Publications by authors named "Liliana Vercelli"

40 Publications

Large genotype-phenotype study in carriers of D4Z4 borderline alleles provides guidance for facioscapulohumeral muscular dystrophy diagnosis.

Sci Rep 2020 12 10;10(1):21648. Epub 2020 Dec 10.

Department of Life Sciences, University of Modena and Reggio Emilia, Modena, Italy.

Facioscapulohumeral muscular dystrophy (FSHD) is a myopathy with prevalence of 1 in 20,000. Almost all patients affected by FSHD carry deletions of an integral number of tandem 3.3 kilobase repeats, termed D4Z4, located on chromosome 4q35. Assessment of size of D4Z4 alleles is commonly used for FSHD diagnosis. However, the extended molecular testing has expanded the spectrum of clinical phenotypes. In particular, D4Z4 alleles with 9-10 repeat have been found in healthy individuals, in subjects with FSHD or affected by other myopathies. These findings weakened the strict relationship between observed phenotypes and their underlying genotypes, complicating the interpretation of molecular findings for diagnosis and genetic counseling. In light of the wide clinical variability detected in carriers of D4Z4 alleles with 9-10 repeats, we applied a standardized methodology, the Comprehensive Clinical Evaluation Form (CCEF), to describe and characterize the phenotype of 244 individuals carrying D4Z4 alleles with 9-10 repeats (134 index cases and 110 relatives). The study shows that 54.5% of index cases display a classical FSHD phenotype with typical facial and scapular muscle weakness, whereas 20.1% present incomplete phenotype with facial weakness or scapular girdle weakness, 6.7% display minor signs such as winged scapula or hyperCKemia, without functional motor impairment, and 18.7% of index cases show more complex phenotypes with atypical clinical features. Family studies revealed that 70.9% of relatives carrying 9-10 D4Z4 reduced alleles has no motor impairment, whereas a few relatives (10.0%) display a classical FSHD phenotype. Importantly all relatives of index cases with no FSHD phenotype were healthy carriers. These data establish the low penetrance of D4Z4 alleles with 9-10 repeats. We recommend the use of CCEF for the standardized clinical assessment integrated by family studies and further molecular investigation for appropriate diagnosis and genetic counseling. Especially in presence of atypical phenotypes and/or sporadic cases with all healthy relatives is not possible to perform conclusive diagnosis of FSHD, but all these cases need further studies for a proper diagnosis, to search novel causative genetic defects or investigate environmental factors or co-morbidities that may trigger the pathogenic process. These evidences are also fundamental for the stratification of patients eligible for clinical trials. Our work reinforces the value of large genotype-phenotype studies to define criteria for clinical practice and genetic counseling in rare diseases.
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http://dx.doi.org/10.1038/s41598-020-78578-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7730397PMC
December 2020

Clinical and Molecular Spectrum of Myotonia and Periodic Paralyses Associated With Mutations in in a Large Cohort of Italian Patients.

Front Neurol 2020 29;11:646. Epub 2020 Jul 29.

Neuroimmunology and Neuromuscular Diseases, Fondazione IRCCS Istituto Neurologico Carlo Besta, Milan, Italy.

Four main clinical phenotypes have been traditionally described in patients mutated in SCN4A, including sodium-channel myotonia (SCM), paramyotonia congenita (PMC), Hypokaliemic type II (HypoPP2), and Hyperkaliemic/Normokaliemic periodic paralysis (HyperPP/NormoPP); in addition, rare phenotypes associated with mutations in SCN4A are congenital myasthenic syndrome and congenital myopathy. However, only scarce data have been reported in literature on large patient cohorts including phenotypes characterized by myotonia and episodes of paralysis. We retrospectively investigated clinical and molecular features of 80 patients fulfilling the following criteria: (1) clinical and neurophysiological diagnosis of myotonia, or clinical diagnosis of PP, and (2) presence of a pathogenic SCN4A gene variant. Patients presenting at birth with episodic laryngospasm or congenital myopathy-like phenotype with later onset of myotonia were considered as neonatal SCN4A. PMC was observed in 36 (45%) patients, SCM in 30 (37.5%), Hyper/NormoPP in 7 (8.7%), HypoPP2 in 3 (3.7%), and neonatal SCN4A in 4 (5%). The median age at onset was significantly earlier in PMC than in SCM ( < 0.01) and in Hyper/NormoPP than in HypoPP2 ( = 0.02). Cold-induced myotonia was more frequently observed in PMC ( = 34) than in SCM ( = 23) ( = 0.04). No significant difference was found in age at onset of episodes of paralysis among PMC and PP or in frequency of permanent weakness between PP ( = 4), SCM ( = 5), and PMC ( = 10). PP was more frequently associated with mutations in the S4 region of the NaV1.4 channel protein compared to SCM and PMC ( < 0.01); mutations causing PMC were concentrated in the C-terminal region of the protein, while SCM-associated mutations were detected in all the protein domains. Our data suggest that skeletal muscle channelopathies associated with mutations in SCN4A represent a continuum in the clinical spectrum.
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http://dx.doi.org/10.3389/fneur.2020.00646DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7403394PMC
July 2020

A 5-year clinical follow-up study from the Italian National Registry for FSHD.

J Neurol 2021 Jan 19;268(1):356-366. Epub 2020 Aug 19.

Department of Biomedical, Metabolic and Neural Sciences, University of Modena and Reggio Emilia, via G. Campi 287, 41125, Modena, Italy.

Background: The natural history of facioscapulohumeral muscular dystrophy (FSHD) is undefined.

Methods: An observational cohort study was conducted in 246 FSHD1 patients. We split the analysis between index cases and carrier relatives and we classified all patients using the Comprehensive Clinical Evaluation Form (CCEF). The disease progression was measured as a variation of the FSHD score performed at baseline and at the end of 5-year follow-up (ΔFSHD score).

Findings: Disease worsened in 79.4% (112/141) of index cases versus 38.1% (40/105) of carrier relatives and advanced more rapidly in index cases (ΔFSHD score 2.3 versus 1.2). The 79.1% (38/48) of asymptomatic carriers remained asymptomatic. The highest ΔFSHD score (1.7) was found in subject with facial and scapular weakness at baseline (category A), whereas in subjects with incomplete phenotype (facial or scapular weakness, category B) had lower ΔFSHD score (0.6) p < 0.0001.

Conclusions: The progression of disease is different between index cases and carrier relatives and the assessment of the CCEF categories has strong prognostic effect in FSHD1 patients.
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http://dx.doi.org/10.1007/s00415-020-10144-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7815626PMC
January 2021

Cytokine Profile in Striated Muscle Laminopathies: New Promising Biomarkers for Disease Prediction.

Cells 2020 06 23;9(6). Epub 2020 Jun 23.

Neurology IV-Neuroimmunology and Neuromuscular Diseases Unit, Fondazione IRCCS Istituto Neurologico Carlo Besta, 20133 Milan, Italy.

