., Vezzadini G., Misaggi G., Forlani C., Avanzi S., Serena S., Claudia C., Marilena V., Alberto L., Diego G., Alessandro G., Iemolo F., Sanzaro E., D'Asta G., Proietto M., Carnemolla A., Razza G., Spadaro D., Bertolotti M., Mussi C., Neviani F., Roberto C., Valentina G., Linda M., Francesca V., Tarozzi A., Balestri F., Mannarino G., Bigolari M., Natale A., Grassi S., Bottaro C., Stefanelli S., Bovone U., Tortorolo U., Quadri R., Leone G., Ponzetto M., Frasson P., Annoni G., Bruni A., Confalonieri R., Corsi M., Moretti D., Teruzzi F., Umidi S., Mazzola P., Perego S., Persico I., Olivieri G., Bonfanti A., Hajnalka S., Galeazzi M., Massariello F., Anzuini A., Caffarra P., Barocco F., Spallazzi M., Paolo C.G., Simonetta M., Chioatto P., Bortolamei S., Soattin L., Ruotolo G., Beneamino B., Giuseppe B., Bertazzoli M., Rota E., Adobati A., Scarpa A., Granziera S., Zuccher P., Dal Fabbro A., Zara D., Lo Nigro A., Franchetti L., Toniolo M., Marcuzzo C., Rollone M., Guerriero F., Sgarlata C., Masse A., Zatti G., Piatti M., Graci J., Benati G., Boschi F., Biondi M., Fiumi N., Erika T., Locatelli S.M., Mauri S., Beretta M., Margheritis L., Desideri G., Liberatore E., Carucci A.C., Bonino P., Caput M., Antonietti M.P., Polistena G., De La Pierre F., Mari M., Massignani P., Tombesi F., Selvaggio F., Verbo B., Bodoni P., Marchionni N., Sabatini T., Mussio E., Magni E., Bianchetti A., Crucitti A., Titoldini G., Cossu B., Fascendini S., Licini C., Tomasoni A., Calderazzo M., Daniela T., Valentina L., Melotti R.M., Lilli A., Buda S., Adversi M., Noro G., Turco R., Ubezio M.C., Mantovani A.R., Viola M.C., Serrati C., Pretta S., Infante M., Gentile S., D'Ambrosio V., Mazzanti P., Brambilla C., Sportelli S., Platto C., Faraci B., Quattrocchi D., Pernigotti L.M., Pisu C., Sicuro F., Zagnoni P., Ghiglia S., Mosca M., Corazzin I., Deola M., Di Agordo O., Biagini C.A., Bencini F., Cantini C., Tonon E., Pierinelli S., Onofrj M., Thomas A., Filomena B., Bonanni L., Gabriella C., Comi G., Magnani G., Santangelo R., Mazzeo S., Francesca C., Giordano C., Roberto S., Barbieri C., Giroldi L., Bandini F., Masina M., Malservisi S., Cicognani A., Ricca L., Piccininni M., Tassinari T., Brogi D., Sugo A., Alessandra F., Sonia M., Valerio V., Andrea U.C., Enrico C., Vera R.F., Assunta S., Gianmaria Z., Mauro P., Barone A., Razzano M., Giuseppe I., Angela B., Francesco S., D'Agostino V., Federico G., Lucia P., Antonella V., Di Cello E., Cristina C., Nadia C., Maria S., Luciano A., Chiara C., Bini P., Pignata M., Enrico B., Maria V., Giovanni C., Giorgio C., Piera R., Alberto Z., Ceccon A., Magrin L., Marin S., Barbara S., Matteo M., Caterina P.M., Carla R., Federica G., Clara T., Melania C., Giampaolo B., Stefano G., Valeria G., Lucia M., Giovambattista D., Ester L., Cecilia C.A., Maurizio T., Nadia B., Grillo A., Arenare F., Tonino M., David K., Giorgio V.P., Ubaldo B., Vincenzo S., Stefano M., Marino F., Flavio M.T.B., Paolo A., Monica M., Francesco B., Roberto F., Paolo B., Durante-Mangoni E., Testoni M., Di Stefano F., Loredana S., Daniela M., Valeria S., Fabiano M., Di Giorgio A., De Cosmo S., Greco A., D'Onofrio G., Daniele S., Gianluca R., Renzo G., Sergio M., Morena B., Vitali M., Marina P., De Colle P., Cristina S., Orlandini F., La Regina M., Desiree A., Mario B., Paola P., Padulo F., Cristina M., Dario R., Giancarla M., Guido R., Elena M., Marileda N., Igor B., Nicole B., Elena R., Paolillo C., Riccardi A., Claudia B., Barbara R., Silvia V., Oliver B., Mauro C., Eleonora M., Giuseppe P., Rosaria T., Maria C., Dell'Acqua D., Stefania C., Massimo P., Luca S., De Feo M., Paola V., Lia S., Sandro C., De Santis V., Erminia B., Paola C., Romina R., Minisola S., Luciano C., Pasquale A., Ilaria L., Guglielmo S., Marco E., Sara R., Paola A., Claudio A., Francesco R., Alessandro C., Simona M., Lara F., Paola R., Simonetta C., Antonella C., Generoso U., Fernando G., Giuliano C., Emanuela S., Mariolina S., Alessandro D., Giulia L., Famularo S., Sandini M., Pinotti E., Gianotti L., Antonella B., Giulia P., Sante G., Rossi A., Rubele S., Sant S., Marco V., Danila C., Fabio R., Bandirali M.P., Nicoletta C., Laura B., Paolo T., Luciano T., Leonello A., Margherita S., De Notaris S., Del Santo P., Laura R., Fabiana T., Giovanna C., Antonino A., Felice C., Danilo F., Di Bella G., Francesco L., Salini S., Angela B.M., Giorgetta C., Giovanni G., Gerardo B., Silvio R., Letizia S., Davide B., Rosaria R.M., Maria D.A., Raffaele P., Palmieri V.O., Palasciano G., Belfiore A., Portincasa P., Carlo S., D'Introno A., Valiani V., Carolina B., Tiziana C., Paola T., Ugo P., Giacomo P., Castellano M., Anna G., Elisa C., Federica C., Antonietta C.M., Luigi M., Fabio L., Salvatore B., Gelosa G., Viviana A.T., Piras V., Andrea C., Alessandra B., Coen D., Magliola R., Milanesio D., Muzzulini C.L., Paolo F., Marinella T., Sofia C.M., Marta B., Siano P., Capo G., Napoletano R., Cecilia P., Mancini C., Del Buono C., De Bartolomeo G., Addolorata M., Carmen C., Giovanni V.A., Moschettini G., Franco M., Mulli O.F., Daniela R., D'Amico G., Mirella P., Endrizzi C., Trotta L., Ciarambino T., Orazio Z., Emanuela T., Marta S., Thomas F., Giacomo T., Di Fazio I., Andrea B., Giuseppe O., Emanuela F., Serena A., D'Imporzano E., Serena B., Erika N., Elena S., Manuela P., Francesca A., Angelo T., Aloisi, G., Marengoni, A., Morandi, A., Zucchelli, A., Cherubini, A., Mossello, E., Bo, M., Di Santo, S.G., Mazzone, A., Trabucchi, M., Cappa, S., Fimognari, F.L., Incalzi, R.A., Gareri, P., Perticone, F., Campanini, M., Montorsi, M., Latronico, N., Zambon, A., Bellelli, G., Rispoli, V., Malara, A., Spadea, F., Di Cello, S., Ceravolo, F., Fabiano, F., Chiaradia, G., Gabriele, A., Lenino, P., Andrea, T., Settembrini, V., Capomolla, D., Citrino, A., Scriva, A., Bruno, I., Secchi, R., De Martino, E., Muccinelli, R., Villa Del Sole, R.S.A., Lupi, G., Paonessa, P., Fabbri, A., Passuti, M.T., Castellari, S., Po, A., Gaggioli, G., Varesi, M., Moneti, P., Capurso, S., Latini, V., Ghidotti, S., Riccardelli, F., Macchi, M., Rigo, R., Claudio, P., Angelo, B., Flavio, C., Benedetta, B., Boffelli, S., Cassinadri, A., Franzoni, S., Spazzini, E., Andretto, D., Tonini, G., Andreani, L., Coralli, M., Balotta, A., Cancelliere, R., Ballardini, G., Simoncelli, M., Mancini, A., Strazzacapa, M., Fabio, S., De Filippi, F., Giudice, C., Dentizzi, C., Azzini, M., Cazzadori, M., Mastroeni, V., Bertassello, P., Benati, H.S.C., Nesta, E., Tobaldini, C., Guerini, F., Elena, T., Mombelloni, P., Fontanini, F., Gabriella, L., Pizzorni, C., Oliverio, M., Del Grosso, L.L., Giavedoni, C., Bidoli, G., Mazzei, B., Corsonello, A., Fusco, S., Vena, S., De Vuono, T., Maiuri, G., Luca, F.F., Andrea, A., Giovanni, S., Rossella, N., Castegnaro, E., De Rosa, S., Benvenuti, E., Del Lungo, I., Giardini, S., Giulietti, C., Mauro, D.B., Eleonora, B., Martina, P., Irene, F., Riccardo, B., Federica, S., Bertoletti, E., D'Amico, F., Caronzolo, F., Grippa, A., Lombardo, G., Pipicella, T., Antonino, S., Francesco, C., Valeria, P.G., Daniela, L., Domenico, C., Giorgio, B., March, A., Nitti, M.T., Felici, A., Pavan, S., Piazzani, F., Lunelli, A., Dimori, S., Margotta, A., Soglia, T., Postacchini, D., Brunelli, R., Santini, S., Francavilla, M., Macchiati, I., Sorvillo, F., Giuli, C., Mecocci, P., Longo, A., Addesi, D., Rosa, P.C., Bencardino, G., Falbo, T., Grillo, N., Marco, F., Mirella, F., Fantò, F., Isaia, G., Pezzilli, S., Bergamo, D., Furno, E., Rrodhe, S., Lucarini, S., Dijk, B., Dall'Acqua, F., Calvani, D., Becheri, D., Giuseppe, M., Costanza, M., Vito, A., Francesca, B., Magherini, L., Novella, M., Franca, B., Gambardella, P.M.L., Valente, C., Ospedale, N., Ilaria, B., Alice, F., Porrino, P., Ceci, G., Giuliana, B., Michela, T., Eleonora, C., Ettore, E., Camellini, C., Servello, A., Grassi, A., Rozzini, R., Tironi, S., Grassi, M.G., Troisi, E., Carlo, C., Gabriella, D.S.S., Flaminia, F., Federica, R., Beatrice, P., Sofia, T., Gabutto, A., Quazzo, L., Rosatello, A., Suraci, D., Tagliabue, B., Perrone, C., Ferrara, L., Castagna, A., Tremolada, M.L., Giuseppe, C., Stefano, B., Davide, O., Piano, S., Serviddio, G., Lo Buglio, A., Gurrera, T., Merlo, V., Rovai, C., Cotroneo, A.M., Carlucci, R., Abbaldo, A., Monzani, F., Qasem, A.A., Bini, G., Tafuto, S., Galli, G., Bruni, A.C., Mancuso, G., Mancuso, G., Calipari, D., Bernardini, B., Corsini, C., Michele, C., Sara, D.F., Cagnin, A., Fragiacomo, F., Pompanin, S., Piero, A., Marco, C., Zurlo, A., Guerra, G., Pala, M., Menozzi, L., Delli Gatti, C., Magon, S., Roberto, M., Alfredo, D.G., Fabio, F., Ruana, T., Elisa, M., Benedetta, B., Christian, M., Marco, P., Massimo, G., Di Francesco, V., Faccioli, S., Pellizzari, L., Giorgia, F., Barbagallo, G., Lunardelli, M.L., Martini, E., Ferrari, E., Macchiarulo, M., Corneli, M., Bacci, M., Battaglia, G., Anastasio, L., Lo Storto, M.S., Seresin, C., Simonato, M., Loreggian, M., Cestonaro, F., Durando, M., Latella, R., Mazzoleni, M., Russo, G., Ponte, M., Valchera, A., Salustri, G., Petritola, D., Costa, A., Sinforiani, E., Cotta, M.R., Piano, S., Pizio, R.N., Cester, A., Formilan, M., Pietro, B., Carbone, P., Cazzaniga, I., Appollonio, I., Cereda, D., Stabile, A., Xhani, R., Acampora, R., Tremolizzo, L., Federico, P., Antonio, C., Valerio, P., Cesare, B., Zhirajr, M., Giovanni, V., Maria, A., Mariaelena, S., Bottacchi, E., Bucciantini, E., Di Giovanni, M., Franchi, F., Lucchetti, L., Mariani, C., Grande, G., Rapazzini, P., Marco, M., Romanelli, G., Franco, N., Alessio, M., Stefano, B., Nicola, L., Laura, P., Nazario, P., Carlo, C., Chiara, G., Soccorso, P., Andrea, S., Luca, B., Francesca, S., Roberto, A., Marco, F., Anna, C., Francesco, C., Infettive, M., Fugazza, L., Guerrini, C., De Paduanis, G., Iallonardo, L., Palumbo, P., Zuliani, G., Ortolani, B., Capatti, E., Soavi, C., Bianchi, L., Francesconi, D., Miselli, A., Gloria, B., Tommaso, R., Chiara, P., Agata, M.M., Dall'Agata, M., Luca, M., Gianluca, G., Suardi, T., Zaccarini, C., Manuela, R., Mirra, G., Muti, E., Bottura, R., Gianpaolo, M., Secreto, P., Bisio, E., Cecchettani, M., Naldi, T., Pallavicino, A., Pugliese, M., Iozzo, R.C., Grassi, G., Michele, B., Dell'Oro, R., Fosca, Q.T., Giorgio, G.C., Giovanni, P., Ernesto, C., Mannironi, A., Giorli, E., Oberti, S., Fierro, B., Piccoli, T., Giacalone, F., Mandas, A., Serchisu, L., Costaggiu, D., Pinna, E., Orrù, F., Mannai, M., Cordioli, Z., Pelizzari, L., Turcato, E., Arduini, P., Cacace, C., Chiloiro, R., Cimino, R., Ruberto, C., Giovanni, R., Pietro, G., Laura, G., Alberto, C., Carmen, R., Dal Santo, P., Andriolli, A., Burattin, G., Rossi, L., Antonino, C.G.A., Giuseppe, C., Tezza, F., Maddalena, P., Laura, S., Crippa, P., Aloisio, P., Di Monda, T., Malighetti, A., Galbassini, G., Ivaldi, C., Russo, A.M., Bennati, E., Pino, E., Zavarise, G., Pesci, A., Suigo, G., Faverio, P., Andrea, G., Sabrina, P., Zanasi, M., Moniello, G., Rostagno, C., Cartei, A., Polidori, G., Ungar, A., Melis, M.R., Martellini, E., Enrico, M., Monica, T., Antonella, G., Giovanna, L., Migliorini, M., Caramelli, F., Battiston, B., Berardino, M., Cavallo, S., Alessandro, M., Anna, S., Lombardi, B., D'Ippolito, P., Furini, A., Villani, D., Clara, R., Guarneri, M., Paolucci, S., Bassi, A., Coiro, P., De Angelis, D., Morone, G., Venturiero, V., Palleschi, L., Raganato, P., Di Niro, G., Rosa, C.A., Loredana, B., Imoscopi, A., Isaia, G., Tibaldi, V., Bottignole, G.G., Calvi, E., Clementi, C., Zanocchi, M., Agosta, L., Criasia, A., Spertino, E., Nortarelli, A., Provenzano, G., Principato, P., Rizzo, A., Cellura, E., Mari, D., Romano, F.Y., Rosini, F., Mansi, M., Rossi, S., Riccardelli, A., Inzaghi, L., Bonini, G., Rossi, P., Potena, A., Lichii, M., Candiani, T., Grimaldi, W., Bertani, E., Alessandra, P., Calogero, P., Pinto, D., Bernardi, R., Nicolino, F., Galetti, C., Gianstefani, A., Giulia, C., Lorenzo, M., Odetti, P., Monacelli, F., Prefumo, M., Fiammetta, M., Canepa, M., Minaglia, C., Paolisso, G., Rizzo, M.R., Prestano, R., Dalise, A.M., Barra, D., Dal Bosco, L., Asprinio, V., Dallape, L., Perina, E., Bartoli, I.R., Pluderi, A., Maina, A., Pecoraro, E., Sciarra, M., Prudente, A., Paola, M., Francesca, M., Manuel, V., Luisella, C., Maria, P.L., Tina, S., Benini, L., Levato, F., Mhiuta, V., Alius, F., Davidoaia, D., Giardini, V., Garancini, M., Bellamoli, C., Terranova, L., Bozzini, C., Tosoni, P., Provoli, E., Cascone, L., Dioli, A., Ferrarin, G., Bucci, A., Bua, G., Fenu, S., Bianchi, G., Casella, S., Romano, V., Maurizio, P., Mascherona, I., Belotti, G., Cavaliere, S., Cuni, E., Merciuc, N., Oberti, R., Veneziani, S., Capoferri, E., De Bernardi, E., Colombo, K., Bravi, M., Nicoletta, N., D'Arcangelo, P., Montenegro, N., Galli, G., Montanari, R., Lamanna, P., Gasperini, B., Isabella, M., Stefania, D., Gaia, A., Filippo, C., Palamà, C., Di Emidio, C., Scarpini, E., Arighi, A., Fumagalli, G., Basilico, P., De Amicis Margherita, M., Marta, M., Diletta, M., Granata, A., Ranalli, C., Cammilli, A., Cavallini, M.C., Tricca, M., Natella, D., Gabbani, L., Tesi, F., Martella, L., Imbrici, R., Guerrini, G., Scotuzzi, A.M., Sozzi, F., Valenti, L., Chiarello, A., Monia, M., Pilotto, A., Prete, C., Senesi, B., Meta, A.C., Pendenza, E., Pasqualetti, G., Polini, A., Tognini, S., Ballino, E., Dell'Aquila, G., Gasparrini, P.M., Marotti, E., Migale, M., Scrimieri A., Falsetti, L., Salvi A., Toigo, G., Ceschia, G., Rosso, A., Tongiorgi, C., Scarpa, C., Maurizio, P., De Dominicis, L., Pucci, E., Renzi, S., Cartechini, E., Tomassini, P.F., Del Gobbo, M., Ugenti, F., Romeo, P., Nardelli, A., Lauretani, F., Visioli, S., Montanari, I., Ermini, F., Giordano, A., Pigato, G., Simeone, E., Colameco, F., Cecamore, A., Scurti, R., Lupinetti, M.C., Barbujani, M., Perazzi, B., Giampieri, M., Amoruso, R., Piccinini, M., Ferrari, C., Gambetti, C., Sfrappini, M., Semeraro, L., Striuli, R., Mariani, C., Pelliccioni, G., Marinelli, D., Fabi, K., Rossi, T., Pesallaccia, M., Sabbatini, D., Gobbi, B., Cerqua, R., Tagliani, G., Schlauser, E., Caser, L., Caramello, E., Sandigliano, F., Rosso, G., Ferrari, A., Bendini, C., Luisa, D.M., Casella, M., Prampolini, R., Scevola, M., Vitale, E., Roberto, B., Carlo, F., Sergio, F., Alberto, S., Daniela, Z., Giulia, B., Serena, G., Michele, B., Maugeri, D., Sorace, R., Anzaldi, M., De Gesu, R., Morrone, G., Davolio, F., Fabbo, A., Palmieri, M., Barbagallo, G., Zoli, M., Forti, P., Pirazzoli, L., Fabbri, E., Terenzi, L., Bergolari, F., Wenter, C., Ruffini, I., Insam, M., Abraham, E., Kirchlechner, C., Cucinotta, D., Antonino, L., Basile, G., Grazia, A.M., Parise, P., Boccali, A., Amici, S., Gambacorta, M., Lasagni, A., Lovati, R., Giovinazzo, F., Kimak, E., Zappa, P., Medici, F., Lo Castro, M., Mauro, F., De Luca, A., Sancesario, G., Martorana, A., Scaricamazza, B., Toniolo, S., Di Lorenzo, F., Liguori, C., Lasco, A., Vita, N., Giomi, M., Forte, F., Padovani, A., Rozzini, L., Ceraso, A., Salvatore, C., Cottino, M., Vitali, S., Marelli, E., Tripi, G., Miceli, S., Urso, G., Grioni, G., Vezzadini, G., Misaggi, G., Forlani, C., Avanzi, S., Serena, S., Claudia, C., Marilena, V., Alberto, L., Diego, G., Alessandro, G., Iemolo, F., Giordano, A., Sanzaro, E., D'Asta, G., Proietto, M., Carnemolla, A., Razza, G., Spadaro, D., Bertolotti, M., Mussi, C., Neviani, F., Roberto, C., Valentina, G., Linda, M., Francesca, V., Tarozzi, A., Balestri, F., Mannarino, G., Bigolari, M., Natale, A., Grassi, S., Bottaro, C., Stefanelli, S., Bovone, U., Tortorolo, U., Quadri, R., Leone, G., Ponzetto, M., Frasson, P., Annoni, G., Bruni, A., Confalonieri, R., Corsi, M., Moretti, D., Teruzzi, F., Umidi, S., Mazzola, P., Perego, S., Persico, I., Olivieri, G., Bonfanti, A., Hajnalka, S., Galeazzi, M., Massariello, F., Anzuini, A., Caffarra, P., Barocco, F., Spallazzi, M., Paolo, C.G., Simonetta, M., Andrea, A., Chioatto, P., Bortolamei, S., Soattin, L., Ruotolo, G., Beneamino, B., Giuseppe, B., Bertazzoli, M., Rota, E., Adobati, A., Scarpa, A., Granziera, S., Zuccher, P., Dal Fabbro, A., Zara, D., Lo Nigro, A., Franchetti, L., Toniolo, M., Marcuzzo, C., Piano, S., Rollone, M., Guerriero, F., Sgarlata, C., Massè, A., Zatti, G., Piatti, M., Graci, J., Benati, G., Boschi, F., Biondi, M., Fiumi, N., Erika, T., Locatelli, S.M., Mauri, S., Beretta, M., Margheritis, L., Desideri, G., Liberatore, E., Carucci, A.C., Bonino, P., Caput, M., Antonietti, M.P., Polistena, G., De La Pierre, F., Mari, M., Massignani, P., Tombesi, F., Selvaggio, F., Verbo, B., Bodoni, P., Marchionni, N., Sabatini, T., Mussio, E., Magni, E., Bianchetti, A., Crucitti, A., Titoldini, G., Cossu, B., Fascendini, S., Licini, C., Tomasoni, A., Calderazzo, M., Daniela, T., Valentina, L., Prampolini, R., Melotti, R.M., Lilli, A., Buda, S., Adversi, M., Noro, G., Turco, R., Ubezio, M.C., Mantovani, A.R., Viola, M.C., Serrati, C., Pretta, S., Infante, M., Gentile, S., D'Ambrosio, V., Mazzanti, P., Brambilla, C., Sportelli, S., Platto, C., Faraci, B., Quattrocchi, D., Pernigotti, L.M., Pisu, C., Sicuro, F., Del Grosso, L.L., Zagnoni, P., Ghiglia, S., Mosca, M., Corazzin, I., Deola, M., Di Agordo, O., Biagini, C.A., Bencini, F., Cantini, C., Tonon, E., Pierinelli, S., Onofrj, M., Thomas, A., Filomena, B., Bonanni, L., Gabriella, C., Comi, G., Magnani, G., Santangelo, R., Mazzeo, S., Giuseppe, M., Francesca, C., Giordano, C., Roberto, S., Barbieri, C., Giroldi, L., Bandini, F., Masina, M., Malservisi, S., Cicognani, A., Ricca, L., Ricca, L., Piccininni, M., Tassinari, T., Brogi, D., Sugo, A., Alessandra, F., Sonia, M., Valerio, V., Andrea, U.C., Enrico, C., Vera, R.F., Assunta, S., Gianmaria, Z., Mauro, P., Pietro, B., Roberto, M., Salvatore, C., Barone, A., Razzano, M., Giuseppe, I., Angela, B., Francesco, S., D'Agostino, V., Federico, G., Lucia, P., Antonella, V., Di Cello, E., Cristina, C., Nadia, C., Maria, S., Luciano, A., Chiara, C., Bini, P., Pignata, M., Enrico, B., Maria, V., Giovanni, C., Giorgio, C., Andrea, T., Marco, M., Piera, R., Alberto, Z., Ceccon, A., Magrin, L., Marin, S., Barbara, S., Marco, M., Laura, G., Matteo, M., Marco, P., Caterina, P.M., Carla, R., Federica, G., Clara, T., Melania, C., Giampaolo, B., Stefano, G., Valeria, G., Lucia, M., Giovambattista, D., Ester, L., Cecilia, C.A., Maurizio, T., Vera, R.F., Nadia, B., Grillo, A., Arenare, F., Tonino, M., David, K., Giorgio, V.P., Ubaldo, B., Vincenzo, S., Stefano, M., Marino, F., Flavio, M.T.B., Paolo, A., Monica, M., Francesco, B., Roberto, F., Paolo, B., Durante-Mangoni, E., Testoni, M., Di Stefano, F., Loredana, S., Daniela, M., Valeria, S., Fabiano, M., Di Giorgio, A., De Cosmo, S., Greco, A., D'Onofrio, G., Daniele, S., Gianluca, R., Renzo, G., Sergio, M., Morena, B., Vitali, M., Marina, P., De Colle, P., Cristina, S., Alessandra, F., Orlandini, F., La Regina, M., Desirée, A., Mirella, F., Mario, B., Paola, P., Padulo, F., Cristina, M., Dario, R., Giancarla, M., Guido, R., Elena, M., Prete, C., Marileda, N., Igor, B., Nicole, B., Elena, R., Paolillo, C., Riccardi, A., Claudia, B., Barbara, R., Francesca, M., Silvia, V., Chiara, C., Oliver, B., Mauro, C., Eleonora, M., Giuseppe, P., Rosaria, T., Maria, C., Dell'Acqua, D., Stefania, C., Marco, C., Massimo, P., Luca, S., De Feo, M., Paola, V., Lia, S., Sandro, C., De Santis, V., Erminia, B., Paola, C., Romina, R., Minisola, S., Luciano, C., Pasquale, A., Ilaria, L., Francesca, C., Guglielmo, S., Marco, E., Sara, R., Paola, A., Claudio, A., Francesco, R., Alessandro, C., Simona, M., Lara, F., Paola, R., Simonetta, C., Antonella, C., Generoso, U., Fernando, G., Giuliano, C., Emanuela, S., Mariolina, S., Alessandro, D., Chiara, P., Giulia, L., Alessandro, G., Famularo, S., Sandini, M., Pinotti, E., Gianotti, L., Antonella, B., Giulia, P., Sante, G., Rossi, A., Rubele, S., Sant, S., Marco, V., Danila, C., Fabio, R., Bandirali, M.P., Nicoletta, C., Laura, B., Paolo, T., Luciano, T., Leonello, A., Margherita, S., De Notaris, S., Del Santo, P., Laura, R., Fabiana, T., Giovanna, C., Antonino, A., Felice, C., Giuseppe, B., Danilo, F., Di Bella, G., Francesco, L., Salini, S., Angela, B.M., Giorgetta, C., Giovanni, G., Paola, C., Gerardo, B., Silvio, R., Letizia, S., Sabrina, P., Davide, B., Rosaria, R.M., Maria, D.A., Raffaele, P., Palmieri, V.O., Palasciano, G., Belfiore, A., Portincasa, P., Carlo, S., Vincenzo, S., D'Introno, A., Valiani, V., Carolina, B., Tiziana, C., Giordano, C., Paola, T., Ugo, P., Federica, R., Giacomo, P., Castellano, M., Anna, G., Elisa, C., Federica, C., Antonietta, C.M., Luigi, M., Fabio, L., Salvatore, B., Giuseppe, M., Gelosa, G., Viviana, A.T., Piras, V., Andrea, C., Alessandra, B., Coen, D., Magliola, R., Milanesio, D., Muzzulini, C.L., Paolo, F., Marinella, T., Sofia, C.M., Marta, B., Siano, P., Capo, G., Napoletano, R., Cecilia, P., Mancini, C., Del Buono, C., De Bartolomeo, G., Addolorata, M., Carmen, C., Roberto, C., Giovanni, V.A., Moschettini, G., Franco M., Mulli, O.F., Daniela, R., D'Amico, G., Mirella, P., Endrizzi, C., Trotta, L., Ciarambino, T., Orazio, Z., Emanuela, T., Marta, S., Thomas, F., Giacomo, T., Di Fazio, I., Andrea, B., Giuseppe, O., Emanuela, F., Serena, A., D'Imporzano, E., Anna, C., Serena, B., Erika, N., Roberto, S., Elena, S., Manuela, P., Francesca, A., Angelo, T., Aloisi, G, Marengoni, A, Morandi, A, Zucchelli, A, Cherubini, A, Mossello, E, Bo, M, Di Santo, S, Mazzone, A, Trabucchi, M, Cappa, S, Fimognari, F, Incalzi, R, Gareri, P, Perticone, F, Campanini, M, Montorsi, M, Latronico, N, Zambon, A, Bellelli, G, and Faverio, P

