Unraveling the effect of silent, intronic and missense mutations on VWF splicing: contribution of next generation sequencing in the study of mRNA

Large studies in von Willebrand disease patients, including Spanish and Portuguese registries, led to identification of >250 different mutations. It is a challenge to determine the pathogenic effect of potential splice site mutations on VWF mRNA. This study aimed to elucidate the true effects of 18 mutations on VWF mRNA processing, investigate the contribution of next-generation sequencing to in vivo mRNA study in von Willebrand disease, and compare the findings with in silico prediction. RNA extracted from patient platelets and leukocytes was amplified by RT-PCR and sequenced using Sanger and next generation sequencing techniques. Eight mutations affected VWF splicing: c.1533+1G>A, c.5664+2T>C and c.546G>A (p.=) prompted exon skipping; c.3223-7_3236dup and c.7082-2A>G resulted in activation of cryptic sites; c.3379+1G>A and c.7473G>A (p.=) demonstrated both molecular pathogenic mechanisms simultaneously; and the p.Cys370Tyr missense mutation generated two aberrant transcripts. Of note, the complete effect of 3 mutations was provided by next generation sequencing alone because of low expression of the aberrant transcripts. In the remaining 10 mutations, no effect was elucidated in the experiments. However, the differential findings obtained in platelets and leukocytes provided substantial evidence that 4 of these would have an effect on VWF levels. In this first report using next generation sequencing technology to unravel the effects of VWF mutations on splicing, the technique yielded valuable information. Our data bring to light the importance of studying the effect of synonymous and missense mutations on VWF splicing to improve the current knowledge of the molecular mechanisms behind von Willebrand disease.info:eu-repo/semantics/publishedVersio

Unraveling the effect of silent, intronic and missense mutations on VWF splicing: contribution of next generation sequencing in the study of mRNA

Large studies in von Willebrand disease patients, including Spanish and Portuguese registries, led to the identification of >250 different mutations. It is a challenge to determine the pathogenic effect of potential splice site mutations on VWF mRNA. This study aimed to elucidate the true effects of 18 mutations on VWF mRNA processing, investigate the contribution of next-generation sequencing to in vivo mRNA study in von Willebrand disease, and compare the findings with in silico prediction. RNA extracted from patient platelets and leukocytes was amplified by RT-PCR and sequenced using Sanger and next generation sequencing techniques. Eight mutations affected VWF splicing: c.1533+1G>A, c.5664+2T>C and c.546G>A (p.=) prompted exon skipping; c.3223-7_3236dup and c.7082-2A>G resulted in activation of cryptic sites; c.3379+1G>A and c.7437G>A) demonstrated both molecular pathogenic mechanisms simultaneously; and the p.Cys370Tyr missense mutation generated two aberrant transcripts. Of note, the complete effect of three mutations was provided by next generation sequencing alone because of low expression of the aberrant transcripts. In the remaining 10 mutations, no effect was elucidated in the experiments. However, the differential findings obtained in platelets and leukocytes provided substantial evidence that four of these would have an effect on VWF levels. In this first report using next generation sequencing technology to unravel the effects of VWF mutations on splicing, the technique yielded valuable information. Our data bring to light the importance of studying the effect of synonymous and missense mutations on VWF splicing to improve the current knowledge of the molecular mechanisms behind von Willebrand disease. clinicaltrials.gov identifier:02869074

