Maria Barbara Pasanisi

Scopus Publications

“Natural history of skeletal muscle laminopathies: a 2-year prospective study”
Luca Spiro Santovito, Silvia Bonanno, Maria Barbara Pasanisi, Annamaria Gallone, Federica Ricci, Irene Tramacere, Riccardo Zanin, Stefano Carlo Previtali, and Lorenzo Maggi
Elsevier BV

Serum miR-34a-5p, miR-103a-3p, and miR-376a-3p as possible biomarkers of conversion from relapsing-remitting to secondary progressive multiple sclerosis
Simone Agostini, Roberta Mancuso, Lorenzo Agostino Citterio, Domenico Caputo, Letizia Oreni, Riccardo Nuzzi, Maria Barbara Pasanisi, Marco Rovaris, and Mario Clerici
Elsevier BV

Myelin Basic Protein in Oligodendrocyte-Derived Extracellular Vesicles as a Diagnostic and Prognostic Biomarker in Multiple Sclerosis: A Pilot Study
Cristina Agliardi, Franca Rosa Guerini, Milena Zanzottera, Elisabetta Bolognesi, Silvia Picciolini, Domenico Caputo, Marco Rovaris, Maria Barbara Pasanisi, and Mario Clerici
MDPI AG
Approximately 15% of multiple sclerosis (MS) patients develop a progressive form of disease from onset; this condition (primary progressive-PP) MS is difficult to diagnose and treat, and is associated with a poor prognosis. Extracellular vesicles (EVs) of brain origin isolated from blood and their protein cargoes could function as a biomarker of pathological conditions. We verified whether MBP and MOG content in oligodendrocytes-derived EVs (ODEVs) could be biomarkers of MS and could help in the differential diagnosis of clinical MS phenotypes. A total of 136 individuals (7 clinically isolated syndrome (CIS), 18 PPMS, 49 relapsing remitting (RRMS)) and 70 matched healthy controls (HC) were enrolled. ODEVs were enriched from serum by immune-capture with anti-MOG antibody; MBP and MOG protein cargoes were measured by ELISA. MBP concentration in ODEVs was significantly increased in CIS (p < 0.001), RRMS (p < 0.001) and PPMS (p < 0.001) compared to HC and was correlated with disease severity measured by EDSS and MSSS. Notably, MBP concentration in ODEVs was also significantly augmented in PPMS compared to RRMS (p = 0.004) and CIS (p = 0.03). Logistic regression and ROC analyses confirmed these results. A minimally invasive blood test measuring the concentration of MBP in ODEVs is a promising tool that could facilitate MS diagnosis.

Two Single Nucleotide Polymorphisms in the Purinergic Receptor P2X7 Gene Are Associated with Disease Severity in Multiple Sclerosis
Franca Rosa Guerini, Cristina Agliardi, Elisabetta Bolognesi, Milena Zanzottera, Domenico Caputo, Maria Barbara Pasanisi, Marco Rovaris, and Mario Clerici
MDPI AG
Multiple sclerosis (MS) is a chronic autoimmune inflammatory disease of the central nervous system (CNS) that leads to progressive physical disability. Recent evidence has suggested that P2X7 receptor (P2X7R)-mediated purinergic signalling pathways play a role in MS-associated neuroinflammation, possibly contributing to disease pathogenesis. To evaluate possible associations between P2X7R polymorphisms and MS disease severity, we performed an association study of five non-synonymous SNPs coding variants of the P2X7R gene: rs1718119 Ala348Thr, rs2230911 Thr357Ser, rs2230912 Gln460Arg, rs3751143 Glu496Ala, and rs28360457 Arg307Gln, modulating P2X7R expression in 128 MS patients (relapsing remitting MS, RRMS: n = 94; secondary progressive, SPMS: n = 34). All patients were genotyped, and multiple sclerosis severity score (MSSS) was evaluated in every case; 189 healthy subjects were enrolled as well as controls. Results showed that P2X7R rs1718119(A) 348Thr and rs22390912(G) 464Arg, two SNPs of minor allele frequency (MAF) known to confer gain of function to the P2X7R protein, were associated with significantly higher MSSS in RRMS patients alone (SMRR (p < 0.001, p = 0.01, respectively)). Interestingly, two whole haplotypes resulted in having significant association with MSSS in these same patients. Thus: (1) the P2X7R-4 “ACGAG” haplotype, characterized by the co-presence of the rs1718119-rs2230912 AG MAF alleles, was associated with higher MSSS (Beta: 1.11 p = 0.04), and (2) the P2X7R-1 “GCAAG” complementary haplotype, which contains the rs1718119 and rs2230912 GA wild-type alleles, was more frequently carried by patients with lower MSSS and less severe disease (Beta: −1.54 p < 0.001). Although being preliminary and needing confirmation in an ampler cohort, these results suggest that 348Thr and 464Arg variants have a role as modulators of disease severity in RRMS patients.