Subjects
Male, diagnosis, medications, 0302 clinical medicine, prevention, Aged, 80 and over, Delirium, Drug Prescriptions, Female, Hospital Departments, Humans, Prevalence, Polypharmacy, Prescription Drugs, 80 and over, Medicine, LS4_4, 030212 general & internal medicine, Acute hospital, media_common, Confounding, elderly patients, hip fracture, hospitalized patients, prediction rule, risk factor, dementia, events, medications, prevention, diagnosis, risk-factor, risk factor, hip fracture, Psychiatry and Mental Health, medicine.symptom, events, Drug, medicine.medical_specialty, hospitalized-patients, media_common.quotation_subject, MEDLINE, Socio-culturale, elderly-patients, elderly patients, behavioral disciplines and activities, 03 medical and health sciences, Internal medicine, mental disorders, Dementia, Medical prescription, prediction rule, Aged, business.industry, hospitalized patients, medicine.disease, nervous system diseases, dementia, business, 030217 neurology & neurosurgery
Abstract

Objective This study aimed to evaluate the association between polypharmacy and delirium, the association of specific drug categories with delirium, and the differences in drug-delirium association between medical and surgical units and according to dementia diagnosis. Methods Data were collected during 2 waves of Delirium Day, a multicenter delirium prevalence study including patients (aged 65 years or older) admitted to acute and long-term care wards in Italy (2015-2016); in this study, only patients enrolled in acute hospital wards were selected (n = 4,133). Delirium was assessed according to score on the 4 "A's" Test. Prescriptions were classified by main drug categories; polypharmacy was defined as a prescription of drugs from 5 or more classes. Results Of 4,133 participants, 969 (23.4%) had delirium. The general prevalence of polypharmacy was higher in patients with delirium (67.6% vs 63.0%, P = .009) but varied according to clinical settings. After adjustment for confounders, polypharmacy was associated with delirium only in patients admitted to surgical units (OR = 2.9; 95% CI, 1.4-6.1). Insulin, antibiotics, antiepileptics, antipsychotics, and atypical antidepressants were associated with delirium, whereas statins and angiotensin receptor blockers exhibited an inverse association. A stronger association was seen between typical and atypical antipsychotics and delirium in subjects free from dementia compared to individuals with dementia (typical: OR = 4.31; 95% CI, 2.94-6.31 without dementia vs OR = 1.64; 95% CI, 1.19-2.26 with dementia; atypical: OR = 5.32; 95% CI, 3.44-8.22 without dementia vs OR = 1.74; 95% CI, 1.26-2.40 with dementia). The absence of antipsychotics among the prescribed drugs was inversely associated with delirium in the whole sample and in both of the hospital settings, but only in patients without dementia. Conclusions Polypharmacy is significantly associated with delirium only in surgical units, raising the issue of the relevance of medication review in different clinical settings. Specific drug classes are associated with delirium depending on the clinical setting and dementia diagnosis, suggesting the need to further explore this relationship.