Psicosis epiléptica periictal, una causa de psicosis reversible

Resumen: Introducción: Las psicosis epilépticas se dividen respecto de su relación con las crisis en periictales e interictales. Las psicosis periictales tienen una estrecha relación temporal con las crisis epilépticas y ocurren antes (preictales), durante (ictales) o después de las mismas (postictales). Generalmente, tienen un inicio y final agudo, corta duración y una remisión completa, con riesgo de recurrencia. Las psicosis interictales o crónicas no guardan relación temporal con las crisis epilépticas. Existe otro tipo de psicosis epilépticas que se relaciona con la respuesta al tratamiento de la epilepsia: psicosis epiléptica por fenómeno de normalización forzada (psicosis alternativa) y dentro de esta se encuentra la psicosis epiléptica secundaria a cirugía de la epilepsia. Aunque se ha generalizado la combinación de antiepilépticos y neurolépticos para su manejo, no existen unas pautas estandarizadas de tratamiento en las psicosis epilépticas. Casos clínicos: Presentamos 5 casos de psicosis epilépticas periictales y remarcamos la excelente respuesta al tratamiento con levetiracetam. Consiguiendo un buen control tanto de las crisis como de los episodios psicóticos. Este fármaco resultó inocuo al asociarlo con neurolépticos en nuestros pacientes y no se precisaron dosis elevadas de estos últimos. Conclusiones: La diferenciación de los estados psicóticos asociados con la epilepsia según la relación temporal con las crisis epilépticas tiene utilidad clínica y pronóstica, dado que aporta aspectos importantes respecto al tratamiento y a la evolución de la enfermedad. El tratamiento de los trastornos mentales periictales o agudos se basa en el control de las crisis epilépticas, mientras que el tratamiento de los interictales o crónicos guarda más similitud con el de los trastornos de origen puramente psiquiátrico. El control estricto de las crisis puede, además de mejorar la calidad de vida del paciente y su discapacidad, prevenir el desarrollo de una psicosis interictal, por lo que consideramos que sería necesario establecer un protocolo de tratamiento para estos casos. Abstract: Introduction: Epileptic psychoses are categorised as peri-ictal and interictal according to their relationship with the occurrence of seizures. There is a close temporal relationship between peri-ictal psychosis and seizures, and psychosis may present before (preictal), during (ictal) or after seizures (postictal). Epileptic psychoses usually have acute initial and final phases, with a short symptom duration and complete remission with a risk of recurrence. There is no temporal relationship between interictal or chronic psychosis and epileptic seizures. Another type of epileptic psychosis is related to the response to epilepsy treatment: epileptic psychosis caused by the phenomenon of forced normalisation (alternative psychosis), which includes epileptic psychosis secondary to epilepsy surgery. Although combination treatment with antiepileptic and neuroleptic drugs is now widely used to manage this condition, there are no standard treatment guidelines for epileptic psychosis. Clinical cases: We present 5 cases of peri-ictal epileptic psychosis in which we observed an excellent response to treatment with levetiracetam. Good control was achieved over both seizures and psychotic episodes. Levetiracetam was used in association with neuroleptic drugs with no adverse effects, and our patients did not require high doses of the latter. Conclusions: Categorising psychotic states associated with epilepsy according to their temporal relationship with seizures is clinically and prognostically useful because it provides important information regarding disease treatment and progression. The treatment of peri-ictal or acute mental disorders is based on epileptic seizure control, while the treatment of interictal or chronic disorders has more in common with managing disorders which are purely psychiatric in origin. In addition to improving the patient's quality of life and reducing disability, achieving strict control over seizures may also prevent the development of interictal psychosis. For this reason, we believe that establishing a treatment protocol for such cases is necessary. Palabras clave: Epilepsia, Crisis, Levetiracetam, Periictal, Psicosis, Psicosis epiléptica, Keywords: Epilepsy, Seizure, Levetiracetam, Peri-ictal, Psychosis, Epileptic psychosi