Dietary habits, nutritional status and risk of a first demyelinating event: an incident case-control study in a southern European cohort
Paola Cavalla, Paola Golzio, Daniela Maietta, Chiara Bosa, Maria Barbara Pasanisi, Anastasia Alteno, Valentina Schillaci, Gianfranco Costantini, Paola Durelli, Erica Cuffini,et al.
Springer Science and Business Media LLC

Acute and chronic synaptic pathology in multiple sclerosis gray matter
Marco Vercellino, Stella Marasciulo, Silvia Grifoni, Elena Vallino-Costassa, Chiara Bosa, Maria Barbara Pasanisi, Paola Crociara, Cristina Casalone, Adriano Chiò, Maria Teresa Giordana,et al.
SAGE Publications
Objectives: To investigate the extent of synaptic loss, and the contribution of gray matter (GM) inflammation and demyelination to synaptic loss, in multiple sclerosis (MS) brain tissue. Methods: This study was performed on two different post-mortem series of MS and control brains, including deep GM and cortical GM. MS brain samples had been specifically selected for the presence of active demyelinating GM lesions. Over 1,000,000 individual synapses were identified and counted using confocal microscopy, and further characterized as glutamatergic/GABAergic. Synaptic counts were also correlated with neuronal/axonal loss. Results: Important synaptic loss was observed in active demyelinating GM lesions (−58.9%), while in chronic inactive GM lesions, synaptic density was only mildly reduced compared to adjacent non-lesional gray matter (NLGM) (−12.6%). Synaptic loss equally affected glutamatergic and GABAergic synapses. Diffuse synaptic loss was observed in MS NLGM compared to control GM (−21.2% overall). Conclusion: This study provides evidence, in MS brain tissue, of acute synaptic damage/loss during active GM inflammatory demyelination and of synaptic reorganization in chronically demyelinated GM, affecting equally glutamatergic and GABAergic synapses. Furthermore, this study provides a strong indication of widespread synaptic loss in MS NLGM also independently from focal GM demyelination.

Eculizumab in refractory generalized myasthenia gravis previously treated with rituximab: subgroup analysis of REGAIN and its extension study
Zaeem A. Siddiqi, Richard J. Nowak, Tahseen Mozaffar, Fanny O'Brien, Marcus Yountz, Francesco Patti, and
Wiley
AbstractIntroduction/AimsIndividuals with refractory generalized myasthenia gravis (gMG) who have a history of rituximab use and experience persistent symptoms represent a population with unmet treatment needs. The aim of this analysis was to evaluate the efficacy and safety of eculizumab in patients with refractory anti‐acetylcholine receptor antibody‐positive (AChR+) gMG previously treated with rituximab.MethodsThis post hoc subgroup analysis of the phase 3 REGAIN study (NCT01997229) and its open‐label extension (OLE; NCT02301624) compared baseline characteristics, safety, and response to eculizumab in participants who had previously received rituximab with those who had not. Rituximab use was not permitted within the 6 months before screening or during REGAIN/OLE.ResultsOf 125 REGAIN participants, 14 had received rituximab previously (7 received placebo and 7 received eculizumab). In the previous‐rituximab group, 57% had used at least four other immunosuppressants compared with 16% in the no‐previous‐rituximab group. Myasthenia Gravis Activities of Daily Living total scores from eculizumab baseline to week 130 of eculizumab treatment improved in both the previous‐rituximab and no‐previous‐rituximab groups (least‐squares mean −4.4, standard error of the mean [SEM] 1.0 [n = 9] and least‐squares mean −4.6, SEM 0.3 [n = 67], respectively; difference = 0.2, 95% confidence interval −1.88 to 2.22). In addition, in both groups, most patients who were treated with eculizumab for 130 weeks achieved a Myasthenia Gravis Foundation of America post‐intervention status of minimal manifestations (66.7% and 65.0%, respectively). The eculizumab safety profile was similar between groups and consistent with its established profile.DiscussionEculizumab is an effective therapy for patients with refractory AChR+ gMG, irrespective of whether they had received rituximab treatment previously.

