Clinical application of whole exome sequencing reveals a novel compound heterozygous TK2-mutation in two brothers with rapidly progressive combined muscle-brain atrophy, axonal neuropathy, and status epilepticus

Highlights

• Two boys with encephalomyopathy were examined by whole exome sequencing.

• Identity-by-state analysis reduced disease loci in a non-consanguineous family.

• A novel mutation in the mitochondrial Thymidine kinase 2 (TK2) gene was found.

• Both patients broaden the clinical phenotype associated with TK2 mutations.

• TK2 mutations can cause severe encephalopathy, axonal neuropathy and epilepsy.

Abstract

Mutations in several genes cause mtDNA depletion associated with encephalomyopathy. Due to phenotypic overlap, it is difficult to conclude from clinical phenotype to genetic defect. Here we report on two brothers who presented with rapid fatty muscle degeneration, axonal neuropathy, rapid loss of supratentorial white and gray matter, and status epilepticus. Whole exome sequencing coupled with ‘identity-by-state’ (IBS) analysis revealed a compound heterozygous missense mutation (p.M117V, p.A139V) in the thymidine kinase 2 (TK2) gene that segregated with the phenotype. Both mutations were located in the thymidine binding pouch of the enzyme. Residual mtDNA copy numbers in muscle were 8.5%, but normal in blood and fibroblasts. Our results broaden the clinical phenotype spectrum of TK2 mutations and promote WES as a useful method in the clinical setting for mutation detection, even in untypical cases. If two or more affected siblings from a non-consanguineous family can be investigated, IBS-analysis provides a powerful tool to narrow the number of disease candidates, similarly to autozygosity mapping in consanguineous families.

Introduction

A group of infantile or early-onset disorders, collectively termed mitochondrial DNA (mtDNA) depletion syndromes, is characterized by a massive reduction of mitochondrial DNA content. This is caused by a dysfunction of genes associated with mtDNA-synthesis and maintenance. Many of these patients suffer from a hepato-cerebral syndrome (Al-Hussaini et al., 2014). Encephalopathy concurs with myopathy in another subgroup of patients (Spinazzola et al., 2009), who carry mutations in SUCLA2 (ATP-dependent succinyl-CoA synthase), SUCLG1 (GTP-dependent succinyl-CoA synthase), RRM2B (p53-induced ribonucleotide reductase B subunit) and TK2 (thymidine kinase 2). The mitochondrial thymidine kinase 2 mediates the first step in the phosphorylation of deoxypyrimidine nucleosides required for mtDNA replication in the mitochondrial matrix. Disturbance of these phosphorylation steps results in overall mtDNA depletion (Zhou et al., 2008). The phenotypic spectrum of the TK2-related mtDNA-depletion syndrome (OMIM #609560) is broad and as many as 26 different missense and 9 InDel mutations (HGMD® public database, http://www.hgmd.cf.ac.uk) in 50 patients have been reported so far (Chanprasert et al., 2013). Patients typically present with a progressive muscle disease characterized by generalized hypotonia, proximal muscle weakness, loss of motor skills, feeding and respiratory difficulties (Saada et al., 2001). Respiratory failure due to weakness of the respiratory muscles leads to death in early childhood. TK2 mutations were initially thought to cause a purely myopathic form of mtDNA-depletion syndrome. However, subsequent reports have expanded the clinical spectrum including chronic external ophthalmoplegia, sensorineural hearing loss and early severe muscle weakness with encephalopathy and intractable epilepsy in single cases (Oskoui et al., 2006). Beyond that, milder forms with late- or adult-onset proximal muscle weakness and prolonged survival have been reported (Béhin et al., 2012, Chanprasert et al., 2013). The phenotypic range might be explained by the variability in the amount of residual activity of the mutant enzyme (Poulton et al., 2009).

Achieving an accurate diagnosis can be difficult and requires the coordinated interplay of clinical assessment, muscle histology, respiratory chain enzymology as well as molecular genetic analysis. Whole-exome sequencing (WES) has been used successfully in research to identify novel genes for many Mendelian disorders. Although still improving, WES has already arrived in the clinic and has become a powerful tool in the diagnosis of Mendelian disorders. Success of this approach is more frequent in consanguineous families since the search can be restricted to homozygous variants, but it is also possible for compound heterozygotes, especially if more than one affected patient can be investigated in a single family.

Here we report on two brothers with a fatal clinical course of early onset myopathy with profound loss of muscle mass, axonal neuropathy, respiratory failure as well as severe brain atrophy with status epilepticus. Although disease manifestation and course were dramatic, the diagnosis had been missed for several years because of the deviating phenotype. Finally, WES revealed the diagnosis of mitochondrial depletion syndrome due to a novel TK2 mutation.