title: Antisense oligonucleotide treatment for a pseudoexon-generating mutation in the NPC1 gene causing Niemann-Pick type C disease.
abstract: Niemann-Pick type C disease is an autosomal recessive disorder caused by mutations in either the NPC1 or NPC2 gene. While most of the mutations are missense, a few splicing mutations have also been described. We identified and characterized a novel point mutation c.1554-1009G>A located in intron 9 of the NPC1 gene in a Spanish patient. Sequencing of the cDNA from the patient showed that this intronic mutation creates a cryptic donor splice site resulting in the incorporation of 194 bp of intron 9 as a new exon (pseudoexon) in the mRNA. This new transcript bears a premature termination codon and is degraded by the nonsense-mediated mRNA decay mechanism. Experimental confirmation that the point mutation generates the inclusion of a pseudoexon in the mRNA was obtained using a minigene. A specific antisense morpholino oligonucleotide targeted to the cryptic splice site was designed and transfected into fibroblasts from the patient. Using this approach, normal splicing was restored. These results demonstrate the importance of screening deep intronic regions and support the efficacy of antisense therapeutics for the treatment of diseases caused by pseudoexon-generating mutations.
authors.affiliation: Departament de Genètica, Facultat de Biologia, Universitat de Barcelona, Barcelona, Spain.
authors.forename: Laura
authors.initials: L
authors.lastname: Rodríguez-Pascau
authors.forename: Maria Josep
authors.initials: MJ
authors.lastname: Coll
authors.forename: Lluïsa
authors.initials: L
authors.lastname: Vilageliu
authors.forename: Daniel
authors.initials: D
authors.lastname: Grinberg
mesh_terms: D017398:Alternative Splicing; D002352:Carrier Proteins / Q000235:genetics*; D015316:Genetic Therapy; D006367:HeLa Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008297:Male; D008562:Membrane Glycoproteins / Q000235:genetics*; D000077296:Niemann-Pick C1 Protein; D052556:Niemann-Pick Disease, Type C / Q000235:genetics* / Q000628:therapy*; D016376:Oligonucleotides, Antisense / Q000627:therapeutic use*; D017354:Point Mutation
chemical_list: D002352:Carrier Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008562:Membrane Glycoproteins; C106881:NPC1 protein, human; D000077296:Niemann-Pick C1 Protein; D016376:Oligonucleotides, Antisense