SMA-miRs (MiR-181a- 5p, -324-5p, and -451a) are overexpressed in spinal muscular atrophy skeletal muscle and serum samples
Emanuela Abiusi, Paola Infante, Cinzia Cagnoli, Ludovica Lospinoso Severini, Marika Pane, Giorgia Coratti, Maria Carmela Pera, Adele D'Amico, Federica Diano, Agnese Novelli,et al.
eLife Sciences Publications, Ltd
Background:Spinal muscular atrophy (SMA) is a neuromuscular disorder characterized by the degeneration of the second motor neuron. The phenotype ranges from very severe to very mild forms. All patients have the homozygous loss of the SMN1 gene and a variable number of SMN2 (generally 2–4 copies), inversely related to the severity. The amazing results of the available treatments have made compelling the need of prognostic biomarkers to predict the progression trajectories of patients. Besides the SMN2 products, few other biomarkers have been evaluated so far, including some miRs.Methods:We performed whole miRNome analysis of muscle samples of patients and controls (14 biopsies and 9 cultures). The levels of muscle differentially expressed miRs were evaluated in serum samples (51 patients and 37 controls) and integrated with SMN2 copies, SMN2 full-length transcript levels in blood and age (SMA-score).Results:Over 100 miRs were differentially expressed in SMA muscle; 3 of them (hsa-miR-181a-5p, -324-5p, -451a; SMA-miRs) were significantly upregulated in the serum of patients. The severity predicted by the SMA-score was related to that of the clinical classification at a correlation coefficient of 0.87 (p<10-5).Conclusions:miRNome analyses suggest the primary involvement of skeletal muscle in SMA pathogenesis. The SMA-miRs are likely actively released in the blood flow; their function and target cells require to be elucidated. The accuracy of the SMA-score needs to be verified in replicative studies: if confirmed, its use could be crucial for the routine prognostic assessment, also in presymptomatic patients.Funding:Telethon Italia (grant #GGP12116).

Quantitative Muscle MRI Protocol as Possible Biomarker in Becker Muscular Dystrophy
Lorenzo Maggi, Marco Moscatelli, Rita Frangiamore, Federica Mazzi, Mattia Verri, Alberto De Luca, Maria Barbara Pasanisi, Giovanni Baranello, Irene Tramacere, Luisa Chiapparini,et al.
Springer Science and Business Media LLC

Post-intervention Status in Patients With Refractory Myasthenia Gravis Treated With Eculizumab During REGAIN and Its Open-Label Extension
Renato Mantegazza, Gil I. Wolfe, Srikanth Muppidi, Heinz Wiendl, Kenji P. Fujita, Fanny L. O'Brien, Heather D.E. Booth, James F. Howard, Claudio Gabriel Mazia, Miguel Wilken,et al.
Ovid Technologies (Wolters Kluwer Health)
Objective To evaluate whether eculizumab helps patients with anti–acetylcholine receptor–positive (AChR+) refractory generalized myasthenia gravis (gMG) achieve the Myasthenia Gravis Foundation of America (MGFA) post-intervention status of minimal manifestations (MM), we assessed patients' status throughout REGAIN (Safety and Efficacy of Eculizumab in AChR+ Refractory Generalized Myasthenia Gravis) and its open-label extension. Methods Patients who completed the REGAIN randomized controlled trial and continued into the open-label extension were included in this tertiary endpoint analysis. Patients were assessed for the MGFA post-intervention status of improved, unchanged, worse, MM, and pharmacologic remission at defined time points during REGAIN and through week 130 of the open-label study. Results A total of 117 patients completed REGAIN and continued into the open-label study (eculizumab/eculizumab: 56; placebo/eculizumab: 61). At week 26 of REGAIN, more eculizumab-treated patients than placebo-treated patients achieved a status of improved (60.7% vs 41.7%) or MM (25.0% vs 13.3%; common OR: 2.3; 95% CI: 1.1–4.5). After 130 weeks of eculizumab treatment, 88.0% of patients achieved improved status and 57.3% of patients achieved MM status. The safety profile of eculizumab was consistent with its known profile and no new safety signals were detected. Conclusion Eculizumab led to rapid and sustained achievement of MM in patients with AChR+ refractory gMG. These findings support the use of eculizumab in this previously difficult-to-treat patient population. ClinicalTrials.gov Identifier REGAIN, NCT01997229; REGAIN open-label extension, NCT02301624. Classification of Evidence This study provides Class II evidence that, after 26 weeks of eculizumab treatment, 25.0% of adults with AChR+ refractory gMG achieved MM, compared with 13.3% who received placebo.