Laminopathies are a wide and heterogeneous group of rare human diseases caused by mutations of the LMNA gene or related nuclear envelope genes. The variety of clinical phenotypes and the wide spectrum of histopathological changes among patients carrying an identical mutation in the LMNA gene make the prognostic process rather difficult, and classical genetic screens appear to have limited predictive value for disease development. The aim of this study was to evaluate whether a comprehensive profile of circulating cytokines may be a useful tool to differentiate and stratify disease subgroups, support clinical follow-ups and contribute to new therapeutic approaches. Serum levels of 51 pro- and anti-inflammatory molecules, including cytokines, chemokines and growth factors, were quantified by a Luminex multiple immune-assay in 53 patients with muscular laminopathy (Musc-LMNA), 10 with non-muscular laminopathy, 22 with other muscular disorders and in 35 healthy controls. Interleukin-17 (IL-17), granulocyte colony-stimulating factor (G-CSF) and transforming growth factor beta (TGF-β2) levels significantly discriminated Musc-LMNA from controls; interleukin-1β (IL-1β), interleukin-4 (IL-4) and interleukin-8 (IL-8) were differentially expressed in Musc-LMNA patients compared to those with non-muscular laminopathies, whereas IL-17 was significantly higher in Musc-LMNA patients with muscular and cardiac involvement. These findings support the hypothesis of a key role of the immune system in Musc-LMNA and emphasize the potential use of cytokines as biomarkers for these disorders.
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http://dx.doi.org/10.3390/cells9061532DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7348753PMC
June 2020

Phenotypic Variability Among Patients With D4Z4 Reduced Allele Facioscapulohumeral Muscular Dystrophy.

JAMA Netw Open 2020 05 1;3(5):e204040. Epub 2020 May 1.

Department of Life Sciences, University of Modena and Reggio Emilia, Modena, Italy.

Importance: Facioscapulohumeral muscular dystrophy (FSHD) is considered an autosomal dominant disorder, associated with the deletion of tandemly arrayed D4Z4 repetitive elements. The extensive use of molecular analysis of the D4Z4 locus for FSHD diagnosis has revealed wide clinical variability, suggesting that subgroups of patients exist among carriers of the D4Z4 reduced allele (DRA).

Objective: To investigate the clinical expression of FSHD in the genetic subgroup of carriers of a DRA with 7 to 8 repeat units (RUs).

Design, Setting, And Participants: This multicenter cross-sectional study included 422 carriers of DRA with 7 to 8 RUs (187 unrelated probands and 235 relatives) from a consecutive sample of 280 probands and 306 relatives from the Italian National Registry for FSHD collected between 2008 and 2016. Participants were evaluated by the Italian Clinical Network for FSHD, and all clinical and molecular data were collected in the Italian National Registry for FSHD database. Data analysis was conducted from January 2017 to June 2018.

Main Outcomes And Measures: The phenotypic classification of probands and relatives was obtained by applying the Comprehensive Clinical Evaluation Form which classifies patients in the 4 following categories: (1) participants presenting facial and scapular girdle muscle weakness typical of FSHD (category A, subcategories A1-A3), (2) participants with muscle weakness limited to scapular girdle or facial muscles (category B, subcategories B1 and B2), (3) asymptomatic or healthy participants (category C, subcategories C1 and C2), and (4) participants with myopathic phenotypes presenting clinical features not consistent with FSHD canonical phenotype (category D, subcategories D1 and D2).

Results: A total of 187 probands (mean [SD] age at last neurological examination, 53.5 [15.2] years; 103 [55.1%] men) and 235 relatives (mean [SD] age at last neurologic examination, 45.1 [17.0] years; 104 [44.7%] men) with a DRA with 7 to 8 RUs and a molecular diagnosis of FSHD were evaluated. Of 187 probands, 99 (52.9%; 95% CI, 45.7%-60.1%) displayed the classic FSHD phenotype, whereas 86 (47.1%; 95% CI, 39.8%-54.3%) presented incomplete or atypical phenotypes. Of 235 carrier relatives from 106 unrelated families, 124 (52.8%; 95% CI, 46.4%-59.7%) had no motor impairment, whereas a small number (38 [16.2%; 95% CI, 9.8%-23.1%]) displayed the classic FSHD phenotype, and 73 (31.0%; 95% CI, 24.7%-38.0%) presented with incomplete or atypical phenotypes. In 37 of 106 families (34.9%; 95% CI, 25.9%-44.8%), the proband was the only participant presenting with a myopathic phenotype, while only 20 families (18.9%; 95% CI, 11.9%-27.6%) had a member with autosomal dominant FSHD.

Conclusions And Relevance: This study found large phenotypic variability associated with individuals carrying a DRA with 7 to 8 RUs, in contrast to the indication that a positive molecular test is the only determining aspect for FSHD diagnosis. These findings suggest that carriers of a DRA with 7 to 8 RUs constitute a genetic subgroup different from classic FSHD. Based on these results, it is recommended that clinicians use the Comprehensive Clinical Evaluation Form for clinical classification and, whenever possible, study the extended family to provide the most adequate clinical management and genetic counseling.
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http://dx.doi.org/10.1001/jamanetworkopen.2020.4040DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7195625PMC
May 2020

Interpretation of the Epigenetic Signature of Facioscapulohumeral Muscular Dystrophy in Light of Genotype-Phenotype Studies.

Int J Mol Sci 2020 Apr 10;21(7). Epub 2020 Apr 10.

Department of Biomedical, Metabolic and Neural Sciences, University of Modena and Reggio Emilia, 41125 Modena, Italy.

Facioscapulohumeral muscular dystrophy (FSHD) is characterized by incomplete penetrance and intra-familial clinical variability. The disease has been associated with the genetic and epigenetic features of the D4Z4 repetitive elements at 4q35. Recently, D4Z4 hypomethylation has been proposed as a reliable marker in the FSHD diagnosis. We exploited the Italian Registry for FSHD, in which FSHD families are classified using the Clinical Comprehensive Evaluation Form (CCEF). A total of 122 index cases showing a classical FSHD phenotype (CCEF, category A) and 110 relatives were selected to test with the receiver operating characteristic (ROC) curve, the diagnostic and predictive value of D4Z4 methylation. Moreover, we performed DNA methylation analysis in selected large families with reduced penetrance characterized by the co-presence of subjects carriers of one D4Z4 reduced allele with no signs of disease or presenting the classic FSHD clinical phenotype. We observed a wide variability in the D4Z4 methylation levels among index cases revealing no association with clinical manifestation or disease severity. By extending the analysis to family members, we revealed the low predictive value of D4Z4 methylation in detecting the affected condition. In view of the variability in D4Z4 methylation profiles observed in our large cohort, we conclude that D4Z4 methylation does not mirror the clinical expression of FSHD. We recommend that measurement of this epigenetic mark must be interpreted with caution in clinical practice.
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http://dx.doi.org/10.3390/ijms21072635DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7178248PMC
April 2020

Intrathecal administration of nusinersen in adult and adolescent patients with spinal muscular atrophy and scoliosis: Transforaminal versus conventional approach.

Neuromuscul Disord 2019 10 23;29(10):742-746. Epub 2019 Aug 23.