Epileptic peri-ictal psychosis, a reversible cause of psychosis

Introduction: Epileptic psychoses are categorised as peri-ictal and interictal according to their relationship with the occurrence of seizures. There is a close temporal relationship between peri-ictal psychosis and seizures, and psychosis may present before (preictal), during (ictal) or after seizures (postictal). Epileptic psychoses usually have acute initial and final phases, with a short symptom duration and complete remission with a risk of recurrence. There is no temporal relationship between interictal or chronic psychosis and epileptic seizures. Another type of epileptic psychosis is related to the response to epilepsy treatment: epileptic psychosis caused by the phenomenon of forced normalisation (alternative psychosis), which includes epileptic psychosis secondary to epilepsy surgery. Although combination treatment with antiepileptic and neuroleptic drugs is now widely used to manage this condition, there are no standard treatment guidelines for epileptic psychosis. Clinical cases: We present 5 cases of peri-ictal epileptic psychosis in which we observed an excellent response to treatment with levetiracetam. Good control was achieved over both seizures and psychotic episodes. Levetiracetam was used in association with neuroleptic drugs with no adverse effects, and our patients did not require high doses of the latter. Conclusions: Categorising psychotic states associated with epilepsy according to their temporal relationship with seizures is clinically and prognostically useful because it provides important information regarding disease treatment and progression. The treatment of peri-ictal or acute mental disorders is based on epileptic seizure control, while the treatment of interictal or chronic disorders has more in common with managing disorders which are purely psychiatric in origin. In addition to improving the patient's quality of life and reducing disability, achieving strict control over seizures may also prevent the development of interictal psychosis. For this reason, we believe that establishing a treatment protocol for such cases is necessary. Resumen: Introducción: Las psicosis epilépticas se dividen respecto de su relación con las crisis en periictales e interictales. Las psicosis periictales tienen una estrecha relación temporal con las crisis epilépticas y ocurren antes (preictales), durante (ictales) o después de las mismas (postictales). Generalmente, tienen un inicio y final agudo, corta duración y una remisión completa, con riesgo de recurrencia. Las psicosis interictales o crónicas no guardan relación temporal con las crisis epilépticas. Existe otro tipo de psicosis epilépticas que se relaciona con la respuesta al tratamiento de la epilepsia: psicosis epiléptica por fenómeno de normalización forzada (psicosis alternativa) y dentro de esta se encuentra la psicosis epiléptica secundaria a cirugía de la epilepsia. Aunque se ha generalizado la combinación de antiepilépticos y neurolépticos para su manejo, no existen unas pautas estandarizadas de tratamiento en las psicosis epilépticas. Casos clínicos: Presentamos 5 casos de psicosis epilépticas periictales y remarcamos la excelente respuesta al tratamiento con levetiracetam. Consiguiendo un buen control tanto de las crisis como de los episodios psicóticos. Este fármaco resultó inocuo al asociarlo con neurolépticos en nuestros pacientes y no se precisaron dosis elevadas de estos últimos. Conclusiones: La diferenciación de los estados psicóticos asociados con la epilepsia según la relación temporal con las crisis epilépticas tiene utilidad clínica y pronóstica, dado que aporta aspectos importantes respecto al tratamiento y a la evolución de la enfermedad. El tratamiento de los trastornos mentales periictales o agudos se basa en el control de las crisis epilépticas, mientras que el tratamiento de los interictales o crónicos guarda más similitud con el de los trastornos de origen puramente psiquiátrico. El control estricto de las crisis puede, además de mejorar la calidad de vida del paciente y su discapacidad, prevenir el desarrollo de una psicosis interictal, por lo que consideramos que sería necesario establecer un protocolo de tratamiento para estos casos. Keywords: Epilepsy, Seizure, Levetiracetam, Peri-ictal, Psychosis, Epileptic psychosis, Palabras clave: Epilepsia, Crisis, Levetiracetam, Periictal, Psicosis, Psicosis epiléptic

Mapa epidemiológico transversal de las ataxias y paraparesias espásticas hereditarias en España

