Full-Length SMN Transcript in Extracellular Vesicles as Biomarker in Individuals with Spinal Muscular Atrophy Type 2 Treated with Nusinersen.

BACKGROUND: Three therapeutic strategies have radically changed the therapeutic scenario for spinal muscular atrophy (SMA). However, therapeutic response differs between individuals. There is a need to identify biomarkers to further assess therapeutic response and to better understand which variables determine the extent of response. METHODS: We conducted a study using an optimized digital droplet PCR-based method for the ultra-sensitive detection of SMN transcript in serum EVs from SMA 2 individuals treated with nusinersen over 14 months. In parallel, we investigated levels of serum and CSF neurofilament heavy chain (pNF-H) in the same cohort. RESULTS: Expression of flSMN transcript in EVs of SMA 2 individuals prior to nusinersen was lower than in controls (0.40 vs 2.79 copies/ul; p < 0.05) and increased after 14 months of nusinersen (0.40 vs 1.11 copies/ul; p < 0.05). The increase in flSMN with nusinersen was significantly higher in younger individuals (p < 0.05). Serum pNF-h was higher in non-treated individuals with SMA 2 than in controls (230.72 vs 22.88 pg/ml; p < 0.05) and decreased with nusinersen (45.72 pg/ml at 6 months, 39.02 pg/ml at 14 months). CSF pNF-h in SMA 2 individuals also decreased with nusinersen (248.04 pg/ml prior to treatment, 197.10 pg/dl at 2 months, 104.43 pg/dl at 6 months, 131.03 pg/dl at 14 months). CONCLUSIONS: We identified an increase of flSMN transcript in serum EVs of SMA 2 individuals treated with nusinersen that was more pronounced in the younger individuals. Our results indicate that flSMN transcript expression in serum EVs is a possible biomarker in SMA to predict or monitor the response to treatment.

Pediatric SMA patients with complex spinal anatomy: Implementation and evaluation of a decision-tree algorithm for administration of nusinersen.

The approval of nusinersen for the treatment of spinal muscular atrophy (SMA) has significantly changed the natural history of the disease. Nevertheless, scoliosis secondary to axial muscle weakness occurs at some point in most of patients with SMA and a conventional posterior interlaminar approach for intrathecal administration of nusinersen can be particularly challenging to perform in patients with severe scoliosis and/or previous spine fusion surgeries. We developed a protocol for the administration of nusinersen in pediatric patients, which includes a decision-tree algorithm that categorizes patients according to the estimated technical difficulty for the intrathecal administration. Complex spine patients were defined as those with a Cobb angle greater than 50° and/or a history of spinal surgery, while the rest of patients were considered non-complex. Nusinersen was successfully administered through a conventional non-CT-guided lumbar puncture in all 14 non-complex spine patients (110 out of 110 procedures; 100%). The feasibility of the intrathecal injection in the 15 complex spine patients was assessed by 3D CT. Administration was considered unfeasible in 7 out of these 15 patients according to imaging. In the 8 complex spine patients in whom the administration was considered feasible, conventional non-CT-guided lumbar punctures were successful only in 19 out of 53 procedures (36%). The remaining 34 procedures (64%) were guided by CT scan, all successful. Our work demonstrates that a cut-off point of 50° in Cobb angle and history of spinal surgery can reliably be used to anticipate the need for CT guidance in nusinersen administration.

Association of Initial Maximal Motor Ability With Long-term Functional Outcome in Patients With COL6-Related Dystrophies.

OBJECTIVE: To accurately categorize the phenotypes of individuals with collagen VI-related dystrophies (COL6-RDs) during the first years of life to predict long-term motor function and pulmonary function, to provide phenotype-specific anticipatory care, and to improve clinical trial readiness. METHODS: This retrospective, multicenter, international study analyzed the relationship of long-term motor and pulmonary function with the initial maximal motor ability achieved in individuals with COL6-RD. RESULTS: We studied 119 patients with COL6-RD from Spain (n = 54) and the United States (n = 65). The early maximal motor milestones of ability to rise from the floor unassisted and ability to climb 4 steps without holding onto a railing demonstrated reliability in distinguishing between 3 COL6-RD phenotypic subgroups: (1) Ullrich congenital muscular dystrophy, (2) intermediate COL6-RD, and (3) Bethlem myopathy. Long-term motor function and pulmonary function are strongly correlated with the maximal motor ability achieved during the first years of life. Maximal motor capacity can predict other disease-relevant events such as the age at loss of ambulation and the need for the initiation of nocturnal noninvasive ventilation. CONCLUSION: This work proposes a prospective phenotypic classification for COL6-RDs that will enable an accurate prediction of a patient's COL6-RD phenotype during the first years of life. The ability to establish a patient's COL6-RD phenotypic classification early will enable a more accurate prognosis of future motor and pulmonary function, thus improving anticipatory clinical care, and it will be instrumental in aiding the design of future clinical trials by allowing early stratification of trial cohorts.

Early and long-term effect of the treatment with pyridostigmine in patients with GMPPB-related congenital myasthenic syndrome.