Department of Neuroscience "Rita Levi Montalcini, University of Turin, Via Cherasco 15, 10126, Turin, Italy. Electronic address:

Spinal deformities and surgical correction of scoliosis can make intrathecal delivery of nusinersen very challenging. We aim to evaluate the feasibility and safety of intrathecal administration of nusinersen either via interlaminar or transforaminal approach in a cohort of adult and adolescent patients with spinal muscular atrophy (SMA). Twelve patients were treated with nusinersen in our center under CT-guidance; after a CT scan of the lumbar column, we identified a safe virtual trajectory for the needle and defined patients to address to the transforaminal approach (seven patients) or the interlaminar approach (five patients). Out of 47 procedures, all injections but one were successful. There was one adverse event (post-lumbar puncture syndrome) in the interlaminar approach group (out of 20 procedures) and four adverse events in TFA group (out of 27 procedures) including one serious adverse event, a subarachnoid hemorrhage that required hospitalization. Transforaminal approach can be considered an effective option for nusinersen administration but potentially associated with serious complications, therefore it should be recommended in very selected patients.
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http://dx.doi.org/10.1016/j.nmd.2019.08.007DOI Listing
October 2019

New pharmacotherapies for genetic neuromuscular disorders: opportunities and challenges.

Expert Rev Clin Pharmacol 2019 Aug 2;12(8):757-770. Epub 2019 Jul 2.

c Department of Neuroscience "Rita Levi Montalcini"; Hospital Città della Salute e della Scienza, University of Turin , Turin , Italy.

: Genetic neuromuscular diseases (NMDs) constitute a heterogeneous group of rare conditions, including some of the most disabling conditions in childhood. Recently, advanced technologies have greatly expanded preclinical and clinical research, and specific therapies have been developed. : We provide an overview of novel pharmacological approaches to the main NMDs, including Duchenne muscular dystrophy (DMD), spina muscular atrophy (SMA), X-linked myotubular myopathy, Pompe disease (PD), and myotonic dystrophy type 1, with attention to both achievements and unresolved therapeutic challenges. We conducted a selected review of relevant publications in the last five years identified through PubMed and Scopus. Additional information was derived from the website of clinicaltrials.gov and from the authors' direct knowledge of research activities. : For the first time, targeted therapies have received conditional regulatory approval and have been introduced into clinical care: enzyme replacement therapy for PD, gene expression modulation for DMD and SMA, and gene therapy for SMA. Though not curative, these treatments can improve functioning and increase survival. Issues still to be addressed include: early recognition, definition of new emerging phenotypes, development of more sensitive outcome measures, long-term risk-benefit estimates, high costs sustainability, and criteria for therapy initiation and discontinuation.
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http://dx.doi.org/10.1080/17512433.2019.1634543DOI Listing
August 2019

Response to a letter to the editor.

Neurobiol Aging 2019 06 1;78:195-196. Epub 2019 Mar 1.

Department of Neuroscience "Rita Levi Montalcini", University of Torino, Torino, Italy; Neurology 1, Department of Neuroscience and Mental Health, AOU Città della Salute e della Scienza di Torino, Torino, Italy.

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http://dx.doi.org/10.1016/j.neurobiolaging.2019.02.015DOI Listing
June 2019

Assessing the Role of Anti rh-GAA in Modulating Response to ERT in a Late-Onset Pompe Disease Cohort from the Italian GSDII Study Group.

Adv Ther 2019 05 16;36(5):1177-1189. Epub 2019 Mar 16.

Department of Clinical and Experimental Medicine, UOC di Neurologia e Malattie Neuromuscolari, University of Messina, Messina, Italy.

Introduction: Patients with late-onset Pompe disease (LOPD) receiving enzyme replacement therapy (ERT) may develop IgG antibodies against alglucosidase alpha (anti-rhGAA) in the first 3 months of treatment. The exact role of these antibodies in modulating efficacy of ERT in this group of patients is still not fully understood. To assess whether anti rh-GAA antibodies interfere with ERT efficacy, we studied a large Italian cohort of LOPD patients.

Methods: We analyzed clinical findings and performed serial measurements of IgG anti rh-GAA antibody titers from 64 LOPD patients treated with ERT. The first examination (T0) was completed on average at 17.56 months after starting ERT, while the follow-up (T1) was collected on average at 38.5 months. Differences in T0-T1 delta of the six-minute walking test (6MWT), MRC sum score (MRC), gait, stairs and chair performance (GSGC) and forced vital capacity (FVC) were considered and then related to the antibody titers.

Results: Almost 22% of the patients never developed antibodies against GAA, while 78.1% had a positive titer (31.2% patients developed a low titer, 43.8% a medium titer and 3.1% a high titer). No statistical significance was found in relating the T0-T1 delta differences and antibody titers, except for MRC sum score values in a subgroup of patients treated < 36 months, in which those with a null antibody titer showed a greater clinical improvement than patients with a positive titer.

Conclusion: Our results confirm that in a large cohort of LOPD patients, anti rh-GAA antibody generation did not significantly affect either clinical outcome or ERT efficacy. However, in the first 36 months of treatment, a possible interference of low-medium antibody titers with the clinical status could be present. Therefore, a careful and regular evaluation of antibody titers, especially in cases with evidence of clinical decline despite ERT, should be performed.
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http://dx.doi.org/10.1007/s12325-019-00926-5DOI Listing
May 2019

Lipomatosis Incidence and Characteristics in an Italian Cohort of Mitochondrial Patients.

Front Neurol 2019 27;10:160. Epub 2019 Feb 27.

Department of Clinical and Experimental Medicine, UOC Neurologia e Malattie Neuromuscolari, University of Messina, Messina, Italy.

Lipomas have often been associated with mtDNA mutations and were mainly observed in patients with mutation in mitochondrial tRNAlysine which is also the most frequent mutation associated with MERRF. Up to date, no systematic studies have been developed in order to assess the incidence of lipomas in large cohorts of mitochondrial patients.The aim of this study is to analyze the incidence and characteristics of lipomas among an Italian cohort of patients with mitochondrial diseases. A retrospective, database-based study (Nation-wide Italian Collaborative Network of Mitochondrial Diseases) of patients with lipomas was performed. A total of 22 (1.7%) patients with lipomas have been identified among the 1,300 mitochondrial patients, enrolled in the Italian database. In about 18% multiple systemic lipomatosis (MSL) was the only clinical manifestation; 54% of patients showed a classical MERRF syndrome. Myopathy, alone or in association with other symptoms, was found in 27% of patients. Lactate was elevated in all the 12 patients in which was measured. Muscle biopsy was available in 18/22 patients: in all of them mitochondrial abnormalities were present. Eighty six percent had mutations in mtDNA coding for tRNA lysine. In most of patients, lipomas were localized along the cervical-cranial-thoracic region. In 68% of the patients were distributed symmetrically. Only two patients had lipomas in a single anatomical site (1 in right arm and 1 in gluteus maximum). MSL is often overlooked by clinicians in patients with mitochondrial diseases where the clinical picture could be dominated by a severe multi-systemic involvement. Our data confirmed that MSL is a rare sign of mitochondrial disease with a strong association between multiple lipomas and lysine tRNA mutations. MSL could be considered, even if rare, a red flag for mitochondrial disorders, even in patients with an apparently isolated MSL.
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http://dx.doi.org/10.3389/fneur.2019.00160DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6402385PMC
February 2019

Muscle pain in mitochondrial diseases: a picture from the Italian network.

