Comprehensive genetic diagnosis of patients with Duchenne/Becker muscular dystrophy (DMD/BMD) and pathogenicity analysis of splice site variants in the DMD gene

Abstract

Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) are caused by mutations in the DMD gene. The aim of this study is to identify pathogenic DMD variants in probands and reduce the risk of recurrence of the disease in affected families. Variations in 100 unrelated DMD/BMD patients were detected by multiplex ligation-dependent probe amplification (MLPA) and next-generation sequencing (NGS). Pathogenic variants in DMD were successfully identified in all cases, and 11 of them were novel. The most common mutations were intragenic deletions (69%), with two hotspots located in the 5' end (exons 2-19) and the central of the DMD gene (exons 45-55), while point mutations were observed in 22% patients. Further, c.1149+1G>A and c.1150−2A>G were confirmed by hybrid minigene splicing assay (HMSA). This two splice site mutations would lead to two aberrant DMD isoforms which give rise to severely truncated protein. Therefore, the clinical use of MLPA, NGS, and HMSA is an effective strategy to identify variants. Importantly, eight embryos were terminated pregnancies according to prenatal diagnosis and a healthy boy was successfully delivered by preimplantation genetic diagnosis (PGD). Early and accurate genetic diagnosis is essential for prenatal diagnosis/PGD to reduce the risk of recurrence of DMD in affected families.

Whole Exome Sequencing Aids the Diagnosis of Fetal Skeletal Dysplasia

“…To our knowledge, only one fetus with a deletion of exons 17–29 of the DMD gene was reported to present prenatal phenotypes including fetal growth restriction and oligohydramnios (Lin et al, 2017 ). And the deletion of exons 49–53 of the DMD gene were detected in three patients with DMD/BMD as reported previously (Covone et al, 1991 ; Murugan et al, 2010 ; Yang et al, 2019 ). However, our patient was dead, and we could not get more information.…”

The multifaceted view of heart problem in Duchenne muscular dystrophy

“…Genetic analysis verifies the presence of DMD mutations. Currently, the multiplex ligation-dependent probe amplification (MLPA), is the most commonly used first-line screening method, able to identify exonic deletions and duplications of DMD both in male patients and female carriers, conferring an advantage over multiplex polymerase chain reaction (PCR) [49]. Cases with negative MLPA results and/ or unrecognized mutation require further evaluation by DMD sequencing.…”

“…Cases with negative MLPA results and/ or unrecognized mutation require further evaluation by DMD sequencing. Next-generation sequencing (NGS) is increasingly used for detection of large deletions/duplications, point mutations (nonsense, missense, splice site mutation) and small insertions/deletions (indels) [49].…”

Duchenne muscular dystrophy: problems of differential diagnosis and treatment

“…DMD є одним із найбільших генів людини (приблизно 2,6 млн пар ДНК) та містить 79 екзонів. Ген кодує кілька ізоформних білків, найважливіший з яких м'язовий дистрофін [26]. Дистрофін відповідає за з'єднання цито скелету м'язового волокна з основою базальної пластинки (позаклітинного матриксу) через дистрофін асоційований глікопротеїдний ком плекс у сарколемі (dystrophin associated protein complex -DAPC).…”