Resume: Introducción: Las ataxias (AT) y paraparesias espásticas hereditarias (PEH) son síndromes neurodegenerativos raros. Nos proponemos conocer la prevalencia de las AT y PEH en España en 2019. Pacientes y métodos: Estudio transversal, multicéntrico, descriptivo y retrospectivo de los pacientes con AT y PEH, desde marzo de 2018 a diciembre de 2019 en toda España. Resultados: Se obtuvo información de 1933 pacientes procedentes de 11 Comunidades Autónomas, de 47 neurólogos o genetistas. Edad media: 53,64 años ± 20,51 desviación estándar (DE); 938 varones (48,5%), 995 mujeres (51,5%). En 920 pacientes (47,6%) no se conoce el defecto genético. Por patologías, 1.371 pacientes (70,9%) diagnosticados de AT, 562 diagnosticados de PEH (29,1%). La prevalencia estimada de AT es 5,48/100.000 habitantes, y la de PEH es 2,24 casos/100.000 habitantes. La AT dominante más frecuente es la SCA3. La AT recesiva más frecuente es la ataxia de Friedreich (FRDA). La PEH dominante más frecuente es la SPG4, y la PEH recesiva más frecuente es la SPG7. Conclusiones: La prevalencia estimada de AT y PEH en nuestra serie es de 7,73 casos/100.000 habitantes. Estas frecuencias son similares a las del resto del mundo. En el 47,6% no se ha conseguido un diagnóstico genético. A pesar de las limitaciones, este estudio puede contribuir a estimar los recursos, visibilizar estas enfermedades, detectar las mutaciones más frecuentes para hacer los screenings por comunidades, y favorecer los ensayos clínicos. Abstract: Introduction: Ataxia and hereditary spastic paraplegia are rare neurodegenerative syndromes. We aimed to determine the prevalence of these disorders in Spain in 2019. Patients and methods: We conducted a cross-sectional, multicentre, retrospective, descriptive study of patients with ataxia and hereditary spastic paraplegia in Spain between March 2018 and December 2019. Results: We gathered data from a total of 1933 patients from 11 autonomous communities, provided by 47 neurologists or geneticists. Mean (SD) age in our sample was 53.64 (20.51) years; 938 patients were men (48.5%) and 995 were women (51.5%). The genetic defect was unidentified in 920 patients (47.6%). A total of 1371 patients (70.9%) had ataxia and 562 (29.1%) had hereditary spastic paraplegia. Prevalence rates for ataxia and hereditary spastic paraplegia were estimated at 5.48 and 2.24 cases per 100 000 population, respectively. The most frequent type of dominant ataxia in our sample was SCA3, and the most frequent recessive ataxia was Friedreich ataxia. The most frequent type of dominant hereditary spastic paraplegia in our sample was SPG4, and the most frequent recessive type was SPG7. Conclusions: In our sample, the estimated prevalence of ataxia and hereditary spastic paraplegia was 7.73 cases per 100 000 population. This rate is similar to those reported for other countries. Genetic diagnosis was not available in 47.6% of cases. Despite these limitations, our study provides useful data for estimating the necessary healthcare resources for these patients, raising awareness of these diseases, determining the most frequent causal mutations for local screening programmes, and promoting the development of clinical trials

Mapa epidemiológico transversal de las ataxias y paraparesias espásticas hereditarias en España

Introduction: Ataxia and hereditary spastic paraplegia are rare neurodegenerative syndromes. We aimed to determine the prevalence of these disorders in Spain in 2019. Patients and methods: We conducted a cross-sectional, multicentre, retrospective, descriptive study of patients with ataxia and hereditary spastic paraplegia in Spain between March 2018 and December 2019. Results: We gathered data from a total of 1933 patients from 11 autonomous communities, provided by 47 neurologists or geneticists. Mean (SD) age in our sample was 53.64 (20.51) years; 938 patients were men (48.5%) and 995 were women (51.5%). The genetic defect was unidentified in 920 patients (47.6%). A total of 1371 patients (70.9%) had ataxia and 562 (29.1%) had hereditary spastic paraplegia. Prevalence rates for ataxia and hereditary spastic paraplegia were estimated at 5.48 and 2.24 cases per 100 000 population, respectively. The most frequent type of dominant ataxia in our sample was SCA3, and the most frequent recessive ataxia was Friedreich ataxia. The most frequent type of dominant hereditary spastic paraplegia in our sample was SPG4, and the most frequent recessive type was SPG7. Conclusions: In our sample, the estimated prevalence of ataxia and hereditary spastic paraplegia was 7.73 cases per 100 000 population. This rate is similar to those reported for other countries. Genetic diagnosis was not available in 47.6% of cases. Despite these limitations, our study provides useful data for estimating the necessary healthcare resources for these patients, raising awareness of these diseases, determining the most frequent causal mutations for local screening programmes, and promoting the development of clinical trials. (c) 2021 Sociedad Espanola de Neurologia. Published by Elsevier Espana, S.L.U. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/ 4.0/)