J Neurol 2019 Apr 2;266(4):953-959. Epub 2019 Feb 2.

Neurological Clinic, University of Pisa, Pisa, Italy.

Muscle pain may be part of many neuromuscular disorders including myopathies, peripheral neuropathies and lower motor neuron diseases. Although it has been reported also in mitochondrial diseases (MD), no extensive studies in this group of diseases have been performed so far. We reviewed clinical data from 1398 patients affected with mitochondrial diseases listed in the database of the "Nation-wide Italian Collaborative Network of Mitochondrial Diseases", to assess muscle pain and its features. Muscle pain was present in 164 patients (11.7%). It was commonly observed in subjects with chronic progressive external ophthalmoplegia (cPEO) and with primary myopathy without cPEO, but also-although less frequently-in multisystem phenotypes such as MELAS, MERFF, Kearns Sayre syndrome, NARP, MNGIE and Leigh syndrome. Patients mainly complain of diffuse exercise-related muscle pain, but focal/multifocal and at rest myalgia were often also reported. Muscle pain was more commonly detected in patients with mitochondrial DNA mutations (67.8%) than with nuclear DNA changes (32.2%). Only 34% of the patients showed a good response to drug therapy. Interestingly, patients with nuclear DNA mutations tend to have a better therapeutic response than patients with mtDNA mutations. Muscle pain is present in a significant number of patients with MD, being one of the most common symptoms. Although patients with a myopathic phenotype are more prone to develop muscle pain, this is also observed in patients with a multi system involvement, representing an important and disabling symptom having poor response to current therapy.
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http://dx.doi.org/10.1007/s00415-019-09219-xDOI Listing
April 2019

Longitudinal evaluation of SMN levels as biomarker for spinal muscular atrophy: results of a phase IIb double-blind study of salbutamol.

J Med Genet 2019 05 28;56(5):293-300. Epub 2018 Dec 28.

Muscle Pathology and Neuroimmunology Unit, Neurological Institute Carlo Besta, Milano, Italy.

Background: Spinal muscular atrophy (SMA) is an autosomal recessive neuromuscular disorder, due to the loss of function of the survival motor neuron () gene. The first treatment for the condition, recently approved, is based on the reduction of exon 7 skipping in mRNAs produced by a highly homologous gene (). The primary objective of the present study was to evaluate the applicability of the dosage of SMN gene produts in blood, as biomarker for SMA, and the safety of oral salbutamol, a beta2-adrenergic agonist modulating levels.

Methods: We have performed a 1-year multicentre, double-blind, placebo-controlled study with salbutamol in 45 adult patients with SMA. Patients assumed 4 mg of salbutamol or placebo/three times a day. Molecular tests were copy number, SMN transcript and protein levels. We have also explored the clinical effect, by the outcome measures available at the time of study design.

Results: Thirty-six patients completed the study. Salbutamol was safe and well tolerated. We observed a significant and progressive increase in full-length levels in peripheral blood of the salbutamol-treated patients (p<0.00001). The exploratory analysis of motor function showed an improvement in most patients.

Conclusions: Our data demonstrate safety and molecular efficacy of salbutamol. We provide the first longitudinal evaluation of SMN levels (both transcripts and protein) in placebo and in response to a compound modulating the gene expression: transcript dosage in peripheral blood is reliable and may be used as pharmacodynamic marker in clinical trials with systemic compounds modifying levels.

Trial Registration Number: EudraCT no. 2007-001088-32.
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http://dx.doi.org/10.1136/jmedgenet-2018-105482DOI Listing
May 2019

Copy Number Variants Account for a Tiny Fraction of Undiagnosed Myopathic Patients.

Genes (Basel) 2018 Oct 26;9(11). Epub 2018 Oct 26.

Dipartimento di Medicina di Precisione, Università degli Studi della Campania "Luigi Vanvitelli", 80138 Napoli, Italy.

Next-generation sequencing (NGS) technologies have led to an increase in the diagnosis of heterogeneous genetic conditions. However, over 50% of patients with a genetically inherited disease are still without a diagnosis. In these cases, different hypotheses are usually postulated, including variants in novel genes or elusive mutations. Although the impact of copy number variants (CNVs) in neuromuscular disorders has been largely ignored to date, missed CNVs are predicted to have a major role in disease causation as some very large genes, such as the dystrophin gene, have prone-to-deletion regions. Since muscle tissues express several large disease genes, the presence of elusive CNVs needs to be comprehensively assessed following an accurate and systematic approach. In this multicenter cohort study, we analyzed 234 undiagnosed myopathy patients using a custom array comparative genomic hybridization (CGH) that covers all muscle disease genes at high resolution. Twenty-two patients (9.4%) showed non-polymorphic CNVs. In 12 patients (5.1%), the identified CNVs were considered responsible for the observed phenotype. An additional ten patients (4.3%) presented candidate CNVs not yet proven to be causative. Our study indicates that deletions and duplications may account for 5⁻9% of genetically unsolved patients. This strongly suggests that other mechanisms of disease are yet to be discovered.
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http://dx.doi.org/10.3390/genes9110524DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6267442PMC
October 2018

microRNAs as biomarkers in Pompe disease.

Genet Med 2019 03 12;21(3):591-600. Epub 2018 Jul 12.

Department of Translational Medical Sciences, Federico II University, Naples, Italy.

Purpose: We studied microRNAs as potential biomarkers for Pompe disease.

Methods: We analyzed microRNA expression by small RNA-seq in tissues from the disease murine model at two different ages (3 and 9 months), and in plasma from Pompe patients.

Results: In the mouse model we found 211 microRNAs that were differentially expressed in gastrocnemii and 66 in heart, with a different pattern of expression at different ages. In a preliminary analysis in plasma from six patients 55 microRNAs were differentially expressed. Sixteen of these microRNAs were common to those dysregulated in mouse tissues. These microRNAs are known to modulate the expression of genes involved in relevant pathways for Pompe disease pathophysiology (autophagy, muscle regeneration, muscle atrophy). One of these microRNAs, miR-133a, was selected for further quantitative real-time polymerase chain reaction analysis in plasma samples from 52 patients, obtained from seven Italian and Dutch biobanks. miR-133a levels were significantly higher in Pompe disease patients than in controls and correlated with phenotype severity, with higher levels in infantile compared with late-onset patients. In three infantile patients miR-133a decreased after start of enzyme replacement therapy and evidence of clinical improvement.

Conclusion: Circulating microRNAs may represent additional biomarkers of Pompe disease severity and of response to therapy.
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http://dx.doi.org/10.1038/s41436-018-0103-8DOI Listing
March 2019

The analysis of myotonia congenita mutations discloses functional clusters of amino acids within the CBS2 domain and the C-terminal peptide of the ClC-1 channel.

Hum Mutat 2018 09 4;39(9):1273-1283. Epub 2018 Jul 4.