GMPPB mutations cause congenital myasthenic syndromes (CMS) overlapping with muscular dystrophy. Treatment with pyridostigmine has been reported to be effective in those patients. Nevertheless, results of functional motor assessments to determine its precise impact on the short and long term were not available. We describe the response to treatment with pyridostigmine in three siblings with GMPPB-related CMS using functional motor scales performed regularly over a period of 40 months. The beneficial effect of the treatment was outstanding within the first hours, with all the scales showing a dramatic increase in only two days. This remarkable improvement remained steady during 12 months but a moderate decrease was subsequently detected in two of the three patients. Despite this decline in the scores of the scales at the end of follow up, the functional motor status of the patients was still significantly better than it was before starting treatment. The introduction of pyridostigmine at an early age of the disease in one of the patients, before the onset of scoliosis, may have had a protective effect on it.

Longitudinal Study of Three microRNAs in Duchenne Muscular Dystrophy and Becker Muscular Dystrophy.

Our objective was to investigate the potential of three microRNAs, miR-181a-5p, miR-30c-5p, and miR-206 as prognostic biomarkers for long-term follow up of Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) patients. We analyzed the expression of three microRNAs in serum of 18 patients (DMD 13, BMD 5) and 13 controls using droplet digital PCR. Over 4 years a minimum of two and a maximum of three measurements were performed at different time points in the same patient. Correlations between microRNA serum levels, age, and functional outcome measures were analyzed. We show the individual evolution of the levels of the three microRNAs in 12 patients and also the effect of corticosteroid treatment on microRNAs expression. We measure the expression of three microRNAs in the muscle of six DMD patients and also the expression of target genes for miR-30c. We found that levels of miR-30c and miR-206 remained significantly elevated in DMD patients relative to controls over the entire study length. The introduction of the corticosteroid treatment did not significantly influence the levels of these microRNAs. We report a trend for microRNA levels to decrease with age. Moreover, miR-206 expression levels are capable to distinguish DMD from BMD patients according to ROC analysis. We found miR-30c expression decreased in the muscle of DMD patients and marked upregulation of the target genes for this microRNA. MiR-30c and miR-206 represent sensitive biomarkers for DMD, while miR-206 may have an additional value to distinguish the DMD and BMD phenotype. This may be particularly relevant to assess the effectiveness of treatments aimed at converting the DMD to the less-severe BMD like phenotype.

Differences in Adipose Tissue and Lean Mass Distribution in Patients with Collagen VI Related Myopathies Are Associated with Disease Severity and Physical Ability.

Mutations in human collagen VI genes cause a spectrum of musculoskeletal conditions in children and adults collectively termed collagen VI-related myopathies (COL6-RM) characterized by a varying degree of muscle weakness and joint contractures and which include Ullrich Congenital Muscular Dystrophy (UCMD) and Bethlem Myopathy (BM). Given that collagen VI is one of the most abundant extracellular matrix proteins in adipose tissue and its emerging role in energy metabolism we hypothesized that collagen VI deficiency might be associated with alterations in adipose tissue distribution and adipokines serum profile. We analyzed body composition by means of dual-energy X-ray absorptiometry in 30 pediatric and adult COL6-RM myopathy patients representing a range of severities (UCMD, intermediate-COL6-RM, and BM). We found a distinctive pattern of regional adipose tissue accumulation which was more evident in children at the most severe end of the spectrum. In particular, the accumulation of fat in the android region was a distinguishing feature of UCMD patients. In parallel, there was a decrease in lean mass compatible with a state of sarcopenia, particularly in ambulant children with an intermediate phenotype. All children and adult patients that were sarcopenic were also obese. These changes were significantly more pronounced in children with collagen VI deficiency than in children with Duchenne Muscular Dystrophy of the same ambulatory status. High molecular weight adiponectin and leptin were significantly increased in sera from children in the intermediate and BM group. Correlation analysis showed that the parameters of fat mass were negatively associated with motor function according to several validated outcome measures. In contrast, lean mass parameters correlated positively with physical performance and quality of life. Leptin and adiponectin circulating levels correlated positively with fat mass parameters and negatively with lean mass and thus may be relevant to the disease pathogenesis and as circulating markers. Taken together our results indicate that COL6-RM are characterized by specific changes in total fat mass and distribution which associate with disease severity, motor function, and quality of life and which are clinically meaningful and thus should be taken into consideration in the management of these patients.

Digital PCR quantification of miR-30c and miR-181a as serum biomarkers for Duchenne muscular dystrophy.