Consistent improvement with eculizumab across muscle groups in myasthenia gravis
Renato Mantegazza, Fanny L. O'Brien, Marcus Yountz, James F. Howard, and
Wiley
AbstractObjectiveTo assess whether eculizumab, a terminal complement inhibitor, improves patient‐ and physician‐reported outcomes (evaluated using the myasthenia gravis activities of daily living profile and the quantitative myasthenia gravis scale, respectively) in patients with refractory anti‐acetylcholine receptor antibody‐positive generalized myasthenia gravis across four domains, representing ocular, bulbar, respiratory, and limb/gross motor muscle groups.MethodsPatients with refractory anti‐acetylcholine receptor antibody‐positive generalized myasthenia gravis were randomized 1:1 to receive either placebo or eculizumab during the REGAIN study (NCT01997229). Patients who completed REGAIN were eligible to continue into the open‐label extension trial (NCT02301624) for up to 4 years. The four domain scores of each of the myasthenia gravis activities of daily living profile and the quantitative myasthenia gravis scale recorded throughout REGAIN and through 130 weeks of the open‐label extension were analyzed.ResultsOf the 125 patients who participated in REGAIN, 117 enrolled in the open‐label extension; 61 had received placebo and 56 had received eculizumab during REGAIN. Patients experienced rapid improvements in total scores and all four domain scores of both the myasthenia gravis activities of daily living profile and the quantitative myasthenia gravis scale with eculizumab treatment. These improvements were sustained through 130 weeks of the open‐label extension.InterpretationEculizumab treatment elicits rapid and sustained improvements in muscle strength across ocular, bulbar, respiratory, and limb/gross motor muscle groups and in associated daily activities in patients with refractory anti‐acetylcholine receptor antibody‐positive generalized myasthenia gravis.

‘Minimal symptom expression’ in patients with acetylcholine receptor antibody-positive refractory generalized myasthenia gravis treated with eculizumab
, John Vissing, Saiju Jacob, Kenji P. Fujita, Fanny O’Brien, and James F. Howard
Springer Science and Business Media LLC
Abstract Background The efficacy and tolerability of eculizumab were assessed in REGAIN, a 26-week, phase 3, randomized, double-blind, placebo-controlled study in anti-acetylcholine receptor antibody-positive (AChR+) refractory generalized myasthenia gravis (gMG), and its open-label extension. Methods Attainment of ‘minimal symptom expression’ was evaluated using patient-reported outcome measures of gMG symptoms [MG activities of daily living scale (MG-ADL), 15-item MG quality of life questionnaire (MG-QOL15)] at the completion of REGAIN and during the open-label extension. ‘Minimal symptom expression’ was defined as MG-ADL total score of 0–1 or MG-QOL15 total score of 0–3. Results At REGAIN week 26, more eculizumab-treated patients achieved ‘minimal symptom expression’ versus placebo [MG-ADL: 21.4% vs 1.7%; difference 19.8%; 95% confidence interval (CI) 8.5, 31.0; p = 0.0007; MG-QOL15: 16.1% vs 1.7%; difference 14.4%; 95% CI 4.3, 24.6; p = 0.0069]. During the open-label extension, the proportion of patients in the placebo/eculizumab group who achieved ‘minimal symptom expression’ increased after initiating eculizumab treatment and was sustained through 130 weeks of open-label eculizumab (MG-ADL: 1.7 to 27.8%; MG-QOL15: 1.7 to 19.4%). At extension study week 130, similar proportions of patients in the eculizumab/eculizumab and placebo/eculizumab groups achieved ‘minimal symptom expression’ (MG-ADL: 22.9% and 27.8%, respectively, p = 0.7861; MG-QOL15: 14.3% and 19.4%, respectively, p = 0.7531). The long-term tolerability of eculizumab was consistent with previous reports. Conclusions Patients with AChR+ refractory gMG who receive eculizumab can achieve sustained ‘minimal symptom expression’ based on patient-reported outcomes. ‘Minimal symptom expression’ may be a useful tool in measuring therapy effectiveness in gMG. Trial registration ClinicalTrials.gov NCT01997229, NCT02301624.

