title: Preventing formation of toxic N-terminal huntingtin fragments through antisense oligonucleotide-mediated protein modification.
abstract: Huntington's disease (HD) is a progressive autosomal dominant disorder, caused by a CAG repeat expansion in the HTT gene, which results in expansion of a polyglutamine stretch at the N-terminal end of the huntingtin protein. Several studies have implicated the importance of proteolytic cleavage of mutant huntingtin in HD pathogenesis and it is generally accepted that N-terminal huntingtin fragments are more toxic than full-length protein. Important cleavage sites are encoded by exon 12 of HTT. Here we report proof of concept using antisense oligonucleotides to induce skipping of exon 12 in huntingtin pre-mRNA, thereby preventing the formation of a 586 amino acid N-terminal huntingtin fragment implicated in HD toxicity. In vitro studies showed successful exon skipping and appearance of a shorter huntingtin protein. Cleavage assays showed reduced 586 amino acid N-terminal huntingtin fragments in the treated samples. In vivo studies revealed exon skipping after a single injection of antisense oligonucleotides in the mouse striatum. Recent advances to inhibit the formation of mutant huntingtin using oligonucleotides seem promising therapeutic strategies for HD. Nevertheless, huntingtin is an essential protein and total removal has been shown to result in progressive neurodegeneration in vivo. Our proof of concept shows a completely novel approach to reduce mutant huntingtin toxicity not by reducing its expressing levels, but by modifying the huntingtin protein.
authors.affiliation: 1 Department of Human Genetics, Leiden University Medical Center , The Netherlands .
authors.forename: Melvin M
authors.initials: MM
authors.lastname: Evers
authors.forename: Hoang-Dai
authors.initials: HD
authors.lastname: Tran
authors.forename: Ioannis
authors.initials: I
authors.lastname: Zalachoras
authors.forename: Onno C
authors.initials: OC
authors.lastname: Meijer
authors.forename: Johan T
authors.initials: JT
authors.lastname: den Dunnen
authors.forename: Gert-Jan B
authors.initials: GJ
authors.lastname: van Ommen
authors.forename: Annemieke
authors.initials: A
authors.lastname: Aartsma-Rus
authors.forename: Willeke M C
authors.initials: WM
authors.lastname: van Roon-Mom
mesh_terms: D000818:Animals; D053178:Caspase 6 / Q000378:metabolism; D002460:Cell Line; D005091:Exons; D006801:Humans; D000071058:Huntingtin Protein; D006816:Huntington Disease / Q000235:genetics* / Q000378:metabolism / Q000628:therapy*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000235:genetics* / Q000378:metabolism*; D016376:Oligonucleotides, Antisense / Q000235:genetics* / Q000627:therapeutic use*; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D046188:Protein Modification, Translational; D050486:Serotonin Plasma Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism; D052416:Targeted Gene Repair / Q000379:methods*; D019680:Trinucleotide Repeat Expansion
chemical_list: C086055:HTT protein, human; D000071058:Huntingtin Protein; D050505:Mutant Proteins; D009419:Nerve Tissue Proteins; D016376:Oligonucleotides, Antisense; D010446:Peptide Fragments; D050486:Serotonin Plasma Membrane Transport Proteins; C493231:Slc6a4 protein, mouse; C505514:CASP6 protein, human; D053178:Caspase 6