Published
2019

14. Co-occurring malformations of cortical development and SCN1A gene mutations

Author

Antonia Parmeggiani, Renzo Guerrini, Roland Coras, Anna Galuppi, Elena Parrini, Tom Pieper, Thomas Schmitt-Mechelke, Gerhard Kluger, Pasquale Striano, Flavio Giordano, Carmen Barba, Dana Craiu, Ingmar Blümcke, C. Barba, E. Parrini, R.Cora, A. Galuppi, D. Craiu, G. Kluger, A. Parmeggiani, T. Pieper, T. Schmitt-Mechelke, P. Striano, F. Giordano, I. Blumcke, and and R. Guerrini

Subjects
Male, Pathology, medicine.medical_specialty, Adolescent, Mutation, Missense, Neuropathology, SCN1A mutations, Focal cortical dysplasia, Epilepsy, SCN1A mutation, Dravet syndrome, Malformation of cortical development, Periventricular nodular heterotopia, Cerebral Cortex, Child, Child, Preschool, Female, Humans, NAV1.1 Voltage-Gated Sodium Channel, RNA Splice Sites, Neurology (clinical), Neurology, medicine, Epilepsy surgery, Preschool, business.industry, Seizure types, Cortical dysplasia, medicine.disease, Dysplasia, Mutation, Abnormality, Missense, business
Abstract

Summary Objective To report on six patients with SCN1A mutations and malformations of cortical development (MCDs) and describe their clinical course, genetic findings, and electrographic, imaging, and neuropathologic features. Methods Through our database of epileptic encephalopathies, we identified 120 patients with SCN1A mutations, of which 4 had magnetic resonance imaging (MRI) evidence of MCDs. We collected two further similar observations through the European Task-force for Epilepsy Surgery in Children. Results The study group consisted of five males and one female (mean age 7.4 ± 5.3 years). All patients exhibited electroclinical features consistent with the Dravet syndrome spectrum, cognitive impairment, and autistic features. Sequencing analysis of the SCN1A gene detected two missense, two truncating, and two splice-site mutations. Brain MRI revealed bilateral periventricular nodular heterotopia (PNH) in two patients and focal cortical dysplasia (FCD) in three, and disclosed no macroscopic abnormality in one. In the MRI-negative patient, neuropathologic study of the whole brain performed after sudden unexpected death in epilepsy (SUDEP), revealed multifocal micronodular dysplasia in the left temporal lobe. Two patients with FCD underwent epilepsy surgery. Neuropathology revealed FCD type IA and type IIA. Their seizure outcome was unfavorable. All four patients with FCD exhibited multiple seizure types, which always included complex partial seizures, the area of onset of which co-localized with the region of structural abnormality. Significance MCDs and SCN1A gene mutations can co-occur. Although epidemiology does not support a causative role for SCN1A mutations, loss or impaired protein function combined with the effect of susceptibility factors and genetic modifiers of the phenotypic expression of SCN1A mutations might play a role. MCDs, particularly FCD, can influence the electroclinical phenotype in patients with SCN1A-related epilepsy. In patients with MCDs and a history of polymorphic seizures precipitated by fever, SCN1A gene testing should be performed before discussing any epilepsy surgery option, due to the possible implications for outcome.

Published
2014

15. [D-Pen(p-tBuBzl)5]NPS, a novel ligand for the neuropeptide S receptor: structure activity and pharmacological studies

Author

Chiara Ruzza, S. Salvadori, P. Grieco, Remo Guerrini, Davide Malfacini, Michela Pela, Girolamo Calo, Claudio Trapella, L. del Zoppo, C., Ruzza, L., del Zoppo, D., Malfacini, M., Pelà, C., Trapella, Grieco, Paolo, S., Salvadori, G., Calò, and R., Guerrini

Subjects
Chemistry, Stereochemistry, Organic Chemistry, technology, industry, and agriculture, Ligand (biochemistry), In vitro, SAR study, NPSR antagonist, Biochemistry, Neuropeptide S, In vivo, Structure–activity relationship, General Pharmacology, Toxicology and Pharmaceutics, Receptor, Neuropeptide S receptor, G protein-coupled receptor
Abstract

Neuropeptide S (NPS) regulates different biological functions by selectively activating a G protein coupled receptor named NPSR. Previous studies identified [d-Pen5]NPS as a NPSR antagonist. This study investigated the structure activity relationship of [d-Pen5]NPS by alkylation of the thiol group of d-Pen with different aliphatic and aromatic moieties. Nine compounds were synthesised and assayed in calcium mobilization studies, performed in HEK293 cells expressing the recombinant mouse NPSR. All compounds behaved as pure NPSR antagonists of moderate to high potency, in a similar manner to the reference peptide. The most potent compounds were obtained using alkylating groups as cyclic aromatic moieties. [d-Pen(p- t BuBzl)5]NPS, the best compound of this series, was further investigated in vivo, where it fully prevented the stimulatory effects of supraspinal NPS on mouse locomotor activity. [d-Pen(p- t BuBzl)5]NPS is proposed as a novel pharmacological tool which could be useful to investigate in vitro and in vivo biological functions regulated by the NPS/NPSR receptor system.

Published
2014

16. Racemic synthesis and solid phase peptide synthesis application of the chimeric Valine/Leucine derivative 2-amino-3,3,4-trimethyl-pentanoic acid

Author

Pela', Michela, Del Zoppo, L., Allegri, L., Marzola, Erika, Trapella, Claudio, Ruzza, Chiara, Calo', Girolamo, Perissutti, E., Frecentese, F., Salvadori, Severo, Guerrini, Remo, M., Pelà, L., Del Zoppo, L., Allegri, E., Marzola, C., Trapella, C., Ruzza, G., Calo???, Perissutti, Elisa, Frecentese, Francesco, S., Salvadori, and R., Guerrini

Subjects
Solid phase peptide synthesis, Leucine, Valine
Abstract

The synthesis of non natural amino acid 2-​amino-​3,​3,​4-​trimethylpentanoic acid (Ipv)​, ready for solid phase peptide synthesis, has been developed. Copper(I) chloride Michael addn., followed by a Curtius rearrangement are the key steps for the Ipv synthesis. The racemic valine​/leucine chimeric amino acid was then successfully inserted in position 5 of neuropeptide S (NPS) and the diastereomeric mixt. sepd. by reverse phase HPLC. The two diastereomeric NPS derivs. were tested for intracellular calcium mobilization using HEK293 cells stably expressing the mouse NPS receptor where they behaved as partial agonist and pure antagonist.

Published
2014

17. Clinical and polygraphic study of familial paroxysmal kinesigenic dyskinesia with PRRT2 mutation

Author

Paolo Tinuper, Renzo Guerrini, Francesca Bisulli, Antonio E. Elia, Lidia Di Vito, Davide Mei, Carla Marini, Margherita Fabbri, M. Fabbri, C. Marini, F. Bisulli, L. Di Vito, A. Elia, R. Guerrini, D. Mei, and P. Tinuper

Subjects
Adult, medicine.medical_specialty, Pediatrics, Neurology, Adolescent, Video Recording, Nerve Tissue Proteins, Frameshift mutation, Epilepsy, ATP1A2, Chorea, Gene duplication, otorhinolaryngologic diseases, medicine, Humans, Paroxysmal kinesigenic dyskinesia, Genetics, Aged, 80 and over, Polymorphism, Genetic, business.industry, Female prevalence, Membrane Proteins, General Medicine, Paroxysmal dyskinesia, Middle Aged, medicine.disease, Pedigree, Dystonia, Mutation (genetic algorithm), Mutation, Female, PRRT2, Neurology (clinical), business
Abstract

Paroxysmal kinesigenic dyskinesia is a neurological condition characterised by brief attacks of involuntary movements triggered by sudden voluntary movements.We describe the clinical, polygraphic, and genetic features of an Italian family with paroxysmal kinesigenic dyskinesia.Paroxysmal kinesigenic dyskinesia manifested as brief choreoathetosic-dystonic attacks precipitated by sudden movements, varying in severity and frequency, amongst the four affected family members. The disorder follows an autosomal dominant transmission and affects female members. Mutation of SLC2A1, MR1, CACNA1A, and ATP1A2 genes was excluded by direct sequencing. Mutation analysis of the PRRT2 gene revealed a single nucleotide duplication, c.649dupC, resulting in the frameshift mutation p.Arg217Profs*8 in all affected members.Paroxysmal kinesigenic dyskinesia is the most common type of paroxysmal movement disorder and is often misdiagnosed clinically as epilepsy. We describe a family with paroxysmal kinesigenic dyskinesia associated with PRRT2 gene mutation, mild intrafamilial clinical heterogeneity, and benign course. [Published with video sequences].

Published
2013

18. Genome-wide association analysis of genetic generalized epilepsies implicates susceptibility loci at 1q43, 2p16.1,2q22.3 and 17q21.32

Author

Steffens, M., Leu, C., Ruppert, A., Zara, F., Striano, P., Robbiano, A., Capovilla, G., Tinuper, P., Gambardella, A., Bianchi, A., La neve, A., Crichiutti, G., de kovel, C. G., Trenité, D. K. -N., de haan, G., Lindhout, D., Gaus, V., Schmitz, B., Janz, D., Weber, Y. G., Becker, F., Lerche, H., Steinhoff, B. J., Kleefuß-Lie, A. A., Kunz, W. S., Surges, R., Elger, C. E., Muhle, H., Von spiczak, S., Ostertag, P., Helbig, I., Stephani, U., Møller, R. S., Hjalgrim, H., Dibbens, L. M., Bellows, S., Oliver, K., Mullen, S., Scheffer, I. E., Berkovic, S. F., Everett, K. V., Gardiner, M. R., Marini, Chiara, Guerrini, R., Lehesjoki, A., Siren, A., Guipponi, M., Malafosse, A., Thomas, P., Nabbout, R., Baulac, S., Leguern, E., Guerrero, R., Serratosa, J. M., Reif, P. S., Rosenow, F., Mörzinger, M., Feucht, M., Zimprich, F., Kapser, C., Schankin, C. J., Suls, A., Smets, K., De jonghe, P., Jordanova, A., Caglayan, H., Yapici, Z., Yalcin, D. A., Baykan, B., Bebek, N., Ozbek, U., Gieger, C., Wichmann, H., Balschun, T., Ellinghaus, D., Franke, A., Meesters, C., Becker, T., Wienker, T. F., Hempelmann, A., Schulz, H., Rüschendorf, F., Leber, M., Pauck, S. M., Trucks, H., Toliat, M. R., Nürnberg, P., Avanzini, G., Koeleman, B. P., Sander, T., Weckhuysen, S., Claes, L., Deprez, L., Van Dyck, T., Deconinck, T., De Jonghe, P., Velizarova, R., Dimova, P., Radionova, M., Tournev, I., Kancheva, D., Kaneva, R., Lehesjoki, A. -E., von Spiczak, S., Martin Klein, K., Oertel, W. H., Hamer, H. M., Marini, C., Mei, D., Norci, V., Pezzella, M., La Neve, A., Vigliano, P., Vanadia, F., Vignoli, A., Coppola, A., Striano, S., Egeo, G., Teresa Giallonardo, M., Franceschetti, S., Belcastro, V., Benna, P., Coppola, G., De Palo, A., Ferlazzo, E., Vecchi, M., Martinelli, V., Bisulli, F., Beccaria, F., Del Giudice, E., Mancardi, M., Stranci, G., Scabar, A., Gobbi, G., Giordano, I., de Haan, G. -J., Giraldez, B. G., Ozbeck, U., Ozdemir, O., Ugur, S., Kocasoy-Orhan, E., Yücesan, E., Cine, N., Gokyigit, A., Gurses, C., Gul, G., Ozkara, C., Yalcin, O., Turkdogan, D., Dizdarer, G., Agan, K., Steffens, Michael, Leu, Costin, Ruppert, Ann-Kathrin, Zara, Frederico, Dibbens, Leanne Michelle, Sander, Thomas, EPICURE Consortium, Epicure, Consortium, DEL GIUDICE, Ennio, Steffens, M, Leu, C, Ruppert, Ak, Zara, F, Striano, P, Robbiano, A., Coppola, Antonietta, E. P. I. C. U. R. E. Consortium, E. M. I.Net Consortium, M. Steffen, C. Leu, A. Ruppert, F. Zara, P. Striano, A. Robbiano, G. Capovilla, P. Tinuper, A. Gambardella, A. Bianchi, A. L. Neve, G. Crichiutti, C. G. F, D. K. Trenité, G. d. Haan, D. Lindhout, V. Gau, B. Schmitz, D. Janz, Y. G. Weber, F. Becker, H. Lerche, B. J. Steinhoff, A. A. Kleefuß-Lie, W. S. Kunz, R. Surge, C. E. Elger, H. Muhle, S. v. Spiczak, P. Ostertag, I. Helbig, U. Stephani, R. S. Møller, H. Hjalgrim, L. M. Dibben, S. Bellow, K. Oliver, S. Mullen, I. E. Scheffer, S. F. Berkovic, K. V. Everett, M. R. Gardiner, C. Marini, R. Guerrini, A. Lehesjoki, A. Siren, M. Guipponi, A. Malafosse, P. Thoma, R. Nabbout, S. Baulac, E. Leguern, R. Guerrero, J. M. Serratosa, P. S. Reif, F. Rosenow, M. Mörzinger, M. Feucht, F. Zimprich, C. Kapser, C. J. Schankin, A. Sul, K. Smet, P. D. Jonghe, A. Jordanova, H. Caglayan, Z. Yapici, D. A. Yalcin, B. Baykan, N. Bebek, U. Ozbek, C. Gieger, H. Wichmann, T. Balschun, D. Ellinghau, A. Franke, C. Meester, T. Becker, T. F. Wienker, A. Hempelmann, H. Schulz, F. Rüschendorf, M. Leber, S. M. Pauck, H. Truck, M. R. Toliat, P. Nürnberg, G. Avanzini, B. P. C, and T. Sander

Subjects
Candidate gene, Juvenile, Genome-wide association study, Alleles, Epilepsy, ZEB2 protein, human, VRK2 protein, human, 0302 clinical medicine, genetics [Genetic Predisposition to Disease], genetics, Humans, Myoclonic Epilepsy, genetics [Epilepsy, Generalized], SCN1A protein, human, Genetics (clinical), Genetics, 0303 health sciences, genetics [Epilepsy, Absence], Myoclonic Epilepsy, Juvenile, genetics, Genetic Predisposition to Disease, General Medicine, Protein-Serine-Threonine Kinases, 3. Good health, Chemistry, Absence, genetics, Epilepsy, genetics [Myoclonic Epilepsy, Juvenile], Epilepsy, Generalized, genetics [Receptor, Muscarinic M3], genetics, NAV1.1 Voltage-Gated Sodium Channel, genetics [Homeodomain Proteins], Single-nucleotide polymorphism, genetics [NAV1.1 Voltage-Gated Sodium Channel], Protein Serine-Threonine Kinases, Biology, genetics [Protein-Serine-Threonine Kinases], 03 medical and health sciences, ddc:570, Genetic variation, medicine, Humans, Genetic Predisposition to Disease, genetics, Repressor Protein, Allele, Molecular Biology, Alleles, Zinc Finger E-box Binding Homeobox 2, 030304 developmental biology, Homeodomain Proteins, Receptor, Muscarinic M3, genetics, Protein-Serine-Threonine Kinase, genetics, Receptor, Generalized, genetics, Genome-Wide Association Study, Homeodomain Protein, Heritability, medicine.disease, NAV1.1 Voltage-Gated Sodium Channel, Repressor Proteins, genetics [Repressor Proteins], Muscarinic M3, Epilepsy, Absence, Myoclonic epilepsy, Human medicine, Juvenile myoclonic epilepsy, 030217 neurology & neurosurgery, Genome-Wide Association Study
Abstract

Genetic generalized epilepsies (GGEs) have a lifetime prevalence of 0.3\% and account for 20-30\% of all epilepsies. Despite their high heritability of 80\%, the genetic factors predisposing to GGEs remain elusive. To identify susceptibility variants shared across common GGE syndromes, we carried out a two-stage genome-wide association study (GWAS) including 3020 patients with GGEs and 3954 controls of European ancestry. To dissect out syndrome-related variants, we also explored two distinct GGE subgroups comprising 1434 patients with genetic absence epilepsies (GAEs) and 1134 patients with juvenile myoclonic epilepsy (JME). Joint Stage-1 and 2 analyses revealed genome-wide significant associations for GGEs at 2p16.1 (rs13026414, P(meta) = 2.5 × 10(-9), OR[T] = 0.81) and 17q21.32 (rs72823592, P(meta) = 9.3 × 10(-9), OR[A] = 0.77). The search for syndrome-related susceptibility alleles identified significant associations for GAEs at 2q22.3 (rs10496964, P(meta) = 9.1 × 10(-9), OR[T] = 0.68) and at 1q43 for JME (rs12059546, P(meta) = 4.1 × 10(-8), OR[G] = 1.42). Suggestive evidence for an association with GGEs was found in the region 2q24.3 (rs11890028, P(meta) = 4.0 × 10(-6)) nearby the SCN1A gene, which is currently the gene with the largest number of known epilepsy-related mutations. The associated regions harbor high-ranking candidate genes: CHRM3 at 1q43, VRK2 at 2p16.1, ZEB2 at 2q22.3, SCN1A at 2q24.3 and PNPO at 17q21.32. Further replication efforts are necessary to elucidate whether these positional candidate genes contribute to the heritability of the common GGE syndromes.

