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JCI Insight. 2019 Mar 21;4(6). pii: 124403. doi: 10.1172/jci.insight.124403. eCollection 2019 Mar 21.

A recurrent COL6A1 pseudoexon insertion causes muscular dystrophy and is effectively targeted by splice-correction therapies.

Author information

1
Neuromuscular and Neurogenetic Disorders of Childhood Section, National Institute of Neurological Disorders and Stroke, NIH, Bethesda, Maryland, USA.
2
Center for Biochemistry, Faculty of Medicine and Center for Molecular Medicine Cologne, University of Cologne, Cologne, Germany.
3
Dubowitz Neuromuscular Centre, UCL Institute of Child Health & Great Ormond Street Hospital for Children, London, United Kingdom.
4
Genetics and Genomic Medicine Programme, UCL Great Ormond Street Institute of Child Health, London, United Kingdom.
5
Analytical and Translation Genetics Unit, Massachusetts General Hospital, Boston, Massachusetts, USA.
6
Medical and Population Genetics, Broad Institute of MIT and Harvard, Cambridge, Massachusetts, USA.
7
Courant Institute of Mathematical Sciences, New York University, New York, USA.
8
Metabolic Disease Unit, Edmond and Lily Safra Children's Hospital, Sheba Medical Center, Tel-Hashomer, Israel.
9
Department of Neurology and Pediatrics, University of Utah, Salt Lake City, Utah, USA.
10
Pathology Department and Biobanc de l'Hospital Infantil Sant Joan de Déu per a la Investigació, Hospital Sant Joan de Déu, Barcelona, Spain.
11
Neuromuscular Unit, Neuropediatrics Department, Hospital Sant Joan de Déu, Institut de Recerca Sant Joan de Déu, Esplugues de Llobregat, Barcelona, Spain.
12
CIBERER (ISCIII), Madrid, Spain.
13
Peptide/Protein Sequencing Facility, National Institute of Neurological Disorder and Stroke, NIH, Bethesda, Maryland, USA.
14
Sorbonne Université, Inserm, Association Institut de Myologie, Centre de Recherche en Myologie, UMRS974, Paris, France.
15
Medical Genetics Unit, Department of Medical Science, University of Ferrara, Ferrara, Italy.
16
Bio21 Advanced Microscopy Facility, The University of Melbourne, Melbourne, Australia.
17
Centre for Molecular Medicine and Therapeutics, Murdoch University, Perth, Australia.
18
Perron Institute for Neurological and Translational Science, University of Western Australia, Perth, Australia.
19
Murdoch Children's Research Institute, Parkville, Australia.
20
Department of Paediatrics, University of Melbourne, Parkville, Australia.
21
NIHR Great Ormond Street Hospital Biomedical Research Centre, London, United Kingdom.

Abstract

The clinical application of advanced next-generation sequencing technologies is increasingly uncovering novel classes of mutations that may serve as potential targets for precision medicine therapeutics. Here, we show that a deep intronic splice defect in the COL6A1 gene, originally discovered by applying muscle RNA sequencing in patients with clinical findings of collagen VI-related dystrophy (COL6-RD), inserts an in-frame pseudoexon into COL6A1 mRNA, encodes a mutant collagen α1(VI) protein that exerts a dominant-negative effect on collagen VI matrix assembly, and provides a unique opportunity for splice-correction approaches aimed at restoring normal gene expression. Using splice-modulating antisense oligomers, we efficiently skipped the pseudoexon in patient-derived fibroblast cultures and restored a wild-type matrix. Similarly, we used CRISPR/Cas9 to precisely delete an intronic sequence containing the pseudoexon and efficiently abolish its inclusion while preserving wild-type splicing. Considering that this splice defect is emerging as one of the single most frequent mutations in COL6-RD, the design of specific and effective splice-correction therapies offers a promising path for clinical translation.

KEYWORDS:

Collagens; Extracellular matrix; Muscle Biology; Neuromuscular disease; Therapeutics

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