Exercise training alone or in combination with high-protein diet in patients with late onset Pompe disease: Results of a cross over study
Annalisa Sechi, Lucrezia Zuccarelli, Bruno Grassi, Rita Frangiamore, Ramona De Amicis, Mauro Marzorati, Simone Porcelli, Annarita Tullio, Anna Bacco, Simona Bertoli,et al.
Springer Science and Business Media LLC
Abstract Background Late onset Pompe disease (LOPD) is a lysosomal neuromuscular disorder which can progressively impair the patients’ exercise tolerance, motor and respiratory functions, and quality of life. The available enzyme replacement therapy (ERT) does not completely counteract disease progression. We investigated the effect of exercise training alone, or associated with a high-protein diet, on the exercise tolerance, muscle and pulmonary functions, and quality of life of LOPD patients on long term ERT. Methods The patients were asked to participate to a crossover randomized study comprehending a control period (free diet, no exercise) followed by 2 intervention periods: exercise or exercise + diet, each lasting 26 weeks and separated by 13 weeks washout periods. Exercise training included moderate-intensity aerobic exercise on a cycle ergometer, stretching and balance exercises, strength training. The diet was composed by 25–30% protein, 30–35% carbohydrate and 35–40% fat. Before and after each period patients were assessed for: exercise tolerance test on a cycle-ergometer, serum muscle enzymes, pulmonary function tests and SF36 questionnaire for quality of life. Compliance was evaluated by training and dietary diaries. Patients were contacted weekly by researchers to optimize adherence to treatments. Results Thirteen LOPD patients, median age 49 ± 11 years, under chronic ERT (median 6.0 ± 4.0 years) were recruited. Peak aerobic power (peak pulmonary O2 uptake) decreased after control, whereas it increased after exercise, and more markedlyafter exercise + diet. Serum levels of lactate dehydrogenase (LDH) significantly decreased after exercise + diet; both creatine kinase (CK) and LDH levels were significantly reduced after exercise + diet compared to exercise. Pulmonary function showed no changes after control and exercise, whereas a significant improvement of forced expiratory volume in 1 sec (FEV1) was observed after exercise + diet. SF36 showed a slight improvement in the “mental component” scale after exercise, and a significant improvement in “general health” and “vitality” after exercise + diet. The compliance to prescriptions was higher than 70% for both diet and exercise. Conclusions Exercise tolerance (as evaluated by peak aerobic power) showed a tendency to decrease in LOPD patients on long term ERT. Exercise training, particularly if combined with high-protein diet, could reverse this decrease and result in an improvement, which was accompanied by improved quality of life. The association of the two lifestyle interventions resulted also in a reduction of muscle enzyme levels and improved pulmonary function.

Kappa free light chains index in the differential diagnosis of Multiple Sclerosis from Neuromyelitis optica spectrum disorders and other immune-mediated central nervous system disorders
P. Cavalla, P. Caropreso, S. Limoncelli, C. Bosa, M.B. Pasanisi, V. Schillaci, A. Alteno, G. Costantini, M.T. Giordana, G. Mengozzi,et al.
Elsevier BV

Correction to: Eculizumab improves fatigue in refractory generalized myasthenia gravis (Quality of Life Research, (2019), 28, 8, (2247-2254), 10.1007/s11136-019-02148-2)
, Henning Andersen, Renato Mantegazza, Jing Jing Wang, Fanny O’Brien, Kaushik Patra, and James F. Howard
Springer Science and Business Media LLC