Published
2012
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19. Molecular Mechanisms Generating and Stabilizing Terminal 22q13 Deletions in 44 Subjects with Phelan/McDermid Syndrome

Author

Roberto Giorda, Susan Marelli, Santina Reitano, Alfredo Brusco, Renzo Guerrini, Maria Grazia Patricelli, Nieves Martínez Guardia, María Luisa Martínez-Frías, Maria Clara Bonaglia, Stefano D'Arrigo, Donatella Greco, Frédérique Béna, Antonella Cecconi, Sara Osimani, Bernardo Dalla Bernardina, Maria Luisa Martínez-Fernández, Pierluigi Politi, Cristina De Agostini, Elisa Mani, Lucia Grillo, Elisabetta Lapi, Sabrina Giglio, Stefania Bigoni, Francesca Novara, Claudio Zucca, Corrado Romano, Silvana Beri, Renato Borgatti, Rita Grasso, Roberto Ciccone, Marco Fichera, Anna Bremer, Emanuela Priolo, Anna Baroncini, Emanuela Maserati, Irene Bagnasco, Stefania Gimelli, Lucio Nitsch, Cristina Motta, Britt-Marie Anderlid, Ornella Galesi, Annalisa Vetro, Orsetta Zuffardi, Francesca L. Sciacca, Chiara Pantaleoni, Giulia Arrigo, Leonardo Zoccante, Ursula Giussani, Daria Riva, Maria Teresa Bonati, Francesca Darra, Massimo Molteni, Eleonora Di Gregorio, Cristiano Termine, Armand Bottani, M. C., Bonaglia, R., Giorda, S., Beri, C. D., Agostini, F., Novara, M., Fichera, L., Grillo, O., Galesi, A., Vetro, R., Ciccone, M. T., Bonati, S., Giglio, R., Guerrini, S., Osimani, S., Marelli, C., Zucca, R., Grasso, R., Borgatti, E., Mani, C., Motta, M., Molteni, C., Romano, D., Greco, S., Reitano, A., Baroncini, E., Lapi, A., Cecconi, G., Arrigo, M. G., Patricelli, C., Pantaleoni, S., D'Arrigo, D., Riva, F., Sciacca, B. D., Bernardina, L., Zoccante, F., Darra, C., Termine, E., Maserati, S., Bigoni, E., Priolo, A., Bottani, S., Gimelli, F., Bena, A., Brusco, E. d., Gregorio, I., Bagnasco, U., Giussani, Nitsch, Lucio, P., Politi, M. L., Martinez Fria, M. L., Martinez Fernandez, N. M., Guardia, A., Bremer, B., Anderlid, and O., Zuffardi

Subjects
Male, Parents, Cancer Research, Chromosomes, Human, Pair 22, Ring chromosome, 22q13 deletion syndrome, Chromosome Disorders, Germline, Translocation, Genetic, Chromosomal Disorders, Phelan/McDermid Syndrome, Ring Chromosomes, Child, SHANK3, Genetics (clinical), Sequence Deletion, Genetics, Aged, 80 and over, Comparative Genomic Hybridization, deletion 22q13, Chromosomal Deletions and Duplications, Middle Aged, Phelan McDermid, Child, Preschool, Cytogenetic Analysis, Translocations, Medicine, Female, Chromosome Deletion, Haploinsufficiency, Research Article, Adult, Monosomy, SCAFFOLDING PROTEIN SHANK3, CHROMOSOME BREAKAGE, SUBTELOMERIC PROBES, ALZHEIMERS-DISEASE, MENTAL-RETARDATION, ALPHA-THALASSEMIA, TELOMERE CAPTURE, MONOSOMY 1P36, ARRAY CGH, GENE, lcsh:QH426-470, Adolescent, Molecular Sequence Data, Biology, Cytogenetics, Young Adult, medicine, Humans, Molecular Biology, Ecology, Evolution, Behavior and Systematics, Aged, Clinical Genetics, Base Sequence, Breakpoint, Infant, Newborn, Infant, Human Genetics, medicine.disease, Telomere, lcsh:Genetics, microdeletion, array-CGH, Chromosome 22
Abstract

In this study, we used deletions at 22q13, which represent a substantial source of human pathology (Phelan/McDermid syndrome), as a model for investigating the molecular mechanisms of terminal deletions that are currently poorly understood. We characterized at the molecular level the genomic rearrangement in 44 unrelated patients with 22q13 monosomy resulting from simple terminal deletions (72%), ring chromosomes (14%), and unbalanced translocations (7%). We also discovered interstitial deletions between 17–74 kb in 9% of the patients. Haploinsufficiency of the SHANK3 gene, confirmed in all rearrangements, is very likely the cause of the major neurological features associated with PMS. SHANK3 mutations can also result in language and/or social interaction disabilities. We determined the breakpoint junctions in 29 cases, providing a realistic snapshot of the variety of mechanisms driving non-recurrent deletion and repair at chromosome ends. De novo telomere synthesis and telomere capture are used to repair terminal deletions; non-homologous end-joining or microhomology-mediated break-induced replication is probably involved in ring 22 formation and translocations; non-homologous end-joining and fork stalling and template switching prevail in cases with interstitial 22q13.3. For the first time, we also demonstrated that distinct stabilizing events of the same terminal deletion can occur in different early embryonic cells, proving that terminal deletions can be repaired by multistep healing events and supporting the recent hypothesis that rare pathogenic germline rearrangements may have mitotic origin. Finally, the progressive clinical deterioration observed throughout the longitudinal medical history of three subjects over forty years supports the hypothesis of a role for SHANK3 haploinsufficiency in neurological deterioration, in addition to its involvement in the neurobehavioral phenotype of PMS., Author Summary Terminal chromosome deletions are among the most commonly observed rearrangements detected by cytogenetics and may result in several well-known genetic syndromes. We used 22q13 deletions to study how these types of chromosome abnormalities arise. Children with Phelan/McDermid syndrome, caused by deletion of the terminal portion of chromosome 22, experience developmental delay, absent or severely delayed speech, and frequent behavioral problems. Lack of one copy of SHANK3, a key gene for the correct development and organization of brain synapses, is very likely the basis of the syndrome's major neurological features. Deletion of additional genes probably causes more complex phenotypes in subjects with larger deletions. We also studied patients who only lack a portion of SHANK3 and demonstrated that small, hard-to-detect deletions of this gene may cause substantial clinical problems. Until now, the 22q distal deletion had been only diagnosed in very young people. We studied a large group of patients of different ages and discovered that all adult patients face progressive cognitive decline. Our study demonstrates that deletion of the terminal portion of chromosome 22, a prototype for terminal deletions in human chromosomes, can occur in several ways. Mosaic deletions of different size can also form in early embryogenesis.

Published
2011

20. Mutational analysis of EFHC1 gene in Italian families with juvenile myoclonic epilepsy

Author

Annesi, F., Gambardella, A., Michelucci, R., Bianchi, A., Marini, C., Canevini, M. P., Capovilla, G., Elia, M., Buti, D., Chifari, R., Striano, Pasquale, Rocca, F. E., Castellotti, B., Cali, F., Labate, A., Lepiane, E., Besana, D., Sofia, V., Tabiadon, G., Tortorella, G., Vigliano, P., Vignoli, A., Beccaria, F., Annesi, G., Striano, S., Aguglia, U., Guerrini, R., Quattrone, A., Annesi, F, Gambardella, A, Michelucci, R, Bianchi, A, Marini, C, Canevini, Mp, Capovilla, G, Elia, M, Buti, D, Chifari, R, Striano, P, Rocca, Fe, Castellotti, B, Cali, F, Labate, A, Lepiane, E, Besana, D, Sofia, V, Tabiadon, G, Tortorella, G, Vigliano, P, Vignoli, A, Beccaria, F, Annesi, G, Striano, Salvatore, Aguglia, U, Guerrini, R, Quattrone, A., F., Annesi, A., Gambardella, R., Michelucci, A., Bianchi, C., Marini, M. P., Canevini, G., Capovilla, M., Elia, D., Buti, R., Chifari, P., Striano, F. E., Rocca, B., Castellotti, F., Cali, A., Labate, E., Lepiane, D., Besana, V., Sofia, G., Tabiadon, G., Tortorella, P., Vigliano, A., Vignoli, F., Beccaria, G., Annesi, U., Aguglia, R., Guerrini, and A., Quattrone

Subjects
Adult, Male, Adult, Calcium-Binding Protein, European Continental Ancestry Group, DNA Mutational Analysis, Mutation, Missense, Juvenile, White People, genetics, Family, Female, Genetic Heterogeneity, Genetic Testing, Humans, Italy, Adult, Calcium-Binding Proteins, genetics, Chromosome Mapping, DNA Mutational Analysis, European Continental Ancestry Group, epidemiology, Male, Middle Aged, Mutation, Missense, genetics, Mutation, genetics, Myoclonic Epilepsy, epidemiology/ethnology/genetics, Pedigree, Phenotype, Genetic Heterogeneity, Humans, genetics, Family, Genetic Testing, epidemiology/ethnology/genetics, Calcium-Binding Proteins, Myoclonic Epilepsy, Juvenile, Chromosome Mapping, Middle Aged, Pedigree, Phenotype, Myoclonic Epilepsy, Italy, Mutation, epidemiology, Female
Abstract

Mutations in the EFHC1 gene have been reported in six juvenile myoclonic epilepsy (JME) families from Mexico and Belize. In this study, we screened 27 unrelated JME Italian families for mutations in the EFHC1 gene.Twenty-seven families (86 affected individuals, 52 women) with at least two affected members with JME were selected. DNA was isolated from peripheral blood lymphocytes by standard methods and each exon of the EFHC1 gene was amplified and sequenced using intronic primers.Two heterozygous mutations were identified in three unrelated families. One (R353 W) was a novel missense mutation, while the F229 L mutation was previously described (say which on of the two occurred in two families). Both mutations cosegregated with the disease. In a fourth family, the variant 545G-->A (resulting in the amino acid substitution R182 H) cosegregated with JME.The results of our study extend the distribution of EFHC1 mutations to the white population and confirm the high level of genetic heterogeneity associated with JME.

Published
2007

21. Familial occurrence of febrile seizures and epilepsy in severe myoclonic epilepsy of infancy (SMEI) patients with SCN1A mutations

Author

Elena Freri, Pasquale Striano, Francesca Madia, Antonia Parmeggiani, Maria Margherita Mancardi, Francesca Darra, Chiara Pantaleoni, Sara Scapolan, Giuseppe Gobbi, Pierangelo Veggiotti, Maurizio Elia, Elena Gennaro, Federico Zara, Bernardo Dalla Bernardina, Antonino Romeo, Renzo Guerrini, Marilena Vecchi, Federico Vigevano, Angela Pistorio, Roberta Paravidino, Margherita Santucci, Roberto Gaggero, Amedeo Bianchi, Tiziana Granata, Enrico Bertini, Giuseppe Capovilla, M. M. Mancardi, P. Striano, E. Gennaro, F. Madia, R. Paravidino, S. Scapolan, B. dalla Bernardina, E. Bertini, A. Bianchi, G. Capovilla, F. Darra, M. Elia, E. Freri, G. Gobbi, T. Granata, R. Guerrini, C. Pantaleoni, A. Parmeggiani, A. Romeo, M. Santucci, M. Vecchi, P. Veggiotti, F. Vigevano, A. Pistorio, R. Gaggero, and F. Zara

Subjects
Proband, Pediatrics, medicine.medical_specialty, Concordance, Epilepsies, Myoclonic, Nerve Tissue Proteins, Neurological disorder, Epilepsies, Seizures, Febrile, Voltage-gated sodium channel α subunit type A, Sodium Channels, Central nervous system disease, Febrile, Epilepsy, Myoclonic, epidemiology/genetics, Epilepsy, epidemiology/genetics, Family, Humans, Mutation, genetics, Nerve Tissue Proteins, genetics, Pedigree, Seizures, epidemiology/genetics, Sodium Channels, genetics, Seizures, Voltage-gated sodium channel αsubunit type A, Genetics, medicine, Severe myoclonic epilepsy of infancy, Humans, Family, Family history, epidemiology/genetics, business.industry, Matched control, medicine.disease, Pedigree, NAV1.1 Voltage-Gated Sodium Channel, Neurology, Anesthesia, Mutation, Myoclonic epilepsy, Neurology (clinical), business
Abstract

Summary: Purpose: The role of the familial background in severe myoclonic epilepsy of infancy (SMEI) has been traditionally emphasized in literature, with 25–70% of the patients having a family history of febrile seizures (FS) or epilepsy. We explored the genetic background of SMEI patients carrying SCN1A mutations to further shed light on the genetics of this disorder. Methods: We analyzed the occurrence of FS and epilepsy among first- and second-degree relatives (N = 867) of 74 SMEI probands with SCN1A mutations (70 de novo, four inherited) and compared data with age-matched and ethnically matched control families. Familial clustering and syndromic concordance within the affected relatives in both groups were investigated. Results: The frequency of FS or epilepsy in relatives of SMEI patients did not significantly differ from that in controls (FS: 13 of 867 vs. 12 of 674, p = 0.66; epilepsy: 15 of 867 vs. six of 674, p = 0.16). Different forms of epilepsy were identified in both relatives of SMEI probands and controls. Twenty-eight relatives with FS and epilepsy were distributed in 20 (27%) of 74 SMEI families; among the controls, 18 affected relatives were clustered in 13 (18.5%) of 70 families. No pedigree showed several affected members, including the four with inherited mutations. Conclusions: A substantial epileptic family background is not present in our SMEI patients with SCN1A mutations. These data do not confirm previous observations and would not support polygenic inheritance in SMEI. The investigation of the family background in additional series of SMEI patients will further shed light on the genetics of this syndrome.

Published
2006

22. Solution structure of amyloid beta-peptide (25-35) in different media

Author

Anna Maria D'Ursi, Severo Salvadori, Remo Guerrini, Delia Picone, Maria Rosaria Armenante, Giuseppe Sorrentino, A. M., D'Ursi, M. R., Armenante, R., Guerrini, S., Salvadori, G., Sorrentino, and Picone, Delia

Subjects
chemistry.chemical_classification, Models, Molecular, Circular dichroism, Amyloid beta-Peptides, Magnetic Resonance Spectroscopy, biology, Amyloid, Stereochemistry, Amyloid beta, Chemistry, Protein Conformation, Circular Dichroism, Sequence (biology), Peptide, Nuclear magnetic resonance spectroscopy, Peptide Fragments, NO, Solutions, Transmembrane domain, Protein structure, mental disorders, Drug Discovery, biology.protein, Molecular Medicine, Electrophoresis, Polyacrylamide Gel
Abstract

The design of molecules able to interact with the amyloid peptides either as inhibitors of fibril formation or as inhibitors of amyloid membrane pore formation represents one of the most relevant approaches in the development of anti-Alzheimer therapies. Abeta-(25-35), sequence GSNKGAIIGLM, is a highly toxic synthetic derivative of amyloid beta-peptides (Abeta-peptides), which forms fibrillary aggregates. Here, we report the NMR and CD investigation of Abeta-(25-35) in a membrane-mimicking environment and in isotropic mixtures of water and fluoro-alcohols to scan its conformational properties as a function of the medium. The analysis of the 3D structures in the mentioned conditions indicates a propensity of the peptide to behave as a typical transmembrane helix in the lipidic environment. In media characterized by different polarity, it loses the structural regularity at specific points of the sequence as a function of the environment. Furthermore, a comparison with the solution structure of full-length amyloid peptides suggests a role for the 25-27 kink region, which appears to be a general feature of all peptides under the solution conditions explored.