Department of Biomedical Sciences and Human Oncology, University of Bari Aldo Moro, Polyclinic, Bari, Italy.

Myotonia congenita (MC) is a skeletal-muscle hyperexcitability disorder caused by loss-of-function mutations in the ClC-1 chloride channel. Mutations are scattered over the entire sequence of the channel protein, with more than 30 mutations located in the poorly characterized cytosolic C-terminal domain. In this study, we characterized, through patch clamp, seven ClC-1 mutations identified in patients affected by MC of various severities and located in the C-terminal region. The p.Val829Met, p.Thr832Ile, p.Val851Met, p.Gly859Val, and p.Leu861Pro mutations reside in the CBS2 domain, while p.Pro883Thr and p.Val947Glu are in the C-terminal peptide. We showed that the functional properties of mutant channels correlated with the clinical phenotypes of affected individuals. In addition, we defined clusters of ClC-1 mutations within CBS2 and C-terminal peptide subdomains that share the same functional defect: mutations between 829 and 835 residues and in residue 883 induced an alteration of voltage dependence, mutations between 851 and 859 residues, and in residue 947 induced a reduction of chloride currents, whereas mutations on 861 residue showed no obvious change in ClC-1 function. This study improves our understanding of the mechanisms underlying MC, sheds light on the role of the C-terminal region in ClC-1 function, and provides information to develop new antimyotonic drugs.
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http://dx.doi.org/10.1002/humu.23581DOI Listing
September 2018

Differential diagnosis of vacuolar muscle biopsies: use of p62, LC3 and LAMP2 immunohistochemistry.

Acta Myol 2017 Dec 1;36(4):191-198. Epub 2017 Dec 1.

Center for Neuromuscular Diseases "Paolo Peirolo", Department of Neuroscience "Rita Levi Montalcini", University of Turin, Italy.

Intrafibral vacuoles are the morphological hallmark in a wide variety of human skeletal muscle disorders with different etiology. In most cases, differential diagnosis is feasible with a routine histochemical work up of muscle biopsy. Ultrastructural analysis is an important confirmatory tool, but it is not widely available. Immunohistochemical stainings for p62, LAMP2 and LC3 are commonly available as tissutal marker for autophagy. We compared the immunohistochemical patterns for autophagic markers p62, LC3 and LAMP2 with routine histochemical markers in 39 biopsies from patients with definite diagnoses of glycogen storage disease type 2 (LOPD or Pompe disease, PD), sporadic inclusion body myositis (sIBM), oculo-pharyngeal muscular dystrophy (OPMD) and necrotizing myopathy (NM). Moreover, we also analyzed muscles of 10 normal controls. In PD group, LC3 and LAMP2 showed an higher percentage of positive fibers, whereas p62 was limited to a minority of fibers, thus paralleling the results of histochemical stainings; in NM group, LAMP2 and LC-3 were diffusely and unspecifically expressed in necrotic fibers, with p62 significantly expressed only in two cases. OPMD biopsies did not reveal any significant positivity. The most interesting results were observed in sIBM group, where p62 was expressed in all cases, even in fibers without other markers positivity. There results, although limited to a small number of cases, suggest that the contemporary use of p62, LAMP2 and LC-3 staining may have an adjunctive role in characterizing muscle fiber vacuoles, revealing autophagic pathway activation and providing further clues for the understanding of pathogenetic mechanisms.s.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5953231PMC
December 2017

Elevated TGF β2 serum levels in Emery-Dreifuss Muscular Dystrophy: Implications for myocyte and tenocyte differentiation and fibrogenic processes.

Nucleus 2018 01;9(1):292-304

j Institute of Molecular Genetics (IGM)-CNR, Unit of Bologna , Bologna , Italy.

Among rare diseases caused by mutations in LMNA gene, Emery-Dreifuss Muscular Dystrophy type 2 and Limb-Girdle muscular Dystrophy 1B are characterized by muscle weakness and wasting, joint contractures, cardiomyopathy with conduction system disorders. Circulating biomarkers for these pathologies have not been identified. Here, we analyzed the secretome of a cohort of patients affected by these muscular laminopathies in the attempt to identify a common signature. Multiplex cytokine assay showed that transforming growth factor beta 2 (TGF β2) and interleukin 17 serum levels are consistently elevated in the vast majority of examined patients, while interleukin 6 and basic fibroblast growth factor are altered in subgroups of patients. Levels of TGF β2 are also increased in fibroblast and myoblast cultures established from patient biopsies as well as in serum from mice bearing the H222P Lmna mutation causing Emery-Dreifuss Muscular Dystrophy in humans. Both patient serum and fibroblast conditioned media activated a TGF β2-dependent fibrogenic program in normal human myoblasts and tenocytes and inhibited myoblast differentiation. Consistent with these results, a TGF β2 neutralizing antibody avoided fibrogenic marker activation and myogenesis impairment. Cell intrinsic TGF β2-dependent mechanisms were also determined in laminopathic cells, where TGF β2 activated AKT/mTOR phosphorylation. These data show that TGF β2 contributes to the pathogenesis of Emery-Dreifuss Muscular Dystrophy type 2 and Limb-Girdle muscular Dystrophy 1B and can be considered a potential biomarker of those diseases. Further, the evidence of TGF β2 pathogenetic effects in tenocytes provides the first mechanistic insight into occurrence of joint contractures in muscular laminopathies.
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http://dx.doi.org/10.1080/19491034.2018.1467722DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5973167PMC
January 2018

Mutation analysis of CHCHD2 and CHCHD10 in Italian patients with mitochondrial myopathy.

Neurobiol Aging 2018 06 14;66:181.e1-181.e2. Epub 2018 Feb 14.

Department of Neuroscience "Rita Levi Montalcini", University of Torino, Torino, Italy; Neurology 1, Department of Neuroscience and Mental Health, AOU Cittá della Salute e della Scienza di Torino, Torino, Italy.

Mutations in CHCHD2 and CHCHD10 were recently reported in a broad spectrum of neurodegenerative diseases, for example, Parkinson's disease, amyotrophic lateral sclerosis, frontotemporal dementia, or mitochondrial myopathy (MM). The aim of the study was to evaluate the prevalence of CHCHD2 and CHCHD10 mutations in Italian MM patients without mitochondrial DNA mutations. The coding regions of CHCHD2 and CHCHD10 were sequenced in 62 MM patients. None of the patients showed CHCHD2 mutations, whereas 1 sporadic MM patient carried a homozygous Pro96Thr substitution in CHCHD10. Muscle biopsy of this patient showed intracellular glycogen accumulation with cytochrome c oxidase negative and ragged red fibers. Our study suggests that the homozygous Pro96Thr mutation in CHCHD10 might be pathogenic but does not support a major role for CHCHD2 in MM pathogenesis.
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http://dx.doi.org/10.1016/j.neurobiolaging.2018.02.007DOI Listing
June 2018

Interpreting Genetic Variants in Titin in Patients With Muscle Disorders.

JAMA Neurol 2018 05;75(5):557-565

Dipartimento di Biochimica, Biofisica e Patologia Generale, Università degli Studi della Campania "Luigi Vanvitelli," Napoli, Italy.