Eculizumab improves fatigue in refractory generalized myasthenia gravis
Henning Andersen, Renato Mantegazza, Jing Jing Wang, Fanny O’Brien, Kaushik Patra, James F. Howard, , Claudio Gabriel Mazia, Miguel Wilken, Fabio Barroso,et al.
Springer Science and Business Media LLC
Abstract Purpose To evaluate the effect of eculizumab on perceived fatigue in patients with anti-acetylcholine receptor antibody-positive, refractory, generalized myasthenia gravis (MG) using the Quality of Life in Neurological Disorders (Neuro-QOL) Fatigue subscale, and to evaluate correlations between improvements in Neuro-QOL Fatigue and other clinical endpoints. Methods Neuro-QOL Fatigue, MG Activities of Daily Living (MG-ADL), Quantitative MG (QMG), and the 15-item MG Quality of Life (MG-QOL15) scales were administered during the phase 3, randomized, placebo-controlled REGAIN study (eculizumab, n = 62; placebo, n = 63) and subsequent open-label extension (OLE). Data were analyzed using repeated-measures models. Correlations between changes in Neuro-QOL Fatigue and in MG-ADL, QMG, and MG-QOL15 scores were determined at REGAIN week 26. Results At REGAIN week 26, eculizumab-treated patients showed significantly greater improvements in Neuro-QOL Fatigue scores than placebo-treated patients (consistent with improvements in MG-ADL, QMG, and MG-QOL15 scores previously reported in REGAIN). Improvements with eculizumab were sustained through OLE week 52. Correlations between Neuro-QOL Fatigue and MG-QOL15, MG-ADL, and QMG scores were strong for eculizumab-treated patients at REGAIN week 26, and strong, moderate, and weak, respectively, for placebo-treated patients. Conclusions Compared with placebo, eculizumab was associated with improvements in perceived fatigue that strongly correlated with improvements in MG-specific outcome measures. Trial ID Registration: NCT01997229, NCT02301624.

Long-term safety and efficacy of eculizumab in generalized myasthenia gravis
Srikanth Muppidi, Kimiaki Utsugisawa, Michael Benatar, Hiroyuki Murai, Richard J. Barohn, Isabel Illa, Saiju Jacob, John Vissing, Ted M. Burns, John T. Kissel,et al.
Wiley
ABSTRACTIntroduction: Eculizumab is effective and well tolerated in patients with antiacetylcholine receptor antibody‐positive refractory generalized myasthenia gravis (gMG; REGAIN; NCT01997229). We report an interim analysis of an open‐label extension of REGAIN, evaluating eculizumab's long‐term safety and efficacy. Methods: Eculizumab (1,200 mg every 2 weeks for 22.7 months [median]) was administered to 117 patients. Results: The safety profile of eculizumab was consistent with REGAIN; no cases of meningococcal infection were reported during the interim analysis period. Myasthenia gravis exacerbation rate was reduced by 75% from the year before REGAIN (P < 0.0001). Improvements with eculizumab in activities of daily living, muscle strength, functional ability, and quality of life in REGAIN were maintained through 3 years; 56% of patients achieved minimal manifestations or pharmacological remission. Patients who had received placebo during REGAIN experienced rapid and sustained improvements during open‐label eculizumab (P < 0.0001). Discussion: These findings provide evidence for the long‐term safety and sustained efficacy of eculizumab for refractory gMG. Muscle Nerve 2019

Aging-associated genes and let-7 microRNAs: a contribution to myogenic program dysregulation in oculopharyngeal muscular dystrophy
Cristina Cappelletti, Barbara Galbardi, Mirella Bruttini, Franco Salerno, Eleonora Canioni, Maria Barbara Pasanisi, Carmelo Rodolico, Teresa Brizzi, Marina Mora, Alessandra Renieri,et al.
Wiley
Oculopharyngeal muscular dystrophy (OPMD) is a late‐onset muscle disease caused by an abnormal (GCN) triplet expansion within the polyadenylate‐binding protein nuclear 1 gene and consequent mRNA processing impairment and myogenic defects. Because a reduced cell proliferation potential and the consequent regeneration failure of aging muscle have been shown to be governed by lethal‐7 (let‐7) microRNA‐mediated mechanisms, in the present study, we evaluated the role of let‐7 in the pathogenesis of OPMD. By a multidisciplinary approach, including conf ocal microscopy, Western blot, and quantitative PCR analyses on muscle biopsies from patients and unaffected individuals, we found a significant increase in let‐7 expression in OPMD muscles associated with an unusual high percentage of paired box 7–positive satellite cells. Furthermore, IL‐6, a cytokine involved in the regulation of satellite cell proliferation and differentiation and a potential target of let‐7, was found strongly down‐regulated in OPMD compared with control muscles. The decrease in IL‐6 transcript levels and protein content was also confirmed in vitro during differentiation of patients' and controls' muscle cells. Overall, our data suggest a key role of let‐7 in the regeneration and degeneration process in OPMD muscle and pointed to IL‐6 as a potential target molecule for new therapeutic approaches for this disorder.—Cappelletti, C., Galbardi, B., Bruttini, M., Salerno, F., Canioni, E., Pasanisi, M. B., Rodolico, C., Brizzi, T., Mora, M., Renieri, A., Maggi, L., Bernasconi, P., Mantegazza, R. Aging‐associated genes and let‐7 microRNAs: a contribution to myogenic program dysregulation in oculopharyngeal muscular dystrophy. FASEB J. 33, 7155–7167 (2019). www.fasebj.org