Published
2004

23. A new benign adult familial myoclonic epilepsy (BAFME) pedigree suggesting linkage to chromosome 2p11.1-q12.2

Author

Maria Paola Canevini, Maurizio Elia, Maurizio De Fusco, Salvatore Striano, Rosanna Chifari, Renzo Guerrini, Pasquale Striano, Giorgio Casari, Striano, P, Chifari, R, Striano, Salvatore, DE FUSCO, M, Elia, M, Guerrini, R, Casari, G, Canevini, Mp, P., Striano, R., Chifari, M. d., Fusco, M., Elia, R., Guerrini, G., Casari, M. P., Canevini, Striano, S, de Fusco, M, and Casari, GIORGIO NEVIO

Subjects
Adult, Male, Benign adult familial myoclonic epilepsy, classification/genetics, Genetic Linkage, Adult, Aged, Chromosomes, Human, Pair 2, Epilepsies, Myoclonic, classification/genetics, Family Health, Female, Genetic Linkage, Humans, Male, Pedigree, Phenotype, Epilepsies, Myoclonic, Epilepsies, Biology, Chromosomes, Epilepsy, Genetic linkage, medicine, Humans, Aged, Genetics, Linkage (software), Family Health, Pair 2, Epilepsie, Genetic heterogeneity, Chromosome, Adult, Aged, Chromosome, medicine.disease, Phenotype, Pedigree, Neurology, Chromosomes, Human, Pair 2, Pair 2, Myoclonic epilepsy, Female, Neurology (clinical)
Abstract

Summary: Benign adult familial myoclonic epilepsy (BAFME) is an autosomal dominant condition characterized by cortical tremor and generalized seizures, mapped on chromosome 8q24 by Japanese authors. Recently the same phenotype also was reported in European families, with linkage on chromosome 2. We present a new family with suggestion of linkage to chromosome 2p11.1-2q12.2 (lod score value, 1.55). This observation would confirm that BAFME is a worldwide, genetically heterogeneous condition, probably with Japanese families linked to 8q24 and European families to 2p11.1-q12.2.

Published
2004

24. Solution conformation of nociceptin

Author

Delia Picone, Teodorico Tancredi, S. Salvadori, Domenico Regoli, Pierandrea Temussi, Lawrence H. Lazarus, Remo Guerrini, Roberta Spadaccini, S., Salvadori, Picone, Delia, T., Tancredi, R., Guerrini, R., Spadaccini, L. H., Lazaru, D., Regoli, and P. A., Temussi

Subjects
Models, Molecular, Magnetic Resonance Spectroscopy, medicine.drug_class, Protein Conformation, Guinea Pigs, Molecular Sequence Data, Biophysics, Dynorphin, Pharmacology, In Vitro Techniques, Biochemistry, Dynorphins, Protein Structure, Secondary, Opioid receptor, Ileum, Cerebellum, polycyclic compounds, medicine, Animals, Amino Acid Sequence, Receptor, Molecular Biology, Endogenous opioid, Sequence (medicine), Chemistry, Cell Biology, Rats, Solutions, Nociceptin receptor, Kinetics, nervous system, Opioid, Opioid Peptides, Receptors, Opioid, Thermodynamics, medicine.drug
Abstract

Nociceptin, a novel heptadecapeptide, interacts with ORL1 a G protein-coupled receptor whose sequence is closely related to that of the kappa opioid receptor but has no opioid activity. We have Investigated the conformational preferences of Nociceptin also in comparison to Dynorphin A, The N-terminal part of Nociceptin has the same conformational preferences of She message of endogenous opioids but the C-terminal part of the sequence is more flexible than the corresponding address of Dynorphin A. [Tyr(1)]-Nociceptin, while retaining nociceptive activity, has also an opioid activity comparable to that of enkephalins. (C) 1997 Academic Press.

Published
1997

25. Selective Opioid Dipeptides

Author

Remo Guerrini, Delia Picone, S. Salvadori, Teodorico Tancredi, Pietro Amodeo, L. H. Lazarus, Roberto Tomatis, Piero Andrea Temussi, Clementina Bianchi, P. A., Temussi, S., Salvadori, P., Amodeo, C., Bianchi, R., Guerrini, R., Tomati, L. H., Lazaru, Picone, Delia, and T., Tancredi

Subjects
Models, Molecular, Protein Conformation, Receptors, Opioid, mu, Biochemistry, Radioligand Assay, Receptors, Opioid, delta, Tetrahydroisoquinolines, chemistry.chemical_classification, Brain, Stereoisomerism, Aromaticity, Dipeptides, Enkephalins, Selectivity, Oligopeptides, Muscle Contraction, medicine.drug, Narcotics, congenital, hereditary, and neonatal diseases and abnormalities, Stereochemistry, Carboxylic acid, Guinea Pigs, Molecular Sequence Data, Biophysics, In Vitro Techniques, Binding, Competitive, Structure-Activity Relationship, Ileum, mental disorders, medicine, Animals, Amino Acid Sequence, Opioid peptide, Molecular Biology, Cell Membrane, Muscle, Smooth, Cell Biology, Enkephalin, Ala(2)-MePhe(4)-Gly(5), Isoquinolines, Combinatorial chemistry, In vitro, Rats, nervous system diseases, body regions, Kinetics, Opioid, chemistry, Enkephalin, D-Penicillamine (2,5), Chirality (chemistry), human activities, Synaptosomes
Abstract

The surprising change of selectivity induced by the change of chirality in peptides containing the tetrahydro-3-isoquinoline carboxylic acid (Tic) in second position, interpreted as a conformational preference induced on the Tyr-Xaa-Phe domain, can instead be attributed to the Tyr-Tic message domain. The relative spatial disposition of the aromatic rings of δ-selective non peptidic opiates is compatible with a message domain, in opioid peptides, of only two residues. This hypothesis was tested through the synthesis of Tyr-L-Tic-NH 2 , Tyr-D-Tic-NH 2 , Tyr-L-Tic-Ala-NH 2 , Tyr-L-Tic-Ala-OH and Tyr-D-Tic-Ala-NH 2 . Peptides containing Tyr-L-Tic- behave as very selective δ antagonists and those containing Tyr-DTic- as non selective agonists. This is the first case of opioid peptides containing a two-residue message domain and of opioid dipeptides with substantial opioid activity.

Published
1994

26. Conversion of Enkephalin and Dermorphin into δ‐Selective Opioid Antagonists by Single‐Residue Substitution

Author

Piero Andrea Temussi, Delia Picone, Sharon D. Bryant, Giuliano Marzola, Severo Salvadori, Teodorico Tancredi, Pietro Amodeo, Lawrence H. Lazarus, Remo Guerrini, T., Tancredi, S., Salvadori, P., Amodeo, Picone, Delia, L. H., Lazaru, S. D., Bryant, R., Guerrini, G., Marzola, and P. A., Temussi

Subjects
Agonist, Models, Molecular, Magnetic Resonance Spectroscopy, Enkephalin, medicine.drug_class, Stereochemistry, Protein Conformation, Carboxylic acid, Narcotic Antagonists, Guinea Pigs, Molecular Sequence Data, Peptide, In Vitro Techniques, Biochemistry, chemistry.chemical_compound, Mice, Radioligand Assay, Naltrindole, Tetrahydroisoquinolines, medicine, Animals, Amino Acid Sequence, Opioid peptide, chemistry.chemical_classification, Chemistry, Brain, Biological activity, Dermorphin, Enkephalins, Isoquinolines, Rats, Opioid Peptides, Oligopeptides, medicine.drug
Abstract

The properties of di- and tri-peptides containing 1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid (Tic) in second position suggest that the message domain of opioid peptides can be composed of only two residues [Temussi, P. A., Salvadori, S., Amodeo, P., Guerrini, R., Tomatis, R., Lazarus, L. H., Picone, D. gr Tancredi, T. (1994) Biochem. Biophys. Res. Commun. 198, 933-939]. As a crucial test of the possibility that the Sr-Tic segment be a message domain in longer peptide sequences, we have inserted it in the sequences of two typical opioid peptides: [Leu]enkephalin, a non-selective agonist, and dermorphin, a selective mu agonist. Here we report the synthesis and biological activity of [L-Tic(2)]enkephalin, [L-Tic(2)]dermorphin, [L-Tic(2)]dermorphin carboxylic acid and [D-Tic(2)]dermorphin: all [L-Tic(2)]peptides were converted from agonists to delta-selective antagonists. The NMR conformational study in a dimethylsulfoxide/water cryoprotective mixture at low temperature shows diagnostic side-chain - side-chain NOEs in the spectra of all [L-Tic(2)]peptides and hints that the 90 degrees arrangement of the the two aromatic rings found in the cis-Tyr-L-Tic moiety, typical of N-methyl naltrindole and other delta-selective opiate antagonists, is responsible for the antagonist activity of all these peptides.

Published
1994

27. Inclusion of pyridoxine dependent epilepsy in expanded newborn screening programs by tandem mass spectrometry: set up of first and second tier tests.

Author

Damiano R, Della Bona M, Procopio E, Guerrini R, Bettiol A, and la Marca G

Abstract

Objectives: Pyridoxine-dependent epilepsy (PDE) is a rare genetic disorder characterized by intractable neonatal seizures responsive to pyridoxine. Diagnosis relies on quantification of α-aminoadipic semialdehyde, piperideine-6-carboxylate and pipecolic acid in urine or plasma in patients with overt symptoms. We developed and validated simple and rapid first- and second-tier methods for two recently published biomarkers of PDE (2S,6S-/2S,6R-oxopropylpiperidine-2-carboxylic acid (2-OPP) and 6-oxopiperidine-2-carboxylic acid (6-oxoPIP)) in extended newborn screening (NBS) programs from neonatal dried blood spots (DBS)., Methods: For the first-line test, DBS specimens were collected from 5,405 newborns who underwent routine NBS and analysed by FIA-MS/MS. For the second-tier test, samples were analysed by LC-MS/MS. The neonatal DBS from two patients with genetically confirmed PDE were also analysed., Results: The reference values for NBS resulted <0.34 μmol/L for 2-OPP and <4.51 μmol/L for 6-oxoPIP. In the second-tier test, limits of detection were 0.07 μmol/L and 0.14 μmol/L, whereas limits of quantification were 0.25 μmol/L and 0.48 μmol/L, respectively, for 2-OPP and for 6-oxoPIP. The tests provided good linearity, reproducibility, accuracy and precision, with acceptable matrix effect and carry-over, according to international validation criteria. The biomarkers in DBS were stable at room temperature, +4 °C and -20 °C for one month. When assessing these biomarkers in two patients with genetically confirmed PDE, the higher sensitivity of 2-OPP as compared to 6-oxoPIP in discriminating PDE emerged., Conclusions: The first-line and second-tier tests developed in this study highlight the potential for including PDE in the NBS panel, early diagnosis and prompt precision treatment initiation., (© 2025 Walter de Gruyter GmbH, Berlin/Boston.)

Published
2025
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28. Psychiatric disorders in children and adolescents with temporal lobe epilepsy: A narrative review.

Author

Cappelletto P, Accolla C, Preti M, Pisano T, Barba C, and Guerrini R

Subjects
Humans, Adolescent, Child, Risk Factors, Prevalence, Epilepsy, Temporal Lobe psychology, Mental Disorders epidemiology
Abstract

People with epilepsy (PWE) are at higher risk of psychiatric disorders (PD), disability, and reduced quality of life than the general population, especially in childhood and adolescence and when seizures originate from the temporal lobe. Temporal Lobe Epilepsy (TLE) is the most common type of focal epilepsy and can be due to structural abnormalities, or non-lesional causes, such as genetic variants. The prevalence of PD is approximately 20%-30% in people with epilepsy in general, and from 40% up to 80% in people with TLE. A higher rate of anxiety and depression disorders has been observed in association with TLE than with extra-temporal epilepsy, or idiopathic generalized epilepsy, or other chronic diseases such as diabetes. However, while the association between psychopathology and TLE has been extensively assessed in adults, only a few studies have focused on its expression in children and adolescents. In this review, we describe the prevalence, characteristics, and risk factors for PD in people with epilepsy in general and with TLE, with a specific focus on the pediatric age. In addition, we provide insights into the current knowledge of the pathophysiological bases of psychiatric symptoms in children and adolescents with TLE. PLAIN LANGUAGE SUMMARY: This review examines the frequency and characteristics of psychiatric disorders in people with temporal lobe epilepsy, with a focus on children and adolescents. Similarly to adults, younger people with epilepsy have higher rates of psychiatric disorders, such as depression and anxiety, than healthy peers or children with other chronic illnesses such as diabetes and asthma. Contributing risk factors include epilepsy duration and severity, and the effects of antiseizure medications, as well as psychological challenges, sociocultural influences, and family dynamics. Psychiatric disorders associated with temporal lobe epilepsy are relatively frequent, probably in relation to the critical role that some limbic structures in the temporal lobe, such as the amygdala and hippocampus, play in regulating emotions and behavior., (© 2024 The Author(s). Epilepsia Open published by Wiley Periodicals LLC on behalf of International League Against Epilepsy.)

Published
2025
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29. Determination of new biomarkers for diagnosis of pyridoxine dependent epilepsy in human plasma and urine by liquid chromatography-mass spectrometry.

Author

Damiano R, Della Bona M, Procopio E, Gasperini S, Guerrini R, Bettiol A, and la Marca G

Subjects
Humans, Chromatography, Liquid, Liquid Chromatography-Mass Spectrometry, Biomarkers urine, Biomarkers blood, Epilepsy diagnosis, Epilepsy urine, Epilepsy blood, Tandem Mass Spectrometry
Abstract

Background: Pyridoxine-dependent epilepsy (PDE) is a rare inborn error of lysine metabolism. To date, diagnosis of PDE relies on the quantification of α-AminoAdipic SemiAldehyde (α- AASA), Piperideine-6-Carboxylate (P6C) and Pipecolic acid (PA) in urine or plasma from patients with overt symptoms. However, these biomarkers are not specific, and their biochemical analysis is challenged by their instability and technical limitations. We set-up and validated a method for the quantification of two new biomarkers of PDE (2S,6S- and 2S,6R-oxopropylpiperidine-2-carboxylic acid, 2-OPP, and 6-oxopiperidine2-carboxylic acid, 6-oxoPIP) on human urine and plasma by LC-MS/MS, to overcome the diagnostic and technical challenges of old biomarkers., Methods: We analysed urine and plasma samples by LC-MS/MS, and validated the method in both biological matrices., Results: We performed the biomarkers extraction from a 10 µL aliquot of urine or plasma in around 15 min using water 100 % for urine, and a solution of water/methanol 50 % for plasma, respectively. The analytical method was validated and gave good linearity (r 2  > 0.999) in the range 0-15 µmol/L for 2-OPP and 0-25 µmol/L for 6-oxoPIP. In both matrices, the biomarkers were stable at different storage temperatures tested., Conclusions: We set-up and validated a reliable method and confirmed its clinical applicability on real samples from PDE patients. This method could be used as routine test for the diagnosis and monitoring of PDE., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Published
2025
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30. Home-based, computer-assisted cognitive rehabilitation for attention in pediatric onset multiple sclerosis: a randomized, multicenter pilot study.