Circulating microRNAs (miRs/miRNAs) are being used as non-invasive biomarkers for diagnosis, prognosis and efficiency of clinical trials. However, to exploit their potential it is necessary to improve and standardize their detection. In a previous study, we identified two microRNAs, miR-30c and miR-181a, that appear to be key regulators of muscular dystrophy. We hypothesized that they could represent useful biomarkers of Duchenne and Becker muscular dystrophies (DMD and BMD). The objective of this study was to assess the absolute levels of miR-30c and miR-181a in sera of DMD and BMD patients using digital PCR (a robust technique for precise and direct quantification of small amounts of nucleic acids without standard curves and external references), and investigate the correlation between miR-30c and miR-181a expressions and several clinical parameters. Our results show that the serum levels of miR-30c and miR-181a increased 7- and 6-fold respectively in DMD patients (n = 21, 2-14 years, ambulant), and 7-fold in BMD patients (n = 5, 9-15 years) compared to controls (n = 22, 2-14 years). No association between miRNA levels and age or corticosteroid treatment was detected in DMD. However, there was a trend towards higher levels of miR-30c in DMD patients with better preserved motor function according to various motor scales and timed tests. We demonstrate that digital PCR is a useful technique for accurate absolute quantification of microRNAs in sera of DMD/BMD patients. We propose miR-30c and miR-181a as reliable serum diagnostic biomarkers for DMD and BMD and miR-30c as a potential novel biomarker to assess disease severity in DMD.

[Fractures in spinal muscular atrophy]. / Fracturas en la atrofia muscular espinal.

AIM: To determine the frequency of fractures in patients with spinal muscular atrophy, their mechanism of production, age at appearance and functional repercussions. PATIENTS AND METHODS: Sixty-five patients with spinal muscular atrophy were studied. Cases of fractures diagnosed by means of X-rays were collected and the following parameters were analysed: type of spinal muscular atrophy, gait, age at the time the fracture occurred, mechanism of production, location, treatment applied and functional repercussion. RESULTS: Thirteen patients (20%) presented a total of 20 fractures (four of them presented two or more fractures). The mean age was 6.35 years. The fractures were mostly located in the femur and the mechanism of production was falls in 12 cases and minor traumatic injury in eight. No vertebral fractures were detected. All of them were treated conservatively. The only patient with a fracture who was able to walk stopped walking after immobilisation. CONCLUSIONS: The existence of fractures in these patients interferes with their quality of life and their level of functioning. It is important to prevent them from occurring during management of the patient and by ensuring a correct posture in the wheelchair with the use of restraint systems. Further studies are needed on the loss of bone mineral density in these patients and their possible relationship with fractures.

TITLE: Fracturas en la atrofia muscular espinal.Objetivo. Determinar la frecuencia de fracturas en pacientes con atrofia muscular espinal, mecanismo de produccion, edad de aparicion y repercusion funcional. Pacientes y metodos. Se estudian 65 pacientes con atrofia muscular espinal. Se recogen las fracturas diagnosticadas mediante radiografia y se analizan los siguientes parametros: tipo de atrofia muscular espinal, marcha, edad en el momento de la fractura, mecanismo de produccion, localizacion, tratamiento aplicado y repercusion funcional. Resultados. Presentaron fracturas 13 pacientes (20%), con un total de 20 (cuatro presentaron dos o mas fracturas). La edad media fue de 6,35 años. La localizacion fue en su mayoria en el femur y el mecanismo de produccion, en 12 casos por caidas y en 8 por traumatismo menor. No detectamos ninguna fractura vertebral. Todas se trataron de manera conservadora. El unico paciente ambulante que presento una fractura dejo de caminar despues de la inmovilizacion. Conclusiones. La existencia de fracturas en estos pacientes interfiere en su calidad de vida y en el nivel funcional. Es importante la prevencion de las mismas en el manejo del paciente y vigilando la correcta postura en la silla de ruedas con sistemas de sujecion Deberian emprenderse mas estudios sobre la perdida de densidad mineral osea en estos pacientes y su posible relacion con las fracturas.

Pituitary dysfunction after traumatic brain injury in children: is there a need for ongoing endocrine assessment?

BACKGROUND: Hypopituitarism has been widely described in adults after traumatic brain injury (TBI); however, the available data in paediatric populations are scarce. Here, we report the results of a prospective, long-term study in children, adolescents and young adults. STUDY GROUP: Thirty-seven children (age, 2 months to 19·9 years) of 51 eligible patients were followed for 1 year. Clinical and baseline endocrine variables were assessed in all 3 and 12 months after TBI; children ≥ 6 years underwent two stimulation tests (glucagon stimulation and megatest). RESULTS: In the group ≥6 years, 11 of 23 patients (47·8%) had a subnormal GH peak 3 months after TBI that persisted in 8 of 23 patients (34%) after 1 year. The GH response showed no correlation with injury severity (GCS, Marshall classification). Growth velocity was normal in all patients, except for one. Body mass index (BMI) SDS increased significantly in the group with low GH response. A suboptimal cortisol was observed in 10 of 23 subjects, which normalized in all but three, 1 year thereafter. All patients but one showed a pubertal response to GnRH testing. No clinical or hormonal abnormalities were detectable in children <6 years. CONCLUSION: Our results recommend to prospectively follow children after TBI: firstly, because the impairment of pituitary function cannot be predicted, and secondly, to avoid the potential consequences of pituitary dysfunction. Prospective clinical trials are needed before recommending a systematic screening after TBI and/or GH therapy either in postpubertal children or in prepubertal children who grow normally.