Longitudinal evaluation of SMN levels as biomarker for spinal muscular atrophy: Results of a phase IIb double-blind study of salbutamol
Francesco Danilo Tiziano, Rosa Lomastro, Emanuela Abiusi, Maria Barbara Pasanisi, Lorena Di Pietro, Stefania Fiori, Giovanni Baranello, Corrado Angelini, Gianni Sorarù, Alessandra Gaiani,et al.
BMJ
BackgroundSpinal muscular atrophy (SMA) is an autosomal recessive neuromuscular disorder, due to the loss of function of the survival motor neuron (SMN1) 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 (SMN2). 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 SMN2 levels.MethodsWe 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 SMN2 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.ResultsThirty-six patients completed the study. Salbutamol was safe and well tolerated. We observed a significant and progressive increase in SMN2 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.ConclusionsOur 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: SMN transcript dosage in peripheral blood is reliable and may be used as pharmacodynamic marker in clinical trials with systemic compounds modifying SMN2levels.Trial registration numberEudraCT no. 2007-001088-32.

Amifampridine phosphate in the treatment of muscle-specific kinase myasthenia gravis: a phase IIb, randomized, double-blind, placebo-controlled, double crossover study
Silvia Bonanno, Maria Barbara Pasanisi, Rita Frangiamore, Lorenzo Maggi, Carlo Antozzi, Francesca Andreetta, Angela Campanella, Greta Brenna, Lorenzo Cottini, and Renato Mantegazza
SAGE Publications
Objective: The aim of this study is to determine the safety and the efficacy of amifampridine phosphate in muscle-specific kinase antibody-positive myasthenia gravis, in a 1:1 randomized, double-blind, placebo-controlled, switchback, double crossover study. Methods: Eligible patients had muscle-specific kinase myasthenia gravis, >18 years of age, and Myasthenia Gravis Foundation of America class II–IV with a score of ⩾9 on Myasthenia Gravis Composite scale. After the run-in phase, during which amifampridine phosphate was titrated to a tolerable and effective dosage, patients were randomized to receive placebo–amifampridine–placebo sequence or amifampridine–placebo–amifampridine sequence daily for 7 days. Then, patients switched treatment arms twice, for a total of 21 days of double-blind treatment. Safety was determined by serial assessments of adverse events/serious adverse events, physical examinations, and clinical and laboratory tests. The co-primary outcome measures included changes from baseline of Quantitative Myasthenia Gravis score and Myasthenia Gravis–specific Activities of Daily Living Profile score. The secondary outcome measures comprised changes from baseline of Myasthenia Gravis Composite score, Myasthenia Gravis Quality of Life scale—15 questions, Fatigue Severity Scale, and Carlo Besta Neurological Institute–Myasthenia Gravis scale. Statistical analyses were assessed using a switchback model for three-period, two-treatment crossover design. Results: A total of 10 patients were screened, enrolled, and treated. Transient paresthesias (60%) were the only amifampridine phosphate–related adverse events reported. Four patients were randomized to receive placebo–amifampridine–placebo sequence and three patients to receive amifampridine–placebo–amifampridine sequence. The co-primary objectives were statistically met (Quantitative Myasthenia Gravis score: p = 0.0003 and Myasthenia Gravis–specific Activities of Daily Living Profile score: p = 0.0006), as well as all the secondary endpoints (Myasthenia Gravis Composite score: p < 0.0001, Myasthenia Gravis Quality of Life scale—15 questions: p = 0.0025, Fatigue Severity Scale: p = 0.0061, and Carlo Besta Neurological Institute–Myasthenia Gravis scale: p = 0.0014). Conclusion: Despite the low number of patients, MuSK-001 study provided evidence that amifampridine phosphate, in the range of 30–60 mg daily dose, was safe and effective in treating muscle-specific kinase myasthenia gravis, suggesting the need for a large multi-center trial to confirm these results.

Interpreting genetic variants in titin in patients with muscle disorders
Marco Savarese, Lorenzo Maggi, Anna Vihola, Per Harald Jonson, Giorgio Tasca, Lucia Ruggiero, Luca Bello, Francesca Magri, Teresa Giugliano, Annalaura Torella,et al.
American Medical Association (AMA)
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.