Author

Masciulli C, Portaccio E, Goretti B, Niccolai C, Simone M, Viterbo RG, Zaffaroni M, Pippolo L, Cocco E, Fenu G, Carta E, Falautano M, Celico C, Pardini M, Mancardi GL, Guerrini R, Melani F, Giovannelli F, Rocca M, Iaffaldano P, Tacchino A, Zaratin P, Filippi M, and Amato MP

Subjects
Humans, Male, Female, Pilot Projects, Double-Blind Method, Adolescent, Child, Attention physiology, Treatment Outcome, Cognitive Dysfunction rehabilitation, Cognitive Dysfunction etiology, Neuropsychological Tests, Multiple Sclerosis rehabilitation, Multiple Sclerosis complications, Multiple Sclerosis psychology, Multiple Sclerosis, Relapsing-Remitting rehabilitation, Cognitive Behavioral Therapy methods, Cognitive Remediation methods, Feasibility Studies, Cognitive Training, Therapy, Computer-Assisted methods
Abstract

Objective: Cognitive impairment affects approximately 30% of pediatric onset Multiple Sclerosis (POMS) patients with a negative impact on everyday life. The aim of this study was to evaluate the feasibility and effectiveness of a home-based, computer-assisted training of attention in patients with POMS., Methods: This was a randomized double-blind study. Subjects were randomized to specific training (ST) or non-specific training (n-ST), delivered through a customized module based on attention exercises (COGNI-TRAcK2). The main feature of the ST is the implementation of working load algorithms and procedures for intensiveness regulation. The effectiveness of the ST on attention was primarily assessed on the Symbol Digit Modalities Test (SDMT), using the Wilcoxon signed-rank test. Secondary objectives included effectiveness on other cognitive tests. The evaluations were performed at baseline, end of training and 3-month follow-up., Results: 22 relapsing-remitting patients were included: data of 8 subjects in n-ST and 5 subjects in ST were available. As for the primary outcome, SDMT score improved in the ST group (from 31.2 at baseline to 42.4 after 3 months, p = 0.043). There was no benefit in other neuropsychological measures., Conclusions: These preliminary findings point to a potential benefit of a home-based, computer-assisted training of attention in patients with POMS., Competing Interests: Declarations. Ethical approval: All subjects gave their informed consent for inclusion before they participated in the study. The study was conducted in accordance with the Declaration of Helsinki, and the protocol was approved by the Ethics Committee. The protocol of the study was written in accordance to “Norma UNI EN ISO 14155 and Good Clinical Practice”. The promoter is committed to the protection of personal sensitive data in accordance with national (D.Lgs. 196/2003) and international law (European Regulation 679/2019). Conflict of interest: E.P. received compensation for travel grants, participation in advisory board and/or speaking activities from Biogen, Merck Serono, Sanofi, Teva, and Novartis and serves on the editorial board of Frontiers in Neurology and Brain Sciences. RG received fees for Advisory Boards from UCB, Zogenix, Biocodex, GW-Jazz, Angelini, Takeda, and Rapport Therapeutics, and was an investigator in the STICLO-Italy trial of stiripentol. F.M. is employees of GeneDx. M.A.R. received consulting fees from Biogen, Bristol Myers Squibb, Eli Lilly, Janssen, Roche; and speaker honoraria from AstraZaneca, Biogen, Bristol Myers Squibb, Bromatech, Celgene, Genzyme, Horizon Therapeutics Italy, Merck Serono SpA, Novartis, Roche, Sanofi and Teva. She receives research support from the MS Society of Canada, the Italian Ministry of Health, the Italian Ministry of University and Research, and Fondazione Italiana Sclerosi Multipla. She is Associate Editor for Multiple Sclerosis and Related Disorders. P.I. has served on scientific advisory boards for Biogen Idec, Bayer, Teva, Roche, Merck Serono, Novartis, and Genzyme and has received funding for travel and/or speaker honoraria from Sanofi Aventis, Genzyme, Biogen Idec, Teva, Merck Serono, and Novartis. M.F. is Editor-in-Chief of the Journal of Neurology, Associate Editor of Human Brain Mapping, Neurological Sciences, and Radiology; received compensation for consulting services from Alexion, Almirall, Biogen, Merck, Novartis, Roche, Sanofi; speaking activities from Bayer, Biogen, Celgene, Chiesi Italia SpA, Eli Lilly, Genzyme, Janssen, Merck-Serono, Neopharmed Gentili, Novartis, Novo Nordisk, Roche, Sanofi, Takeda, and TEVA; participation in Advisory Boards for Alexion, Biogen, Bristol-Myers Squibb, Merck, Novartis, Roche, Sanofi, Sanofi-Aventis, Sanofi-Genzyme, Takeda; scientific direction of educational events for Biogen, Merck, Roche, Celgene, Bristol-Myers Squibb, Lilly, Novartis, Sanofi-Genzyme; he receives research support from Biogen Idec, Merck-Serono, Novartis, Roche, the Italian Ministry of Health, the Italian Ministry of University and Research, and Fondazione Italiana Sclerosi Multipla. M.P.A. served on scientific advisory boards for and has received speaker honoraria and research support from Biogen Idec, Merck Serono, Bayer Schering Pharma, and Sanofi Aventis and serves on the editorial board of Multiple Sclerosis Journal and BMC Neurology., (© 2024. Fondazione Società Italiana di Neurologia.)

Published
2025
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31. Fibroblast transcriptomics uncovers pathogenic genomic variants in individuals with exome-negative childhood onset epilepsy.

Author

Smal N, Millevert C, De Wachter M, De Vriendt E, Eddafir Z, Schoonjans AS, Bayat A, Møller RS, Mei D, Balestrini S, Guerrini R, Meeuwissen MEC, Jansen AC, and Weckhuysen S

Abstract

Objective: This study aims to improve genetic diagnosis in childhood onset epilepsy with neurodevelopmental problems by utilizing RNA sequencing of fibroblasts to identify pathogenic variants that may be missed by exome sequencing and copy number variation analysis., Methods: We enrolled 41 individuals with childhood onset epilepsy and neurodevelopmental problems who previously had inconclusive genetic testing. Fibroblast samples were cultured and analyzed using RNA sequencing to detect aberrant expression, aberrant splicing, and monoallelic expression using the Detection of RNA Outlier Pipeline (DROP) pipeline. Detected events were correlated with phenotypes, and long-read genome sequencing was performed on individuals with strong candidate events to identify the causal genomic variant. A systematic literature review on RNA sequencing in rare disorders was conducted to contextualize our findings., Results: RNA sequencing identified five strong candidate events in four individuals, affecting the genes QRICH1, TSC1, SMARCA1, GNAI1, and PTEN. (Likely) pathogenic genomic variants affecting expression of QRICH1, TSC1, and SMARCA1 were detected, resulting in a diagnostic yield of 7% (3/41). Two variants were not covered in the initial exome sequencing data but were revealed through long-read sequencing. The identification of a pathogenic TSC1 variant led to a previously unrecognized diagnosis of tuberous sclerosis complex. This prompted guideline-based screening, which revealed tuberous sclerosis lesions in the brain and lung, directly impacting clinical care. Notably, two of the three pathogenic events would not have been detected using whole blood due to the lack of expression of the involved genes. The lower yield of this study compared to studies in other rare disorders reflects the genetic heterogeneity of epilepsy and neurodevelopmental disorders, and the inaccessibility of affected tissue., Significance: This research underscores RNA sequencing of cultured fibroblasts as a valuable tool in genetic diagnostics for childhood onset epilepsy, particularly when conventional methods fail. Expanding the control dataset with age-matched samples and incorporating RNA sequencing with nonsense-mediated decay inhibition could further enhance diagnostic yield., (© 2025 International League Against Epilepsy.)

Published
2025
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32. ASSOCIATIONS BETWEEN EPILEPSY-RELATED POLYGENIC RISK AND BRAIN MORPHOLOGY IN CHILDHOOD.

Author

Ngo A, Liu L, Larivière S, Kebets V, Fett S, Weber CF, Royer J, Yu E, Rodríguez-Cruces R, Zhang Z, Ooi LQR, Thomas Yeo BT, Frauscher B, Paquola C, Caligiuri ME, Gambardella A, Concha L, Keller SS, Cendes F, Yasuda CL, Bonilha L, Gleichgerrcht E, Focke NK, Kotikalapudi R, O'Brien TJ, Sinclair B, Vivash L, Desmond PM, Lui E, Vaudano AE, Meletti S, Kälviäinen R, Soltanian-Zadeh H, Winston GP, Tiwari VK, Kreilkamp BAK, Lenge M, Guerrini R, Hamandi K, Rüber T, Bauer T, Devinsky O, Striano P, Kaestner E, Hatton SN, Caciagli L, Kirschner M, Duncan JS, Thompson PM, McDonald CR, Sisodiya SM, Bernasconi N, Bernasconi A, Gan-Or Z, and Bernhardt BC

Abstract

Temporal lobe epilepsy with hippocampal sclerosis (TLE-HS) is associated with a complex genetic architecture, but the translation from genetic risk factors to brain vulnerability remains unclear. Here, we examined associations between epilepsy-related polygenic risk scores for HS (PRS-HS) and brain structure in a large sample of neurotypical children, and correlated these signatures with case-control findings in in multicentric cohorts of patients with TLE-HS. Imaging-genetic analyses revealed PRS-related cortical thinning in temporo-parietal and fronto-central regions, strongly anchored to distinct functional and structural network epicentres. Compared to disease-related effects derived from epilepsy case-control cohorts, structural correlates of PRS-HS mirrored atrophy and epicentre patterns in patients with TLE-HS. By identifying a potential pathway between genetic vulnerability and disease mechanisms, our findings provide new insights into the genetic underpinnings of structural alterations in TLE-HS and highlight potential imaging-genetic biomarkers for early risk stratification and personalized interventions., Competing Interests: POTENTIAL CONFLICTS OF INTEREST All authors declare no competing conflicts of interest.

Published
2025
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33. Cenobamate as add-on treatment for SCN8A developmental and epileptic encephalopathy.

Author

Gjerulfsen CE, Oudin MJ, Furia F, Gverdtsiteli S, Landmark CJ, Trivisano M, Balestrini S, Guerrini R, Aledo-Serrano A, Morcos R, Previtali R, Veggiotti P, Ricci E, Rubboli G, Gardella E, and Møller RS

Abstract

Objectives: Developmental and epileptic encephalopathies (DEEs) caused by pathogenic variants in SCN8A are associated with difficult-to-treat and early-onset seizures, developmental delay/intellectual disability, impaired quality of life, and increased risk of early mortality. High doses of sodium channel blockers are typically used to treat SCN8A-DEE caused by gain-of-function (GoF) variants. However, seizures are often drug resistant, and only a few patients achieve seizure freedom. In this retrospective study, the effect of cenobamate was assessed in patients with SCN8A-DEE., Methods: Across multiple centers and through collaborations with SCN8A patient advocacy organizations, patients with SCN8A-DEE treated with cenobamate for ≥6 months were identified. Data were obtained from patients' caregivers or treating physicians through a (Research Electronic Data Capture) REDCap survey. The functional effect of the SCN8A variants was obtained from the literature or assessed by prediction tools., Results: Twelve patients (3-25 years of age (median 8 years), 9 females) with presumed GoF SCN8A variants were treated with cenobamate for a mean period of 17 months (range 6-42 months). Countable motor seizures were meaningfully reduced in 10 of 12 patients (83%). Six patients experienced a seizure reduction above 70%, of which two achieved seizure freedom. In addition, two patients achieved a reduction in seizures ranging between 50% and 70%. An increase in seizure-free days per patient was also reported. Rescue medication was decreased in six of seven patients (85%) in need. Furthermore, 80% of patients reported non-seizure-related improvements, which included increased alertness, better sleep, and improved muscle tone. Adverse effects were reported by 50% of patients, and half resolved spontaneously or through the reduction of concomitant antiseizure medications., Significance: Our data suggest that cenobamate is a promising and safe treatment for SCN8A-DEE, even during early childhood. As a potential precision approach to treatment, cenobamate significantly reduced seizure burden and improved non-seizure-related symptoms. These positive outcomes may also be achieved in patient cohorts with GoF variants in other voltage-gated sodium channel genes., (© 2025 International League Against Epilepsy.)

Published
2025
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34. Heterozygous UBR5 variants result in a neurodevelopmental syndrome with developmental delay, autism, and intellectual disability.

Author

Sabeh P, Dumas SA, Maios C, Daghar H, Korzeniowski M, Rousseau J, Lines M, Guerin A, Millichap JJ, Landsverk M, Grebe T, Lindstrom K, Strober J, Ait Mouhoub T, Zweier C, Steinraths M, Hebebrand M, Callewaert B, Abou Jamra R, Kautza-Lucht M, Wegler M, Kruszka P, Kumps C, Banne E, Waberski MB, Dieux A, Raible S, Krantz I, Medne L, Pechter K, Villard L, Guerrini R, Bianchini C, Barba C, Mei D, Blanc X, Kallay C, Ranza E, Yang XR, O'Heir E, Donald KA, Murugasen S, Bruwer Z, Calikoglu M, Mathews JM, Lesieur-Sebellin M, Baujat G, Derive N, Pierson TM, Murrell JR, Shillington A, Ormieres C, Rondeau S, Reis A, Fernandez-Jaen A, Au PYB, Sweetser DA, Briere LC, Couque N, Perrin L, Schymick J, Gueguen P, Lefebvre M, Van Andel M, Juusola J, Antonarakis SE, Parker JA, Burnett BG, and Campeau PM

Subjects
Humans, Animals, Male, Female, Child, Child, Preschool, Ubiquitination, Adolescent, Phenotype, Infant, Adult, Ubiquitin-Protein Ligases genetics, Ubiquitin-Protein Ligases metabolism, Intellectual Disability genetics, Caenorhabditis elegans genetics, Developmental Disabilities genetics, Autistic Disorder genetics, Neurodevelopmental Disorders genetics, Heterozygote
Abstract

E3 ubiquitin ligases have been linked to developmental diseases including autism, Angelman syndrome (UBE3A), and Johanson-Blizzard syndrome (JBS) (UBR1). Here, we report variants in the E3 ligase UBR5 in 29 individuals presenting with a neurodevelopmental syndrome that includes developmental delay, autism, intellectual disability, epilepsy, movement disorders, and/or genital anomalies. Their phenotype is distinct from JBS due to the absence of exocrine pancreatic insufficiency and the presence of autism, epilepsy, and, in some probands, a movement disorder. E3 ubiquitin ligases are responsible for transferring ubiquitin to substrate proteins to regulate a variety of cellular functions, including protein degradation, protein-protein interactions, and protein localization. Knocking out ubr-5 in C. elegans resulted in a lower movement score compared to the wild type, supporting a role for UBR5 in neurodevelopment. Using an in vitro autoubiquitination assay and confocal microscopy for the human protein, we found decreased ubiquitination activity and altered cellular localization in several variants found in our cohort compared to the wild type. In conclusion, we found that variants in UBR5 cause a neurodevelopmental syndrome that can be associated with a movement disorder, reinforcing the role of the UBR protein family in a neurodevelopmental disease that differs from previously described ubiquitin-ligase-related syndromes. We also provide evidence for the pathogenic potential loss of UBR5 function with functional experiments in C. elegans and in vitro ubiquitination assays., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

Published
2025
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35. Bilateral Perisylvian Polymicrogyria, Intellectual Disability and Nephronophthisis Associated With Compound Heterozygous Pathogenic Variants in the CEP83  Gene.

Author

Parrini E, Balestrini S, Rutigliano D, Ricci ML, Mei D, and Guerrini R

Subjects
Humans, Male, Child, Preschool, Heterozygote, Cell Cycle Proteins genetics, Exome Sequencing, Phenotype, Cilia genetics, Cilia pathology, Mutation genetics, Abnormalities, Multiple, Intellectual Disability genetics, Intellectual Disability pathology, Kidney Diseases, Cystic genetics, Kidney Diseases, Cystic pathology, Malformations of Cortical Development genetics, Malformations of Cortical Development pathology
Abstract

The centrosomal protein 83 (CEP83) is a centriolar protein involved in primary cilium assembly, an early and critical step in ciliogenesis. Bi-allelic pathogenic variants in the CEP83 gene have been associated with infantile nephronophthisis and, in a few patients, retinitis pigmentosa. We describe a 5-year-old boy with bilateral perisylvian polymicrogyria, intellectual disability, and nephronophthisis in whom, using exome sequencing, we identified the c.1052T>G p.(Leu351*) stopgain variant inherited from the father and the c.2024T>C p.(Leu675Pro) missense variant inherited from the mother, in a compound heterozygous pattern. Polymicrogyria or, in general, malformations of cortical development had not been previously observed in patients with pathogenic CEP83 variants. However, defects in CEP83 can affect the formation and function of cilia or centrosomal structures, resulting in a polymicrogyric pattern overlapping with that associated with pathogenic variants affecting other genes coding for centrosomal components. This observation expands the spectrum of phenotypes associated with the CEP83 gene and adds it to the list of genes associated with bilateral perisylvian polymicrogyria., (© 2024 The Author(s). American Journal of Medical Genetics Part A published by Wiley Periodicals LLC.)