Importance: Mutations in the titin gene (TTN) cause a wide spectrum of genetic diseases. The interpretation of the numerous rare variants identified in TTN is a difficult challenge given its large size.

Objective: To identify genetic variants in titin in a cohort of patients with muscle disorders.

Design, Setting, And Participants: In this case series, 9 patients with titinopathy and 4 other patients with possibly disease-causing variants in TTN were identified. Titin mutations were detected through targeted resequencing performed on DNA from 504 patients with muscular dystrophy, congenital myopathy, or other skeletal muscle disorders. Patients were enrolled from 10 clinical centers in April 2012 to December 2013. All of them had not received a diagnosis after undergoing an extensive investigation, including Sanger sequencing of candidate genes. The data analysis was performed between September 2013 and January 2017. Sequencing data were analyzed using an internal custom bioinformatics pipeline.

Main Outcomes And Measures: The identification of novel mutations in the TTN gene and novel patients with titinopathy. We performed an evaluation of putative causative variants in the TTN gene, combining genetic, clinical, and imaging data with messenger RNA and/or protein studies.

Results: Of the 9 novel patients with titinopathy, 5 (55.5%) were men and the mean (SD) age at onset was 25 (15.8) years (range, 0-46 years). Of the 4 other patients (3 men and 1 woman) with possibly disease-causing TTN variants, 2 (50%) had a congenital myopathy and 2 (50%) had a slowly progressive distal myopathy with onset in the second decade. Most of the identified mutations were previously unreported. However, all the variants, even the already described mutations, require careful clinical and molecular evaluation of probands and relatives. Heterozygous truncating variants or unique missense changes are not sufficient to make a diagnosis of titinopathy.

Conclusions And Relevance: The interpretation of TTN variants often requires further analyses, including a comprehensive evaluation of the clinical phenotype (deep phenotyping) as well as messenger RNA and protein studies. We propose a specific workflow for the clinical interpretation of genetic findings in titin.
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http://dx.doi.org/10.1001/jamaneurol.2017.4899DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5885217PMC
May 2018

The genetic basis of undiagnosed muscular dystrophies and myopathies: Results from 504 patients.

Neurology 2016 07 8;87(1):71-6. Epub 2016 Jun 8.

From the Dipartimento di Biochimica Biofisica e Patologia Generale (M.S., G.D.F., A. Torella, A.G., T.G., F.D.V.B., G.E., G.P., V.N.), Seconda Università di Napoli; Telethon Institute of Genetics and Medicine (M.S., G.D.F., A. Torella, M. Mutarelli, V.S.M., A.G., T.G., G.E., V.N.), Pozzuoli; U.O.C. Neurologia Pediatrica e Malattie Muscolari (C.F., C.M., C.B.), IRCCS Istituto Giannina Gaslini, Genova; Centro Dino Ferrari, Dipartimento di Fisiopatologia Medico-Chirurgica e dei Trapianti (F.M., D.R., G.P.C.), and Neuromuscular and Rare Disease Unit, Dipartimento di Neuroscienze (M. Moggio), Università degli Studi di Milano, Fondazione IRCCS Ca' Granda, Ospedale Maggiore Policlinico, Milan; Dipartimento di Neuroscienze (M. Fanin, E.P.), Università di Padova; Dipartimento di Neuroscienze e Scienze Riproduttive ed Odontostomatologiche (L.R., L.S.), Università degli Studi di Napoli "Federico II," Napoli; Dipartimento di Medicina Clinica e Sperimentale (G.R., G.S.), Università degli Studi di Pisa; Medicina Molecolare (G.A., F.M.S.), IRCCS Fondazione Stella Maris, Pisa; Dipartimento di Medicina Sperimentale (L. Passamano, P.D., R.P., L. Politano) and Dipartimento di Scienze Mediche, Chirurgiche, Neurologiche, Metaboliche, e dell'Invecchiamento (O.F., S.S., G.D.I.), Seconda Università di Napoli; Dipartimento di Neuroscienze (A.R., M. Mora, L.M.), Istituto Besta, Milano; Don Carlo Gnocchi ONLUS Foundation (G.T.), Milano; Dipartimento di Neuroscienze (A.D., E.B.), IRCCS Ospedale Pediatrico Bambino Gesù, Roma, Italy; Center for Medical Genetics (S.J., K.C.) and Department of Neurology (J.D.B.), Ghent University Hospital, Belgium; Dipartimento di Neuroscienze (O.M., C.R., S.M., A. Toscano), Università degli Studi di Messina, Italy; Folkhälsan Institute of Genetics (A.E., P.H., B.U.), University of Helsinki, Finland; Section for Neuromuscular Diseases and Neuropathies (M. Filosto), Unit of Clinical Neurology, University Hospital 'Spedali Civili,' Brescia, Italy; Neuromusc

Objective: To apply next-generation sequencing (NGS) for the investigation of the genetic basis of undiagnosed muscular dystrophies and myopathies in a very large cohort of patients.

Methods: We applied an NGS-based platform named MotorPlex to our diagnostic workflow to test muscle disease genes with a high sensitivity and specificity for small DNA variants. We analyzed 504 undiagnosed patients mostly referred as being affected by limb-girdle muscular dystrophy or congenital myopathy.

Results: MotorPlex provided a complete molecular diagnosis in 218 cases (43.3%). A further 160 patients (31.7%) showed as yet unproven candidate variants. Pathogenic variants were found in 47 of 93 genes, and in more than 30% of cases, the phenotype was nonconventional, broadening the spectrum of disease presentation in at least 10 genes.

Conclusions: Our large DNA study of patients with undiagnosed myopathy is an example of the ongoing revolution in molecular diagnostics, highlighting the advantages in using NGS as a first-tier approach for heterogeneous genetic conditions.
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http://dx.doi.org/10.1212/WNL.0000000000002800DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4932234PMC
July 2016

A novel clinical tool to classify facioscapulohumeral muscular dystrophy phenotypes.

J Neurol 2016 Jun 28;263(6):1204-14. Epub 2016 Apr 28.

Department of Life Sciences, University of Modena and Reggio Emilia, Modena, Italy.

Based on the 7-year experience of the Italian Clinical Network for FSHD, we revised the FSHD clinical form to describe, in a harmonized manner, the phenotypic spectrum observed in FSHD. The new Comprehensive Clinical Evaluation Form (CCEF) defines various clinical categories by the combination of different features. The inter-rater reproducibility of the CCEF was assessed between two examiners using kappa statistics by evaluating 56 subjects carrying the molecular marker used for FSHD diagnosis. The CCEF classifies: (1) subjects presenting facial and scapular girdle muscle weakness typical of FSHD (category A, subcategories A1-A3), (2) subjects with muscle weakness limited to scapular girdle or facial muscles (category B subcategories B1, B2), (3) asymptomatic/healthy subjects (category C, subcategories C1, C2), (4) subjects with myopathic phenotype presenting clinical features not consistent with FSHD canonical phenotype (D, subcategories D1, D2). The inter-rater reliability study showed an excellent concordance of the final four CCEF categories with a κ equal to 0.90; 95 % CI (0.71; 0.97). Absolute agreement was observed for categories C and D, an excellent agreement for categories A [κ = 0.88; 95 % CI (0.75; 1.00)], and a good agreement for categories B [κ = 0.79; 95 % CI (0.57; 1.00)]. The CCEF supports the harmonized phenotypic classification of patients and families. The categories outlined by the CCEF may assist diagnosis, genetic counseling and natural history studies. Furthermore, the CCEF categories could support selection of patients in randomized clinical trials. This precise categorization might also promote the search of genetic factor(s) contributing to the phenotypic spectrum of disease.
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http://dx.doi.org/10.1007/s00415-016-8123-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4893383PMC
June 2016

"Mitochondrial neuropathies": A survey from the large cohort of the Italian Network.