Diagnosis of Duchenne Muscular Dystrophy in Italy in the last decade: Critical issues and areas for improvements
Adele D'Amico, Michela Catteruccia, Giovanni Baranello, Luisa Politano, Alessandra Govoni, Stefano Carlo Previtali, Marika Pane, Maria Grazia D'Angelo, Claudio Bruno, Sonia Messina,et al.
Elsevier BV

Mutations in INPP5K Cause a Form of Congenital Muscular Dystrophy Overlapping Marinesco-Sjögren Syndrome and Dystroglycanopathy
Daniel P.S. Osborn, Heather L. Pond, Neda Mazaheri, Jeremy Dejardin, Christopher J. Munn, Khaloob Mushref, Edmund S. Cauley, Isabella Moroni, Maria Barbara Pasanisi, Elizabeth A. Sellars,et al.
Elsevier BV

Perceived efficacy of salbutamol by persons with spinal muscular atrophy: A mixed methods study
Ambra M. Giovannetti, Maria Barbara Pasanisi, Milda Černiauskaitė, Chiara Bussolino, Matilde Leonardi, and Lucia Morandi
Wiley
Introduction: The aim of this study was to assess the perceived effect of salbutamol in adult patients with spinal muscular atrophy and to evaluate the usefulness of the World Health Organization Disability Assessment Schedule II (WHODAS II) and Fatigue Severity Scale (FSS) for its measurement. Methods: A longitudinal mixed methods study was performed. Ten patients were interviewed and completed WHODAS II and FSS questionnaires to assess disability and fatigue at 2 time‐points. Inductive thematic analysis was used for qualitative data. The non‐parametric Wilcoxon test was performed for quantitative analysis. Results: All participants reported an improvement in their condition after salbutamol consumption. WHODAS II and FSS reliably captured changes in patients' disability and fatigue. Conclusions: The mixed methods design allowed us to identify the functional domains in which participants experienced effects of salbutamol. Patients were satisfied with the treatment as shown by decreased fatigue, improved functioning, and infrequent side effects. Muscle Nerve, 2016 Muscle Nerve 54: 843–849, 2016

DNAJB6 myopathies: Focused review on an emerging and expanding group of myopathies
Alessandra Ruggieri, Simona Saredi, Simona Zanotti, Maria Barbara Pasanisi, Lorenzo Maggi, and Marina Mora
Frontiers Media SA
Mutations in the DNAJB6 gene have been associated with the autosomal dominant limb girdle muscular dystrophy type 1D (LGMD1D), a disorder characterized by abnormal protein aggregates and rimmed vacuoles in muscle fibers. DNAJB6 is a ubiquitously expressed Hsp40 co-chaperone characterized by a J domain that specifies Hsp70 functions in the cellular environment. DNAJB6 is also a potent inhibitor of expanded polyglutamine (polyQ) aggregation preventing aggregate toxicity in cells. In DNAJB6-mutated patients this anti-aggregation property is significantly reduced, albeit not completely lost. To elucidate the pathogenetic mechanisms underlying the DNAJB6-related myopathy, animal models have been created showing that, indeed, conditional muscular expression of a DNAJB6 mutant in the mouse causes a LGMD1D myofibrillary muscle tissue phenotype. Both mutations and phenotypes reported until recently were rather homogeneous, being exclusively missense mutations of a few amino acids of the protein G/F domain, and with a phenotype characterized by adult-onset slowly progressive muscular dystrophy predominantly affecting proximal muscles. Lately, several novel mutations and new phenotypes of DNAJB6 have been described. These mutations once more affect the G/F domain of DNAJB6 with missense changes and a splice site mutation; and the phenotypes include childhood onset and distal involvement of muscles, or childhood-onset LGMD1D with loss of ambulation in early adulthood and respiratory involvement. Thus, the spectrum of DNAJB6-related phenotypes is widening. Although our knowledge about the role of DNAJB6 in the pathogenesis of muscle diseases has made great progression, several questions remain unsolved, including why a ubiquitous protein affects only, or predominantly, skeletal muscle; why only the G/F domain is involved; and what is the possible role of the DNAJB6a isoform. Clarification of these issues will provide clues to implement possible therapeutic strategies for DNAJB6-related myopathies.

Maria Barbara Pasanisi

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