Published
2025
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36. Positively charged residues play a significant role in enhancing the antibacterial activity of calcitermin.

Author

Leveraro S, D'Accolti M, Marzola E, Caselli E, Guerrini R, Rowinska-Zyrek M, Remelli M, and Bellotti D

Subjects
Microbial Sensitivity Tests, Humans, Antimicrobial Cationic Peptides chemistry, Antimicrobial Cationic Peptides pharmacology, Coordination Complexes pharmacology, Coordination Complexes chemistry, Anti-Bacterial Agents pharmacology, Anti-Bacterial Agents chemistry, Zinc chemistry, Zinc pharmacology, Copper chemistry
Abstract

A systematic study on the human antimicrobial peptide calcitermin (VAIALKAAHYHTHKE) and its carefully designed derivatives was undertaken to verify the impact of divalent copper and zinc ions on the stability, coordination and antimicrobial activity of the formed complexes. In this work we investigate the calcitermin mutants where the alanine in position 7 and 8 is substituted with an arginine residue, with the aim of enhancing the antibacterial activity. Additionally, the analogue where alanine in position 7 is replaced with a histidine is considered, to obtain a chelating sequence with four histidines in alternate position; the aim of this change was to increase the cationic properties of the peptide under acidic conditions and possibly enhance its binding ability towards the metal ions. Through a comprehensive analytical approach involving potentiometric titrations, mass spectrometry, UV-Vis spectrophotometry, NMR and circular dichroism, we delved into the formation equilibria and coordination chemistry of the formed copper(II) and zinc(II) complexes. Antimicrobial assays are also performed to assess the bioactivity of the compounds against a broad spectrum of microorganisms, revealing the pivotal role of positively charged residues in enhancing the antibacterial activity of calcitermin. The obtained results serve as an important stepping stone towards the development of novel metal-based antimicrobial agents., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Published
2025
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37. National survey on the prevalence of single-gene aetiologies for genetic developmental and epileptic encephalopathies in Italy.

Author

Mei D, Balestrini S, Parrini E, Gambardella A, Annesi G, De Giorgis V, Gana S, Bassi MT, Zucca C, Elia M, Vetri L, Castellotti B, Ragona F, Mastrangelo M, Pisani F, d'Orsi G, Carella M, Pruna D, Giglio S, Marini C, Cesaroni E, Riva A, Scala M, Licchetta L, Minardi R, Contaldo I, Gambardella ML, Cossu A, Proietti J, Cantalupo G, Trivisano M, De Dominicis A, Specchio N, Tassi L, and Guerrini R

Subjects
Humans, Italy epidemiology, Male, Female, Child, Prevalence, Adolescent, Child, Preschool, Infant, Adult, Infant, Newborn, Middle Aged, Young Adult, Aged, Genetic Predisposition to Disease, Surveys and Questionnaires, Epilepsy genetics, Epilepsy epidemiology
Abstract

Background: We aimed to estimate real-world evidence of the prevalence rate of genetic developmental and epileptic encephalopathies (DEEs) in the Italian population over a 11-year period., Methods: Fifteen paediatric and adult tertiary Italian epilepsy centres participated in a survey related to 98 genes included in the molecular diagnostic workflows of most centres. We included patients with a clinical diagnosis of DEE, caused by a pathogenic or likely pathogenic variant in one of the selected genes, with a molecular diagnosis established between 2012 and 2022. These data were used as a proxy to estimate the prevalence rate of DEEs., Results: We included 1568 unique patients and found a mean incidence proportion of 2.6 patients for 100.000 inhabitants (SD=1.13) with consistent values across most Italian regions. The number of molecular diagnoses showed a continuing positive trend, resulting in more than a 10-fold increase between 2012 and 2022. The mean age at molecular diagnosis was 11.2 years (range 0-75). Pathogenic or likely pathogenic variants in genes with an autosomal dominant inheritance pattern occurred in 77% (n=1207) patients; 17% (n=271) in X-linked genes and 6% (n=90) in genes with autosomal recessive inheritance. The most frequently reported genes in the survey were SCN1A (16%), followed by KCNQ2 (5.6%) and SCN2A (5%)., Conclusion: Our study provides a large dataset of patients with monogenic DEE, from a European country. This is essential for informing decision-makers in drug development on the appropriateness of initiatives aimed at developing precision medicine therapies and is instrumental in implementing disease-specific registries and natural history studies., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2025. Re-use permitted under CC BY. Published by BMJ Group.)

Published
2024
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38. Fenfluramine treatment for Dravet syndrome: Long term real-world analysis demonstrates safety and reduced health care burden.

Author

Boncristiano A, Balestrini S, Doccini V, Specchio N, Pietrafusa N, Trivisano M, Darra F, Cossu A, Battaglia D, Quintiliani M, Gambardella ML, Parente E, Monni R, Matricardi S, Marini C, Ragona F, Granata T, Striano P, Riva A, and Guerrini R

Abstract

Objective: Fenfluramine (FFA), stiripentol (STP), and cannabidiol (CBD) are approved add-on therapies for seizures in Dravet syndrome (DS). We report on the long-term safety and health care resource utilization (HCRU) of patients with DS treated with FFA under an expanded access program (EAP)., Methods: A cohort of 124 patients received FFA for a median of 2.8 years (34.4 months). We compared data on safety and HCRU during FFA treatment with those from a same pre-treatment period. Echocardiography was conducted every 6 months. Information collected included gender, age, and auxological parameters (height, weight, and body mass index [BMI]) at the start (T0) and follow-up (T1); FFA treatment details (start, withdrawal, dosage); adverse events (AEs); and HCRU data including hospital admissions, status epilepticus (SE) episodes, and rescue medication use. We grouped patients by weight: ≤37.4 kg (n = 68, 54.8%) and ≥37.5 kg (n = 56; 45.1%), with FFA dosing adjusted accordingly. Statistical analyses included paired t test, Wilcoxon signed-rank test, Kaplan-Meier analysis, and Bonferroni correction to adjust for multiple testing., Results: Mean age was 47 months at clinical diagnosis and 81 months at T0. The last follow-up average FFA dose was .5 mg/kg/day, with a median of .4 mg/kg/day. FFA led to a 9.5% reduction in prior treatment load. At last follow-up, 118 of 124 (91.5%) remained on FFA. Rescue medication use decreased significantly from 4.5 to 1, hospitalizations from 1 to 0, and SE episodes from 0-240 to 0-180 (p < .001 for all). Seizure freedom was achieved in 9 of 118 patients (7.6%). AEs occurred in 39 of 124 patients (31.5%), with no cardiac issues or deaths. There was an overall mean reduction in BMI, with no statistical significance, and never requiring FFA withdrawal., Significance: FFA is well tolerated, without cardiac toxicity, and reduces treatment load and HCRU, suggesting improved patient management. BMI reduction in young children highlights the need for growth and nutritional monitoring., (© 2024 The Author(s). Epilepsia published by Wiley Periodicals LLC on behalf of International League Against Epilepsy.)

Published
2024
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39. RICTOR variants are associated with neurodevelopmental disorders.

Author

Carapito R, Molitor A, Pavinato L, Skeyni A, Lambert M, Pichot A, Jiang J, Spinnhirny P, Zimmermann L, Boucher P, Chung CWT, Elserafy N, Blair EM, Li D, Elisabeth B, Kotzaeridou U, Karch S, Wagner M, Lunsing RJ, Pfundt R, Boycott KM, Bruel AL, Mau-Them FT, Moutton S, Conti V, Mei D, Cetica V, Guerrini R, Brunet T, Rump P, Mussa A, Brusco A, Lemire G, de Vries BBA, Miao Z, Isidor B, and Bahram S

Abstract

RICTOR is a key component of the mTORC2 signaling complex which is involved in the regulation of cell growth, proliferation and survival. RICTOR is highly expressed in neurons and is necessary for brain development. Here, we report eight unrelated patients presenting with intellectual disability and/or development delay and carrying variants in the RICTOR gene. The phenotypic presentation is diverse with associated features including growth failure, feeding difficulties, abnormal behavior, seizure, hypertonia, brain anomalies and various other congenital organ and skeletal malformations. All patients carried de novo or heterozygous variants inherited from one affected parent, including three missense variants, four loss-of-function variants and one 3 kb deletion encompassing RICTOR. The mTORC2 pathway was hyperactivated in a patient's fibroblasts carrying a missense variant, while the expression of RICTOR remained unchanged, indicating a gain-of-function mechanism. RNA sequencing on RICTOR knock-out mouse embryonic fibroblasts confirmed the potential role of RICTOR in neuronal cell development., Competing Interests: Competing interests: The authors declare no competing interests. Ethics approval: All sequencing experiments were performed upon written informed consent for clinical sequencing and/or by center-specific institutional review board approval for research sequencing., (© 2024. The Author(s), under exclusive licence to European Society of Human Genetics.)

Published
2024
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40. Rare dysfunctional SCN2A variants are associated with malformation of cortical development.

Author

Clatot J, Thompson CH, Sotardi S, Jiang J, Trivisano M, Balestrini S, Ward DI, Ginn N, Guaragni B, Malerba L, Vakrinou A, Sherer M, Helbig I, Somarowthu A, Sisodiya SM, Ben-Shalom R, Guerrini R, Specchio N, George AL Jr, and Goldberg EM

Abstract

Objective: SCN2A encodes the voltage-gated sodium (Na+) channel α subunit Na V 1.2, which is important for the generation and forward and back propagation of action potentials in neurons. Genetic variants in SCN2A are associated with a spectrum of neurodevelopmental disorders. However, the mechanisms whereby variation in SCN2A leads to disease remains incompletely understood, and the full spectrum of SCN2A-related disorders may not be fully delineated., Methods: Here, we identified seven de novo heterozygous variants in SCN2A in eight individuals with developmental and epileptic encephalopathy (DEE) accompanied by prominent malformation of cortical development (MCD). We characterized the electrophysiological properties of Na + currents in human embryonic kidney (HEK) cells transfected with the adult (A) or neonatal (N) isoform of wild-type (WT) and variant Na V 1.2 using manual and automated whole-cell voltage clamp recording., Results: The neonatal isoforms of all SCN2A variants studied exhibit gain of function (GoF) with a large depolarized shift in steady-state inactivation, creating a markedly enhanced window current common across all four variants tested. Computational modeling demonstrated that expression of the Na V 1.2-p.Met1770Leu-N variant in a developing neocortical pyramidal neuron results in hyperexcitability., Significance: These results support expansion of the clinical spectrum of SCN2A-related disorders and the association of genetic variation in SCN2A with MCD, which suggests previously undescribed roles for SCN2A in fetal brain development., (© 2024 The Author(s). Epilepsia published by Wiley Periodicals LLC on behalf of International League Against Epilepsy.)

Published
2024
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41. Probing non-peptide agonists binding at the human nociceptin/orphanin FQ receptor: a molecular modelling study.

Author

Gozzi M, Malfacini D, Albanese V, Pacifico S, Preti D, Guerrini R, Calò G, and Ciancetta A

Abstract

The N/OFQ-NOP receptor is a fascinating peptidergic system with the potential to be exploited for the development of analgesic drugs devoid of side effects associated with classical opioid signalling modulation. To date, up to four X-ray and cryo-EM structures of the NOP receptor in complex with the endogenous peptide agonist N/OFQ and three small molecule antagonists have been solved and released. Despite the available structural information, the details of selective small molecule agonist binding to the NOP receptor in the active state remain elusive. In this study, by leveraging the available structural information and using N/OFQ(1-13)-NH 2 as a reference compound, we developed a computational protocol based on docking followed by short molecular dynamics (MD) simulations that can suggest small molecule agonist binding modes at the NOP receptor that are reproducible and stable over time in the solvated membrane-embedded receptor active state and in agreement with known structure-activity relationship (SAR) data., Competing Interests: The authors declare no competing financial interest., (This journal is © The Royal Society of Chemistry.)

Published
2024
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42. Multiorgan manifestations of COL4A1 and COL4A2 variants and proposal for a clinical management protocol.

Author

Gasparini S, Balestrini S, Saccaro LF, Bacci G, Panichella G, Montomoli M, Cantalupo G, Bigoni S, Mancano G, Pellacani S, Leuzzi V, Volpi N, Mari F, Melani F, Cavallin M, Pisano T, Porcedda G, Vaglio A, Mei D, Barba C, Parrini E, and Guerrini R

Subjects
Humans, Female, Male, Adult, Child, Adolescent, Child, Preschool, Epilepsy genetics, Epilepsy etiology, Epilepsy therapy, Epilepsy pathology, Intellectual Disability genetics, Intellectual Disability pathology, Seizures genetics, Stroke genetics, Stroke pathology, Cataract genetics, Cataract pathology, Middle Aged, Hematuria genetics, Porencephaly genetics, Phenotype, Developmental Disabilities genetics, Developmental Disabilities etiology, Developmental Disabilities pathology, Mutation, Collagen Type IV genetics
Abstract

COL4A1/2 variants are associated with highly variable multiorgan manifestations. Depicting the whole clinical spectrum of COL4A1/2-related manifestations is challenging, and there is no consensus on management and preventative strategies. Based on a systematic review of current evidence on COL4A1/2-related disease, we developed a clinical questionnaire that we administered to 43 individuals from 23 distinct families carrying pathogenic variants. In this cohort, we extended ophthalmological and cardiological examinations to asymptomatic individuals and those with only limited or mild, often nonspecific, clinical signs commonly occurring in the general population (i.e., oligosymptomatic). The most frequent clinical findings emerging from both the literature review and the questionnaire included stroke (203/685, 29.6%), seizures or epilepsy (199/685, 29.0%), intellectual disability or developmental delay (168/685, 24.5%), porencephaly/schizencephaly (168/685, 24.5%), motor impairment (162/685, 23.6%), cataract (124/685, 18.1%), hematuria (63/685, 9.2%), and retinal arterial tortuosity (58/685, 8.5%). In oligosymptomatic and asymptomatic carriers, ophthalmological investigations detected retinal vascular tortuosity (5/13, 38.5%), dysgenesis of the anterior segment (4/13, 30.8%), and cataract (2/13, 15.4%), while cardiological investigations were unremarkable except for mild ascending aortic ectasia in 1/8 (12.5%). Our multimodal approach confirms highly variable penetrance and expressivity in COL4A1/2-related conditions, even at the intrafamilial level with neurological involvement being the most frequent and severe finding in both children and adults. We propose a protocol for prevention and management based on individualized risk estimation and periodic multiorgan evaluations., (© 2024 The Author(s). American Journal of Medical Genetics Part C: Seminars in Medical Genetics published by Wiley Periodicals LLC.)

Published
2024
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43. Enhanced anticancer effect of thymidylate synthase dimer disrupters by promoting intracellular accumulation.

Author

Marverti G, Moschella MG, Belardo A, Lamesta M, Mercanile G, Tagliazucchi L, Aiello D, Venturelli A, Illuminati D, Guerrini R, Losi L, Ponterini G, Costi MP, and D'Arca D

Abstract

Introduction: Thymidylate synthase (TS) plays a crucial role in cellular growth, proliferation, DNA synthesis, and repair, thus gaining attention for targeted therapies in cancer. TS overexpression and the altered pharmacokinetics of anti-TS drugs are among the most prominent causes of cellular resistance. Decreased drug influx and/or efficient efflux result in reduced drug access to the intracellular targets., Results: In this study, we have evaluated and demonstrated the increased cytotoxic efficacy of novel TS dimer disrupters (Ddis) in the presence of specific inhibitors of drug efflux protein pumps in ovarian and colon cancer cells, suggesting that these compounds are substrates of the cellular drug extruders. A second strategy adopted to favor intracellular accumulation was to employ, as a drug delivery system, a molecular tool able to help less lipophilic compounds to cross the cell membrane. The Ddis were delivered through the SAINT-Protein transfection agent. The observed cell-killing effects agreed with the reduction of TS protein level and cell cycle perturbation., Conclusion: Overall, this preclinical study suggests that the innovative TS dimer disrupters can be optimized by increasing their intracellular accumulation by both inhibiting their outflow and/or enhancing cellular uptake., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Marverti, Moschella, Belardo, Lamesta, Mercanile, Tagliazucchi, Aiello, Venturelli, Illuminati, Guerrini, Losi, Ponterini, Costi and D’Arca.)