Neuromuscul Disord 2016 Apr-May;26(4-5):272-6. Epub 2016 Feb 23.

Neurological Clinic, University of Pisa, Pisa, Italy.

Involvement of the peripheral nervous system in mitochondrial disorders has been previously reported. However, the prevalence of peripheral neuropathy in mitochondrial disorders is still unclear. Based on the large database of the "Nation-wide Italian Collaborative Network of Mitochondrial Diseases", we reviewed the clinical data of 1200 patients, with special regard to peripheral neuropathy (mean age at onset 24.3 ± 20.1 years; age at last evaluation 39.8 ± 22.3 years; females 52.7%; childhood onset [before age 16 years] 43.1%). Peripheral neuropathy was present in 143/1156 patients (12.4%), being one of the ten most common signs and symptoms. POLG mutations cause a potentially painful, axonal/mixed, mainly sensory polyneuropathy; TYMP mutations lead to a demyelinating sensory-motor polyneuropathy; SURF1 mutations are associated with a demyelinating/mixed sensory-motor polyneuropathy. The only mtDNA mutation consistently associated with peripheral neuropathy (although less severely than in the above-considered nuclear genes) was the m.8993T > G (or the rarer T > C) changes, which lead to an axonal, mainly sensory polyneuropathy. In conclusion, peripheral neuropathy is one of the most common features of a mitochondrial disorder, and may negatively impact on the quality of life of these patients. Furthermore, the presence or absence of peripheral neuropathy, as well as its specific forms and the association with neuropathic pain (indicative of a POLG-associated disease) can guide the molecular analysis.
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http://dx.doi.org/10.1016/j.nmd.2016.02.008DOI Listing
January 2017

Clinical expression of facioscapulohumeral muscular dystrophy in carriers of 1-3 D4Z4 reduced alleles: experience of the FSHD Italian National Registry.

BMJ Open 2016 Jan 5;6(1):e007798. Epub 2016 Jan 5.

Department of Science of Life, Institute of Biology, University of Modena and Reggio Emilia, Modena, Italy Department of Molecular, Cell and Cancer Biology, University of Massachusetts Medical School, Worcester, Massachusetts, USA.

Objectives: Facioscapulohumeral muscular dystrophy type 1 (FSHD1) has been genetically linked to reduced numbers (≤ 8) of D4Z4 repeats at 4q35. Particularly severe FSHD cases, characterised by an infantile onset and presence of additional extra-muscular features, have been associated with the shortest D4Z4 reduced alleles with 1-3 repeats (1-3 DRA). We searched for signs of perinatal onset and evaluated disease outcome through the systematic collection of clinical and anamnestic records of de novo and familial index cases and their relatives, carrying 1-3 DRA.

Setting: Italy.

Participants: 66 index cases and 33 relatives carrying 1-3 DRA.

Outcomes: The clinical examination was performed using the standardised FSHD evaluation form with validated inter-rater reliability. To investigate the earliest signs of disease, we designed the Infantile Anamnestic Questionnaire (IAQ). Comparison of age at onset was performed using the non-parametric Wilcoxon rank-sum or Kruskal-Wallis test. Comparison of the FSHD score was performed using a general linear model and Wald test. Kaplan-Meier survival analysis was used to estimate the age-specific cumulative motor impairment risk.

Results: No patients had perinatal onset. Among index cases, 36 (54.5%) showed the first signs by 10 years of age. The large majority of patients with early disease onset (26 out of 36, 72.2%) were de novo; whereas the majority of patients with disease onset after 10 years of age were familial (16, 53.3%). Comparison of the disease severity outcome between index cases with age at onset before and over 10 years of age, failed to detect statistical significance (Wald test p value=0.064). Of 61 index cases, only 17 (27.9%) presented extra-muscular conditions. Relatives carrying 1-3 DRA showed a large clinical variability ranging from healthy subjects, to patients with severe motor impairment.

Conclusions: The size of the D4Z4 allele is not always predictive of severe clinical outcome. The high degree of clinical variability suggests that additional factors contribute to the phenotype complexity.
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http://dx.doi.org/10.1136/bmjopen-2015-007798DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4716236PMC
January 2016

Redefining phenotypes associated with mitochondrial DNA single deletion.

J Neurol 2015 May 26;262(5):1301-9. Epub 2015 Mar 26.

Neurological Clinic, University of Pisa, Via Roma 67, 56126, Pisa, Italy,

Progressive external ophthalmoplegia (PEO), Kearns-Sayre syndrome (KSS) and Pearson syndrome are the three sporadic clinical syndromes classically associated with single large-scale deletions of mitochondrial DNA (mtDNA). PEO plus is a term frequently utilized in the clinical setting to identify patients with PEO and some degree of multisystem involvement, but a precise definition is not available. The purpose of the present study is to better define the clinical phenotypes associated with a single mtDNA deletion, by a retrospective study on a large cohort of 228 patients from the database of the "Nation-wide Italian Collaborative Network of Mitochondrial Diseases". In our database, single deletions account for about a third of all patients with mtDNA-related disease, more than previously recognized. We elaborated new criteria for the definition of PEO and "KSS spectrum" (a category of which classic KSS represents the most severe extreme). The criteria for "KSS spectrum" include the resulting multisystem clinical features associated with the KSS features, and which therefore can predict their presence or subsequent development. With the new criteria, we were able to classify nearly all our single-deletion patients: 64.5% PEO, 31.6% KSS spectrum (including classic KSS 6.6%) and 2.6% Pearson syndrome. The deletion length was greater in KSS spectrum than in PEO, whereas heteroplasmy was inversely related with age at onset. We believe that the new phenotype definitions implemented here may contribute to a more homogeneous patient categorization, which will be useful in future cohort studies of natural history and clinical trials.
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http://dx.doi.org/10.1007/s00415-015-7710-yDOI Listing
May 2015

Early Recognition of Late-Onset Pompe Disease by Skeletal Muscle Signs and Symptoms.

J Neuromuscul Dis 2015;2(s1):S2

Neuromuscular Unit, Department of Neurosciences, Hospital Città della Salute e della Scienza of Torino, University of Torino, Italy.

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January 2015

LMNA-associated myopathies: the Italian experience in a large cohort of patients.

Neurology 2014 Oct 1;83(18):1634-44. Epub 2014 Oct 1.

