Influencia del número de copias del gen SMN2 y de la deleción del gen NAIP en el fenotipo de la Atrofia Muscular Espinal

摘要

Spinal Muscular Atrophy (SMA) is one of the most common autosomal recessive illnesses, caused by mutations of the SMN1 gene. SMA is characterized by symmetric proximal muscle weakness secondary to degeneration of the anterior horn cells of the spinal cord. SMA is clinically subdivided into four types, ranging from the severe neonatal presentation to the adult form with minimum weakness. Phenotypic variability has been associated with the number of copies of SMN2, a gene that is highly homologous and close to SMN1. Unlike SMN1, the SMN2 gene produces only 10%-15% of functional protein. Currently several clinic research studies are being developed with drugs that stimulate increased synthesis of functional SMN protein from SMN2, including valproic acid (VPA) and 4-phenylbutyrate. Another gene close to SMN1, called NAIP, is deleted in most patients with severe SMA.ObjectivesThe main objective of this work was to evaluate the influence of the SMN2 copy number and absence of NAIP gene in SMA phenotype in children diagnosed at the Pediatric Hospital J.P Garrahan. For this purpose:- SMN2 copy number was determined through two semiquantitative molecular biology methodologies.- Presence or absence of NAIP gene was established.- A genotype-phenotype correlation study was conducted in SMA children of our population.Patients and MethodologyWe studied 144 children diagnosed with SMA; these were categorized according to the criteria established by the ?Consensus Statement for Standard of Care in Spinal Muscular Atrophy, 2007?. The number of copies of the SMN2 gene was determined by MLPA (Multiplex Ligation-dependet Probe Amplification) and Real Time PCR. The absence of NAIP gene was established by MLPA.ResultsThe SMN2 copy number could be determined in all patients by MLPA. With the real time PCR it could be established in 119/144 patients. The results obtained by both techniques were equivalent in 95% of cases.All children with SMA type I had 2 copies of SMN2, while over 98% of patients with SMA type II and II showed between 3 and 4 SMN2 copies. Only in the group of patients with SMA type III were observed 4 copies of SMN2, being these more frequent in patients with the subtype IIIb.The NAIP gene was absent in 73,2% of children with SMA type I and present in more than 85% of patients with SMA type II and III.ConclusionsThe MLPA methodology proved to be more robust than real-time PCR and provides more information about SMA region.The number of copies of SMN2 and the absence of NAIP gene showed to be phenotypic modifiers of SMA in the study population. These biomarkers do not explain all cases, suggesting the existence of other modifying factors that are still unknown and should be disclosed in future research.This project contributed to the incorporation of a new methodology that increases diagnostic sensitivity (exceeding 95%) and allows detection of carriers in the family group.