1. NGS panel - Genetic testing for nemaline myopathy

Nemaline myopathy

April 28, 2017

Clinical features

Nemaline myopathy (NEM, NM) is one of the most common congenital myopathies characterized by hypotonia, weakness, and absent deep tendon reflexes. Muscle weakness is usually most severe in the face, the neck flexors, and the proximal limb muscles.

Nemaline myopathies (NEM) include at least six forms of myopathies, which are classified by severity, age of onset, and presence or absence of respiratory involvement1:

  • Severe congenital myopathy accounts for ~16% of all individuals with NEM and it has neonatal onset and severe clinical features with poor prognosis1
  • Amish NEM (less than 1% of all NEM cases) that affects only patients of Amish ancestry1, 10;
  • Intermediate congenital NEM (20%) has neonatal onset but the clinical features are less severe and slowly progressive1;
  • Typical congenital NEM that is detected in ~46% of all NEM cases1;
  • Childhood-onset NEM observed in about 13% of all NEM cases;
  • Adult-onset NEM observed in 4% of all NEM cases1.

Nemaline myopathy should be suspected in individuals with the following clinical signs and symptoms1:

  • Predominantly proximal muscle weakness, often with facial muscles involvement
  • Hypotonia and decreased deep tendon reflexes, with unchanged cognition and sensation
  • Feeding and respiratory difficulties, resulting from facial and bulbar weakness
  • Recurrent respiratory infections and restrictive lung disease resulting from respiratory muscles weakness
  • Early onset: neonatal or infantile and childhood onset
    Earlier onset is associated with proximally predominant or generalized muscle weakness
  • A family history of myopathy, consistent with autosomal recessive or autosomal dominant inheritance.

Generally, NEM represents about 17% of all congenital myopathies1, 15. NEM is a rare disorder with an estimated incidence of 1:50,000 live births1, 9.

NEM may be more common in some populations; an incidence in the Amish community of 1:500 has been suggested1, 10, 11. NEM has a clearly documented genetic background; to date seven genes involved in muscle thin filament structure and function, have been linked to NEM: ACTA1, CFL2, MTM1, NEB, TNNT1, TPM2, TPM3 (Table 1).

Overview of the genes in CENTOGENE´s Nemaline myopathy panel

chr. Locus
Protein % of mutations Associated and allelic disorders
ACTA1 102610
Actin a1 sceletal muscle protein 15%-25% 1, 2, 3 >50% of severe lethal congenital NM 1, 4 Nemaline myopathy 3; Scapulohumeroperoneal myopathy; Actin congenital myopathy; Congenital myopathy with fiber-type disproportion 1
CFL2 601443
Cofilin 2 3 families 1, 5, 6 Nemaline myopathy 7 autosomal recessive
MTM1 300415
Myotubularin Rare, 5/504 7 Myotubular myopathy X-linked, Centonuclear myopathy X-linked
NEB 161650
Nebulin Up to 50% 1, 8
Hot spot 2502-bp del (exon 55) in Ashkenazi Jewish 9
Nemaline myopathy 2, autosomal recessive
TNNT1 191041
Troponin T1 Only in Old Amish 1, 10 common Amish nonsense mutation p.Glu180Ter 11 Nemaline myopathy 5 Amish type; Amish Nemaline Myopath
TPM2 190990
Tropomyosin 2 2/54 families (<1%) 1, 12 Nemaline myopathy 4 autosomal dominant; CAP myopathy 2; Arthrogryposis multiplex congenita, distal type 1; Arthrogryposis distal type 2B
TPM3 191030
Tropomyosin 3 3/117 (2%-3%) 1, 13, 14 Nemaline myopathy 1; CAP myopathy 1; Congenital myopathy with fiber-type disproportion

The rationale for CENTOGENE´s selection of 7 NEM-associated genes in CENTOGENE´s Nemaline myopathy panel (Table 1) is as follows:

  • Mutations in ACTA1 cause 20-25% of all nemaline myopathy, but 50% of severe nemaline myopathy1, 2, 4
  • Mutations in CFL2 is a rare cause of nemaline myopathy, having been described in only three families to date5, 6
  • Mutations in the MTM1 gene, encoding the myotubularin protein, required for muscle cell differentiation, cause X-linked myotubular myopathy, also known as centronuclear myopathy7 This form of NEM is mostly slowly progressive myopathy affecting first proximal muscles and so far it has been described in only few families7
  • Mutations of NEB accounts for about 50% of NEM with the usually typical congenital phenotype1, 8. Less commonly, NEB pathogenic variants are associated with a severe neonatal presentation.
  • Mutations of TNNT1 have been described in the Amish population, with variable age of onset and clinical phenotypes. The disease has an incidence of 1:500 in Amish population1, 10. Amish NEM is usually autosomal recessive with neonatal onset and poor prognosis e.g. early lethality.
  • Mutations of TPM2 are mostly considered for mild dominant disease and have only been identified in afew families worldwide12
  • Mutations of cause NEM type 1, mostly early onset NEM with either autosomal recessive or dominant inheritance13. Most patients have respiratory failure and also an additional pathohistological hallmark of the NEM disease, e.g. nemaline rod inclusions15.

There is no cure for nemaline myopathy but a number of procedures and management actions could significantly improve the quality of a patient´s life. A multidisciplinary approach is needed to handle respiratory infections and insufficiency16. Furthermore, scoliosis, joint contractures or speech anomalies must be managed as usual, as well as cardiac dysfunctions. Prognosis depends on the NM type, and life expectancy ranges from few months to a near-normal lifespan1, 16. Specific therapies are currently being developed and some of them are in the clinical trial stage16.

CENTOGENE offers the Nemaline myopathy panel (genes: ACTA1, CFL2, MTM1, NEB, TNNT1, TPM2, TPM3) including full gene sequencing and deletion/duplication analysis of the MTM1 gene. In addition, any of the genes in the Nemaline myopathy panel can also be ordered individually, for full gene sequencing and deletion/duplication analysis.

Differential diagnosis

The differential diagnosis of nemaline myopathy-related-disorders – depending on the major symptoms in the initial case – includes the following diseases1:

  • Congenital myasthenic syndromes
  • Mitochondrial myopathies
  • Prader-Willi syndrome
  • Metabolic myopathies (Pompe disease)
  • Spinal muscular atrophy
  • Secondary nemaline myopathies, including mitochondrial myopathy, dermatomyositis, myotonic dystrophy type 1, and Hodgkin’s disease, and in normal human extraocular muscle.

Testing strategy

CENTOGENE offers advanced, fast and cost-effective strategy to test large NGS panels and diagnose complex phenotypes based on the PCR-free Whole Genome Sequencing and NGS technology. This approach offers an unparalleled advantage by reducing amplification/capture biases and provides sequencing of entire gene at a more uniform coverage.

To confirm/establish the diagnosis, CENTOGENE offers the following testing strategy for nemaline myopathy using NGS Panel Genomic targeted towards this specific phenotype:

Step 1: Whole genome sequencing from a single filter card. The sequencing covers the entire genic region (coding region, exon/intron boundaries, intronic and promoter) for all the genes included in the Nemaline myopathy panel. Copy Number Variants analysis derived from NGS data is also included.

Step 2: If no mutation is identified after analysis of the Nemaline myopathy panel, based on the approval and consent, we further recommend to continue the bioinformatics analysis of the data obtained by whole genome sequencing to cover genes that are either implicated in an overlapping phenotype or could be involved in a similar pathway but not strongly clinically implicated based on the current information in literature.

Referral reasons

The following individuals are candidates for nemaline myopathy panel genetic testing:

  • Individuals with a family history of nemaline myopathy (positive family history) and presentation of the weakness, hypotonia, and depressed or absent deep tendon reflexes
  • Individuals with a negative but suspected family history, in order to perform proper genetic counseling (prenatal analyses are recommended in families with nemaline myopathy-affected individuals).

Test utility

Sequencing, deletion/duplication of the Nemaline myopathy panel genes (ACTA1, CFL2, MTM1, NEB, TNNT1, TPM2, and TPM3) should be performed in all individuals suspected for nemaline myopathy. In parallel, other genes reported to be related with congenital myopathies should also be analyzed for the presence of mutations, due to the overlap in many clinical features caused by those particular genes (e.g. muscle weakness, hypotonia, absent or depressed deep tendon reflex etc.).

Confirmation of a clinical diagnosis through genetic testing can allow for genetic counseling and may direct medical management. Furthermore, genetic counseling can provide a patient and/or family with the natural history of the nemaline myopathy and it can also identify at-risk family members, provide reproductive risks as well as preconception/prenatal options, and allow for appropriate referral for patient support and/or resources.

More information on CENTOGENE´s Nemaline myopathy panel can be found in our genetic test catalogue.