From the Department of Neurology IV (L. Maggi, G.B., D.K., P.B., L.C., R.M., L. Morandi), Neuromuscular Diseases and Neuroimmunology, Fondazione IRCCS Istituto Neurologico Carlo Besta, Milan; Unit of Neuromuscular and Neurodegenerative Disorders (A.D., L.T., E.B.), Department of Neurosciences, IRCCS Bambino Gesù Children's Hospital, Rome; Child Neurology Unit (A.P.), IRCCS Institute of Neurological Sciences, Bologna; Department of Paediatric Neurology (S. Sivo, M.P., E.M.), Catholic University, Rome; Department of Clinical and Experimental Medicine (G.R., G.S.), Section of Neurology, University of Pisa; Department of Neuroscience Rita Levi Montalcini (L.V., T.M.), University of Torino; Cardiomyology and Medical Genetics (P.D., L.P.), Second Naples University; Arrhythmia Unit and Electrophysiology Laboratories (S. Sala), Institute of Experimental Neurology (Inspe) and Department of Neurology (M.S., S.C.P.) and Genomic Unit for the Diagnosis of Human Pathologies, Center for Translational Genomics and Bioinformatics (M.F.), San Raffaele Scientific Institute, Milan; Vita-Salute San Raffaele University (M.F.), Milan; Laboratory of Molecular Biology (M.F., S.B.), Diagnostica e Ricerca San Raffaele, Milan; Department of Neurosciences (E.P.), University of Padova, Padua; Department of Neurosciences (A.T., C.R.), University of Messina; National Research Council of Italy (G.L.), Institute of Molecular Genetics Unit of Bologna and Laboratory of Musculoskeletal Cell Biology IOR, Bologna; and Department of Public Health, Clinical and Molecular Medicine (N.C.), University of Cagliari, Italy.

Objectives: Our aim was to conduct a comparative study in a large cohort of myopathic patients carrying LMNA gene mutations to evaluate clinical and molecular features associated with different phenotypes.

Methods: We performed a retrospective cohort study of 78 myopathic patients with LMNA mutation and 30 familial cases with LMNA mutation without muscle involvement. We analyzed features characterizing the various forms of LMNA-related myopathy through correlation statistics.

Results: Of the 78 patients, 37 (47%) had limb-girdle muscular dystrophy 1B (LGMD1B), 18 (23%) congenital muscular dystrophy (MDCL), 17 (22%) autosomal dominant Emery-Dreifuss muscular dystrophy 2 (EDMD2), and 6 (8%) an atypical myopathy. The myopathic phenotypes shared a similar cardiac impairment. Cardioverter defibrillator or pacemaker was implanted in 41 (53%) myopathic patients compared to 7 (23%) familial cases without muscle involvement (p = 0.005). Heart transplantation was performed in 8 (10.3%) myopathic patients and in none of the familial cases. Ten (12.8%) myopathic patients died; there were no deaths among the familial cases (p = 0.032). Missense mutations were found in 14 patients (82%) with EDMD2 and 14 patients (78%) with MDCL compared to 17 patients (45%) with LGMD1B and 4 (67%) atypical patients. Frameshift mutations were detected in 17 (45%) LGMD1B compared to 3 (18%) EDMD2, 1 (6%) MDCL, and 2 (33%) with atypical myopathy (p = 0.021). Furthermore, frameshift mutations were found in 30 of 73 patients (41%) with heart involvement compared to 4 of 35 (11%) without heart involvement (p = 0.004).

Conclusions: Our data provided new insights in LMNA-related myopathies, whose natural history appears to be dominated by cardiac involvement and related complications.
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http://dx.doi.org/10.1212/WNL.0000000000000934DOI Listing
October 2014

Myoclonus in mitochondrial disorders.

Mov Disord 2014 May 7;29(6):722-8. Epub 2014 Feb 7.

Neurological Clinic, University of Pisa, Pisa, Italy.

Myoclonus is a possible manifestation of mitochondrial disorders, and its presence is considered, in association with epilepsy and the ragged red fibers, pivotal for the syndromic diagnosis of MERRF (myoclonic epilepsy with ragged red fibers). However, its prevalence in mitochondrial diseases is not known. The aims of this study are the evaluation of the prevalence of myoclonus in a big cohort of mitochondrial patients and the clinical characterization of these subjects. Based on the database of the "Nation-wide Italian Collaborative Network of Mitochondrial Diseases," we reviewed the clinical and molecular data of mitochondrial patients with myoclonus among their clinical features. Myoclonus is a rather uncommon clinical feature of mitochondrial diseases (3.6% of 1,086 patients registered in our database). It is not strictly linked to a specific genotype or phenotype, and only 1 of 3 patients with MERRF harbors the 8344A>G mutation (frequently labeled as "the MERRF mutation"). Finally, myoclonus is not inextricably linked to epilepsy in MERRF patients, but more to cerebellar ataxia. In a myoclonic patient, evidences of mitochondrial dysfunction must be investigated, even though myoclonus is not a common sign of mitochondriopathy. Clinical, histological, and biochemical data may predict the finding of a mitochondrial or nuclear DNA mutation. Finally, this study reinforces the notion that myoclonus is not inextricably linked to epilepsy in MERRF patients, and therefore the term "myoclonic epilepsy" seems inadequate and potentially misleading.
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http://dx.doi.org/10.1002/mds.25839DOI Listing
May 2014

The m.3243A>G mitochondrial DNA mutation and related phenotypes. A matter of gender?

J Neurol 2014 Mar 29;261(3):504-10. Epub 2013 Dec 29.

Neurological Clinic, University of Pisa, Via Roma 67, 56100, Pisa, Italy,

The m.3243A>G "MELAS" (mitochondrial encephalopathy with lactic acidosis and stroke-like episodes) mutation is one of the most common point mutations of the mitochondrial DNA, but its phenotypic variability is incompletely understood. The aim of this study was to revise the phenotypic spectrum associated with the mitochondrial m.3243A>G mutation in 126 Italian carriers of the mutation, by a retrospective, database-based study ("Nation-wide Italian Collaborative Network of Mitochondrial Diseases"). Our results confirmed the high clinical heterogeneity of the m.3243A>G mutation. Hearing loss and diabetes were the most frequent clinical features, followed by stroke-like episodes. "MIDD" (maternally-inherited diabetes and deafness) and "PEO" (progressive external ophthalmoplegia) are nosographic terms without any real prognostic value, because these patients may be even more prone to the development of multisystem complications such as stroke-like episodes and heart involvement. The "MELAS" acronym is convincing and useful to denote patients with histological, biochemical and/or molecular evidence of mitochondrial disease who experience stroke-like episodes. Of note, we observed for the first time that male gender could represent a risk factor for the development of stroke-like episodes in Italian m.3243A>G carriers. Gender effect is not a new concept in mitochondrial medicine, but it has never been observed in MELAS. A better elucidation of the complex network linking mitochondrial dysfunction, apoptosis, estrogen effects and stroke-like episodes may hold therapeutic promises.
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http://dx.doi.org/10.1007/s00415-013-7225-3DOI Listing
March 2014