A novel strategy for the identification of pathogenic intronic/complex genomic variants – a cohort study of 20 dystrophinopathy cases.


RNA-seq analysis, targeted long-read sequencing and in silico prediction to unravel pathogenic intronic events and complicated splicing abnormalities in dystrophinopathy

Human Genet 2022 Sep 1

First author

Mariko Okuba


Michaela Yuen

Dystrophinopathies, caused by genetic variants in Dystrophin (DMD), are the most common hereditary neuromuscular condition. Currently, genetic diagnosis of Dystrophinopathies relies on sequencing exonic regions and regions flanking exons of DMD, however, in approximately 1% of patients a pathogenic genetic variant cannot be identified despite reduced DMD protein levels. With effective therapies for Dystrophinopathies becoming increasingly available an accurate genetic diagnosis is critical to determine whether a patient will benefit from an existing therapy or be eligible to participate in clinical trials.

The authors developed a diagnostic pipeline for the identification of intronic pathogenic variants affecting DMD transcription in 20 patients which failed to receive a diagnosis using traditional methods. Genetic investigations included RNA sequencing (to identify abnormal DMD transcripts), targeted long-read DMD sequencing (able to identify complex, large intra-genetic structural rearrangements) and in silico splicing analysis (Splice AI and MaxEntScan; to identify the genetic variant underlying abnormal splicing). The authors identified single nucleotide variants associated with exonization of intronic sequence and exon skipping as well as complex structural rearrangements and repeat expansions affecting DMD transcripts. A genotype-phenotype correlation was highlighted with more residual DMD transcripts associated with a milder phenotype.

The authors conclude that their approach to Dystrophinopathy diagnosis was capable of uncovering pathogenic genomic variations in the 20 cases studied. They also highlight that pathogenic exonization events in DMD are excellent candidates for treatment with personalised exon-skipping therapies.

About the author

Dr. Mariko Okubo is a pediatric neurologist striving to perform high quality research for the benefit of patients affected by debilitating muscle disease. Dr. Mariko Okubo recently completed her postdoctoral position at the Department of Neuromuscular Research, National Center of Neurology and Psychiatry (NCNP) in Japan, and is currently working as a postdoctoral researcher at the Institut de Myologie in France. Her research focus is genetics, muscle pathology and gene therapy. She previously published several papers about Duchenne/Becker Muscular dystrophy focusing on methodology of genetic analysis, mutation spectrum in Japan, and Dystrophin nonsense mutations. 


This article is presented by the

Publication Highlights Committee.

Published on 22 September 2022.


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