Published
2024
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44. A Multi-Angle Approach to Predict Peptide-GPCR Complexes: The N/OFQ-NOP System as a Successful AlphaFold Application Case Study.

Author

Ciancetta A, Malfacini D, Gozzi M, Marzola E, Camilotto R, Calò G, and Guerrini R

Subjects
Opioid Peptides chemistry, Receptors, Opioid chemistry, Receptors, Opioid metabolism, Peptides chemistry, Artificial Intelligence, Nociceptin Receptor, Humans, Protein Conformation, Protein Binding, Nociceptin, Receptors, G-Protein-Coupled chemistry, Receptors, G-Protein-Coupled metabolism, Ligands, Models, Molecular, Amino Acid Sequence, Molecular Docking Simulation
Abstract

With nearly 700 structures solved and a growing number of customized structure prediction algorithms being developed at a fast pace, G protein-coupled receptors (GPCRs) are an optimal test case for validating new approaches for the prediction of receptor active state and ligand bioactive conformation complexes. In this study, we leveraged the availability of hundreds of peptide GPCRs in the active state and both classical homology and artificial intelligence (AI) based protein modeling combined with docking and AI-based peptide structure prediction approaches to predict the nociceptin/orphanin FQ-NOP receptor active state complex (N/OFQ-NOPa). The In Silico generated hypotheses were validated via the design, synthesis, and pharmacological characterization of novel linear N/OFQ(1-13)-NH 2 analogues, leading to the discovery of a novel antagonist ( 3B ; p K B = 6.63) bearing a single ring-constrained residue in place of the Gly 2 -Gly 3 motif of the N/OFQ message sequence (FGGF). While the experimental validation was ongoing, the availability of the Cryo-EM structure of the predicted complex enabled us to unambiguously validate the generated hypotheses. To the best of our knowledge, this is the first example of a peptide-GPCR complex predicted with atomistic accuracy (full complex Cα RMSD < 1.0 Å) and of the N/OFQ message moiety being successfully modified with a rigid scaffold.

Published
2024
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45. Endocrinological features and epileptic encephalopathy in COX deficiency due to SCO1 mutations: case series and review of literature.

Author

Barbato A, Gori G, Sacchini M, Pochiero F, Bargiacchi S, Traficante G, Palazzo V, Tiberi L, Bianchini C, Mei D, Parrini E, Pisano T, Procopio E, Guerrini R, Peron A, and Stagi S

Abstract

Context: Cytochrome C oxidase (COX) is the fourth component of the respiratory chain and is located within the internal membrane of mitochondria. COX deficiency causes an inherited mitochondrial disease with significant genetic and phenotypic heterogeneity. Four clinical subtypes have been identified, each with distinct phenotypes and genetic variants. Mitochondrial complex IV deficiency nuclear type 4 (MC4DN4) is a form of COX deficiency associated with pathogenic variants in the SCO1 gene., Case Description: We describe three patients with MC4DN4 with developmental and epileptic encephalopathy (DEE), hypopituitarism, and SCO1 pathogenic variants. These patients' phenotypes considerably differ from previously reported MC4DN4 phenotypes as they associate DEE with progressive hypopituitarism and survival beyond the first months after birth. Pituitary deficiency in these patients progressively worsened and mainly involved growth hormone secretion and thyroid function., Conclusions: Our findings expand knowledge of phenotypic variability in MC4DN4 and suggest that SCO1 is a candidate gene for genetic hypopituitarism and DEE., Significance Statement: Our paper describes three patients affected by MC4DN4 with hypopituitarism and developmental and epileptic encephalopathy (DEE), two features that have never been associated with this condition. In addition, we reviewed the clinical features of all previous cases of MC4DN4 to give the other clinicians a wide picture of the clinical phenotype of this genetic disease. We hope that the publication of our data may help others to identify this disease and consider the chance to analyze the SCO1 gene in cases of DEE associated with pituitary dysfunction. Our article contributes to expanding the spectrum of genetic hypopituitarism and proposes a model to explain an association between this condition, mitochondrial anomalies, and neurodevelopmental defects.

Published
2024
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46. Double gonosomal mosaicism as an unusual hereditary mechanism in familial GRIN2A -related disorder.

Author

Cetica V, Cavallin M, Ricci ML, Mandorlini C, Bartolini E, Parrini E, and Guerrini R

Subjects
Humans, Male, Female, Mutation genetics, Intellectual Disability genetics, Intellectual Disability pathology, Epilepsy genetics, Epilepsy pathology, Child, Alleles, Adult, Mosaicism, Receptors, N-Methyl-D-Aspartate genetics, Pedigree
Abstract

We aim to describe double gonosomal mosaicism in the GRIN2A gene in a mother who passed on two different pathogenic variants at the same nucleotide to her two affected children. We studied a boy with epilepsy and intellectual disability, along with his sister and mother who exhibited language impairment and learning difficulties without epilepsy. We identified in the proband a splice-site variant in GRIN2A (c.1008-1G>A) inherited from his mother. Subsequent testing of his sister revealed a different change at the same nucleotide c.1008-1G>T, which was also present in the mother's DNA at 3.9% allele frequency. The co-occurrence of two mutational events at the same nucleotide is extremely rare. Since a chance occurrence is unlikely, we hypothesise that a base mismatch may introduce instability triggering a second event. In this family, the mother carries three alleles, of which one is at very low frequency. This complex genetic landscape poses diagnostic challenges since low-level mosaicism may escape detection via conventional methods. Applying specific technology becomes crucial, as double mosaicism might prove to be more prevalent than anticipated severely impacting diagnostic accuracy and genetic counselling., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2024. No commercial re-use. See rights and permissions. Published by BMJ.)

Published
2024
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47. Exome sequencing of 20,979 individuals with epilepsy reveals shared and distinct ultra-rare genetic risk across disorder subtypes.

Author

Chen S, Abou-Khalil BW, Afawi Z, Ali QZ, Amadori E, Anderson A, Anderson J, Andrade DM, Annesi G, Arslan M, Auce P, Bahlo M, Baker MD, Balagura G, Balestrini S, Banks E, Barba C, Barboza K, Bartolomei F, Bass N, Baum LW, Baumgartner TH, Baykan B, Bebek N, Becker F, Bennett CA, Beydoun A, Bianchini C, Bisulli F, Blackwood D, Blatt I, Borggräfe I, Bosselmann C, Braatz V, Brand H, Brockmann K, Buono RJ, Busch RM, Caglayan SH, Canafoglia L, Canavati C, Castellotti B, Cavalleri GL, Cerrato F, Chassoux F, Cherian C, Cherny SS, Cheung CL, Chou IJ, Chung SK, Churchhouse C, Ciullo V, Clark PO, Cole AJ, Cosico M, Cossette P, Cotsapas C, Cusick C, Daly MJ, Davis LK, Jonghe P, Delanty N, Dennig D, Depondt C, Derambure P, Devinsky O, Di Vito L, Dickerson F, Dlugos DJ, Doccini V, Doherty CP, El-Naggar H, Ellis CA, Epstein L, Evans M, Faucon A, Feng YA, Ferguson L, Ferraro TN, Da Silva IF, Ferri L, Feucht M, Fields MC, Fitzgerald M, Fonferko-Shadrach B, Fortunato F, Franceschetti S, French JA, Freri E, Fu JM, Gabriel S, Gagliardi M, Gambardella A, Gauthier L, Giangregorio T, Gili T, Glauser TA, Goldberg E, Goldman A, Goldstein DB, Granata T, Grant R, Greenberg DA, Guerrini R, Gundogdu-Eken A, Gupta N, Haas K, Hakonarson H, Haryanyan G, Häusler M, Hegde M, Heinzen EL, Helbig I, Hengsbach C, Heyne H, Hirose S, Hirsch E, Ho CJ, Hoeper O, Howrigan DP, Hucks D, Hung PC, Iacomino M, Inoue Y, Inuzuka LM, Ishii A, Jehi L, Johnson MR, Johnstone M, Kälviäinen R, Kanaan M, Kara B, Kariuki SM, Kegele J, Kesim Y, Khoueiry-Zgheib N, Khoury J, King C, Klein KM, Kluger G, Knake S, Kok F, Korczyn AD, Korinthenberg R, Koupparis A, Kousiappa I, Krause R, Krenn M, Krestel H, Krey I, Kunz WS, Kurlemann G, Kuzniecky RI, Kwan P, La Vega-Talbott M, Labate A, Lacey A, Lal D, Laššuthová P, Lauxmann S, Lawthom C, Leech SL, Lehesjoki AE, Lemke JR, Lerche H, Lesca G, Leu C, Lewin N, Lewis-Smith D, Li GH, Liao C, Licchetta L, Lin CH, Lin KL, Linnankivi T, Lo W, Lowenstein DH, Lowther C, Lubbers L, Lui CHT, Macedo-Souza LI, Madeleyn R, Madia F, Magri S, Maillard L, Marcuse L, Marques P, Marson AG, Matthews AG, May P, Mayer T, McArdle W, McCarroll SM, McGoldrick P, McGraw CM, McIntosh A, McQuillan A, Meador KJ, Mei D, Michel V, Millichap JJ, Minardi R, Montomoli M, Mostacci B, Muccioli L, Muhle H, Müller-Schlüter K, Najm IM, Nasreddine W, Neaves S, Neubauer BA, Newton CRJC, Noebels JL, Northstone K, Novod S, O'Brien TJ, Owusu-Agyei S, Özkara Ç, Palotie A, Papacostas SS, Parrini E, Pato C, Pato M, Pendziwiat M, Pennell PB, Petrovski S, Pickrell WO, Pinsky R, Pinto D, Pippucci T, Piras F, Piras F, Poduri A, Pondrelli F, Posthuma D, Powell RHW, Privitera M, Rademacher A, Ragona F, Ramirez-Hamouz B, Rau S, Raynes HR, Rees MI, Regan BM, Reif A, Reinthaler E, Rheims S, Ring SM, Riva A, Rojas E, Rosenow F, Ryvlin P, Saarela A, Sadleir LG, Salman B, Salmon A, Salpietro V, Sammarra I, Scala M, Schachter S, Schaller A, Schankin CJ, Scheffer IE, Schneider N, Schubert-Bast S, Schulze-Bonhage A, Scudieri P, Sedláčková L, Shain C, Sham PC, Shiedley BR, Siena SA, Sills GJ, Sisodiya SM, Smoller JW, Solomonson M, Spalletta G, Sparks KR, Sperling MR, Stamberger H, Steinhoff BJ, Stephani U, Štěrbová K, Stewart WC, Stipa C, Striano P, Strzelczyk A, Surges R, Suzuki T, Talarico M, Talkowski ME, Taneja RS, Tanteles GA, Timonen O, Timpson NJ, Tinuper P, Todaro M, Topaloglu P, Tsai MH, Tumiene B, Turkdogan D, Uğur-İşeri S, Utkus A, Vaidiswaran P, Valton L, van Baalen A, Vari MS, Vetro A, Vlčková M, von Brauchitsch S, von Spiczak S, Wagner RG, Watts N, Weber YG, Weckhuysen S, Widdess-Walsh P, Wiebe S, Wolf SM, Wolff M, Wolking S, Wong I, von Wrede R, Wu D, Yamakawa K, Yapıcı Z, Yis U, Yolken R, Yücesan E, Zagaglia S, Zahnert F, Zara F, Zimprich F, Zizovic M, Zsurka G, Neale BM, and Berkovic SF

Abstract

Identifying genetic risk factors for highly heterogeneous disorders like epilepsy remains challenging. Here, we present the largest whole-exome sequencing study of epilepsy to date, with >54,000 human exomes, comprising 20,979 deeply phenotyped patients from multiple genetic ancestry groups with diverse epilepsy subtypes and 33,444 controls, to investigate rare variants that confer disease risk. These analyses implicate seven individual genes, three gene sets, and four copy number variants at exome-wide significance. Genes encoding ion channels show strong association with multiple epilepsy subtypes, including epileptic encephalopathies, generalized and focal epilepsies, while most other gene discoveries are subtype-specific, highlighting distinct genetic contributions to different epilepsies. Combining results from rare single nucleotide/short indel-, copy number-, and common variants, we offer an expanded view of the genetic architecture of epilepsy, with growing evidence of convergence among different genetic risk loci on the same genes. Top candidate genes are enriched for roles in synaptic transmission and neuronal excitability, particularly postnatally and in the neocortex. We also identify shared rare variant risk between epilepsy and other neurodevelopmental disorders. Our data can be accessed via an interactive browser, hopefully facilitating diagnostic efforts and accelerating the development of follow-up studies., Competing Interests: Competing Interests B.M.N is a member of the scientific advisory board at Deep Genomics and Neumora. No other authors have competing interests to declare

Published
2024
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48. Quantitative cytoarchitectural phenotyping of deparaffinized human brain tissues.

Author

Meo DD, Sorelli M, Ramazzotti J, Cheli F, Bradley S, Perego L, Lorenzon B, Mazzamuto G, Emmi A, Porzionato A, Caro R, Garbelli R, Biancheri D, Pelorosso C, Conti V, Guerrini R, Pavone FS, and Costantini I

Abstract

Advanced 3D imaging techniques and image segmentation and classification methods can profoundly transform biomedical research by offering deep insights into the cytoarchitecture of the human brain in relation to pathological conditions. Here, we propose a comprehensive pipeline for performing 3D imaging and automated quantitative cellular phenotyping on Formalin-Fixed Paraffin-Embedded (FFPE) human brain specimens, a valuable yet underutilized resource. We exploited the versatility of our method by applying it to different human specimens from both adult and pediatric, normal and abnormal brain regions. Quantitative data on neuronal volume, ellipticity, local density, and spatial clustering level were obtained from a machine learning-based analysis of the 3D cytoarchitectural organization of cells identified by different molecular markers in two subjects with malformations of cortical development (MCD). This approach will grant access to a wide range of physiological and pathological paraffin-embedded clinical specimens, allowing for volumetric imaging and quantitative analysis of human brain samples at cellular resolution. Possible genotype-phenotype correlations can be unveiled, providing new insights into the pathogenesis of various brain diseases and enlarging treatment opportunities.

Published
2024
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50. Developmental and epileptic encephalopathies.

Author

Scheffer IE, Zuberi S, Mefford HC, Guerrini R, and McTague A

Subjects
Humans, Epilepsy diagnosis, Epilepsy genetics, Epilepsy therapy
Abstract

Developmental and epileptic encephalopathies, the most severe group of epilepsies, are characterized by seizures and frequent epileptiform activity associated with developmental slowing or regression. Onset typically occurs in infancy or childhood and includes many well-defined epilepsy syndromes. Patients have wide-ranging comorbidities including intellectual disability, psychiatric features, such as autism spectrum disorder and behavioural problems, movement and musculoskeletal disorders, gastrointestinal and sleep problems, together with an increased mortality rate. Problems change with age and patients require substantial support throughout life, placing a high psychosocial burden on parents, carers and the community. In many patients, the aetiology can be identified, and a genetic cause is found in >50% of patients using next-generation sequencing technologies. More than 900 genes have been identified as monogenic causes of developmental and epileptic encephalopathies and many cell components and processes have been implicated in their pathophysiology, including ion channels and transporters, synaptic proteins, cell signalling and metabolism and epigenetic regulation. Polygenic risk score analyses have shown that common variants also contribute to phenotypic variability. Holistic management, which encompasses antiseizure therapies and care for multimorbidities, is determined both by epilepsy syndrome and aetiology. Identification of the underlying aetiology enables the development of precision medicines to improve the long-term outcome of patients with these devastating diseases., (© 2024. Springer Nature Limited.)

Published
2024
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